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Sample records for a549 human pulmonary

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

    PubMed

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

    2012-01-01

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

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

    PubMed

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

    2016-02-01

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

  3. 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.

    PubMed

    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.

  4. [Expression and significance of IKBKB in pulmonary adenocarcinoma A549 cells and its cisplatin-resistant variant A549/DDP].

    PubMed

    Qi, Kang; Li, Yang; Li, Xuebing; Zhang, Fang; Shao, Yi; Zhou, Qinghua

    2014-05-01

    Cisplatin-resistance in Lung cancer cells is widespread in the clinical treatment, seriously affecting the effects of the treatment of lung cancer. Therefore, the research of mechanisms of cisplain-resistance has significant meaning for developing new chemotherapeutic drug and solving the cisplain-resistance in clinic treatment. IKBKB is one of the most important catalytic subunits of IKK complexes. It plays an important regulatory role in activation of NF-κB. The aim of this study is to investigate the differential expression of IKBKB gene in human lung adenocarcinoma cells line A549 and the cisplatin-resistant variant A549/DDP and the mechanisms of cisplain-resistance induced by IKBKB gene. MTT assay was employed to determine the sensitivity of A549 and A549/DDP cells line to cisplatin and the effect of IKBKB gene on A549 cell lines' sensitivity to cisplatin. The mRNA level of IKBKB was determined by real-time PCR. Dual luciferase reporter gene experiment was employed to determine the activity of the NF-κB. Apoptosis rate of lung adenocarcinoma cells was determined by flow cytometry. Apoptosis rate and IC50 were significantly different in A549 and A549/DDP cells, the expression of mRNA level of IKBKB gene in A549/DDP was significantly higher than that in A549. Compared with control group, IKBKB gene was able to reduce the cisplain sensitivity of A549 cells. After A549 was transfected with pcDNA3.1/IKBKB plasmid, mRNA level of IKBKB was significantly increased, the sensitivity of cisplain was decreased, the IC50 was increased 2.85 fold, the apoptosis rate was decreased 59%, the activity of NF-κB has been greatly increased. IKBKB inhibits cisplatin-induced apoptosis via the activation of NF-κB pathway. It will be helpful in the development of new anticancer drug and solving the challenge of cisplatin-resistance.

  5. Paracrine control of differentiation in the alveolar carcinoma, A549, by human foetal lung fibroblasts.

    PubMed Central

    Speirs, V.; Ray, K. P.; Freshney, R. I.

    1991-01-01

    Synthesis of pulmonary surfactant (PS) is necessary for normal functioning of the lungs and its production is indicative of normal differentiated lung. The human alveolar carcinoma, A549, has been found to synthesis and secrete PS in vitro. The purpose of this study was to optimise the culture conditions for PS synthesis by A549 as well as to determine the potential role of foetal lung fibroblasts in the induction of PS by glucocorticoids. A549 cells growing in filter wells produced higher levels of PS in response to steroid, a 5-fold increase on the filter well compared to only a 1.5-fold increase when the cells were cultured on a conventional plastic substrate. A549 cells grown in filter wells responded to coculture with fibroblasts whether in direct contact or separated co-culture. A 20-fold increase in PS over control values was observed in separated steroid-treated co-cultures, suggesting the presence of a diffusible factor. A partially purified factor was isolated from fibroblast conditioned medium which was capable of inducing differentiation and other phenotypic changes in A549, namely induction of PS, reduction of plasminogen activator activity and reduction in the in vivo growth of A549 xenografts in nude mice. These results suggest that, under the correct conditions, A549 cells, although transformed, still retain the capacity to respond to differentiation-inducing signals from normal fibroblasts. Images Figure 5 PMID:1654985

  6. Paracrine control of differentiation in the alveolar carcinoma, A549, by human foetal lung fibroblasts.

    PubMed

    Speirs, V; Ray, K P; Freshney, R I

    1991-10-01

    Synthesis of pulmonary surfactant (PS) is necessary for normal functioning of the lungs and its production is indicative of normal differentiated lung. The human alveolar carcinoma, A549, has been found to synthesis and secrete PS in vitro. The purpose of this study was to optimise the culture conditions for PS synthesis by A549 as well as to determine the potential role of foetal lung fibroblasts in the induction of PS by glucocorticoids. A549 cells growing in filter wells produced higher levels of PS in response to steroid, a 5-fold increase on the filter well compared to only a 1.5-fold increase when the cells were cultured on a conventional plastic substrate. A549 cells grown in filter wells responded to coculture with fibroblasts whether in direct contact or separated co-culture. A 20-fold increase in PS over control values was observed in separated steroid-treated co-cultures, suggesting the presence of a diffusible factor. A partially purified factor was isolated from fibroblast conditioned medium which was capable of inducing differentiation and other phenotypic changes in A549, namely induction of PS, reduction of plasminogen activator activity and reduction in the in vivo growth of A549 xenografts in nude mice. These results suggest that, under the correct conditions, A549 cells, although transformed, still retain the capacity to respond to differentiation-inducing signals from normal fibroblasts.

  7. Interleukin-15 increases neutrophil adhesion onto human respiratory epithelial A549 cells and attracts neutrophils in vivo

    PubMed Central

    Pelletier, M; Girard, D

    2005-01-01

    Interleukin-15 (IL-15) is a neutrophil agonist that plays a role in inflammatory disorders, including a variety of pulmonary diseases. Adhesion of neutrophils onto pulmonary cells is a major event leading to development of inflammation. Recently, elevated levels of IL-15 have been associated with different pulmonary diseases. There is no clear evidence that IL-15 modulates cell surface expression of adhesion molecules in neutrophils, or that IL-15 is involved in neutrophil adhesion onto pulmonary cells. Also, it is not clear if IL-15 induces a neutrophilic inflammation in vivo. This study was aimed at elucidation of these issues. Neutrophils were treated with IL-15 and cell surface expression of CD11a, CD11b, CD11c and CD18 was monitored by flow cytometry. The human respiratory epithelial A549 cell line was used as a substrate for the neutrophil adhesion assay and cell surface expression of CD50, CD54 and CD106 was monitored in IL-15-induced A549 cells. The murine air pouch model was used for investigating potential neutrophilic inflammation induced by IL-15 in vivo. IL-15 significantly increased neutrophil cell surface expression of CD11b and CD18 and up-regulated A549 cell surface expression of CD54. Moreover, A549 cells were found to express IL-15R components and adhesion of neutrophils onto A549 cells was increased when neutrophils or A549 cells were treated with IL-15. Finally, IL-15 induced neutrophilic inflammation in vivo and concentrations of IL-6 and CXCL2/MIP-2 were increased in IL-15-induced pouches. IL-15 might participate in inflammatory pulmonary diseases by attracting neutrophils, modulating cell surface expression molecules and increasing neutrophil adhesion onto pulmonary cells. PMID:15996196

  8. [Comparative Proteomic Analysis of Human Lung Adenocarcinoma Cisplatin-resistant Cell Strain A549/CDDP.].

    PubMed

    Shi, Sien; Wang, Jianjun; Zhai, Wei; Gao, Guizhou; Tang, Jian; Fan, Kai; Tang, Zheng; Wei, Liang

    2009-11-20

    Chemotherapy plays an important role in the comprehensive therapy of lung cancer. However, the drug-resistance often causes the failure of the chemotherapy. The aim of this study is to identify differently expressed protein before and after cisplatin resistance of human lung adenocarcinoma cell A549 by proteomic analysis. Cisplatin-resistant cell strain A549/CDDP was established by combining gradually increasing concentration of cisplatin with large dosage impact. Comparative proteomic analysis of A549 and A549/CDDP were carried out by means of two-dimensional gel electrophoresis. The differentially expressed proteins were detected and identified by MALDI-TOF mass spectrometry. Eighty-two differentially expressed proteins were screened by analysis the electrophoretic maps of A549 and A549/CDDP. Six differential proteins were analyzed by peptide mass fingerprinting. Glucose regulating protein 75, ribosomal protein S4, mitochondrial ATP synthase F1 complex beta subunit and immunoglobulin heavy chain variable region were identified. All four differentially expressed proteins were over-expressed in A549/CDDP, whereas low-expressed or no-expressed in A549. These differentially expressed proteins give some clues to elucidate the mechanism of lung cancer cell resistant of cisplatin, providing the basis of searching for potential target of chemotherapy of lung cancer.

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

    PubMed

    Miyayama, Takamitsu; Matsuoka, Masato

    2016-01-01

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

  10. Bostrycin inhibits proliferation of human lung carcinoma A549 cells via downregulation of the PI3K/Akt pathway

    PubMed Central

    2011-01-01

    Background Bostrycin is a novel compound isolated from marine fungi that inhibits proliferation of many cancer cells. However, the inhibitory effect of bostrycin on lung cancers has not been reported. This study is to investigate the inhibitory effects and mechanism of bostrycin on human lung cancer cells in vitro. Methods We used MTT assay, flow cytometry, microarray, real time PCR, and Western blotting to detect the effect of bostrycin on A549 human pulmonary adenocarcinoma cells. Results We showed a significant inhibition of cell proliferation and induction of apoptosis in bostrycin-treated lung adenocarcinoma cells. Bostrycin treatment caused cell cycle arrest in the G0/G1 phase. We also found the upregulation of microRNA-638 and microRNA-923 in bostrycin-treated cells. further, we found the downregulation of p110α and p-Akt/PKB proteins and increased activity of p27 protein after bostrycin treatment in A549 cells. Conclusions Our study indicated that bostrycin had a significant inhibitory effect on proliferation of A549 cells. It is possible that upregulation of microRNA-638 and microRNA-923 and downregulaton of the PI3K/AKT pathway proteins played a role in induction of cell cycle arrest and apoptosis in bostrycin-treated cells. PMID:21303527

  11. [Radiosensitizing effect of erlotinib on human lung adenocarcinoma cell line A549].

    PubMed

    Liu, Xiao-qun; Qiao, Tian-kui

    2013-11-01

    To explore the radiosensitizing effect of erlotinib on human lung adenocarcinoma cell line A549 cells and the related mechanisms. The inhibitory effect of erlotinib on A549 cells was assessed by MTT assay, and its IC50 concentration was calculated. The radiosensitization was evaluated by the method of clone forming assay. Flow cytometry was used to analyze the effect of erlotinib on cell cycle and apoptosis. The growth of A549 cells was inhibited after the cells were exposed to erlotinib for 48 hours. Moreover, the inhibitory rates increased with the increase of erlotinib concentrations, and IC50 was 19.26 µmol/L. In contrast to the irradiation alone group, the survival rates of the cells in erlotinib plus irradiation groups decreased, and erlotinib enhanced the radiosensitivity of the A549 cells. This effect was further increased as cells were exposed to erlotinib for a longer time. In the irradiation alone group and the two groups exposed to erlotinib for 24 hours and 48 hours before irradiation, D0 values were 3.01 Gy, 2.58 Gy and 2.45 Gy respectively, and Dq values were 2.16 Gy, 1.94 Gy and 1.61 Gy, respectively. In the last two groups, SERD0 values were 1.17 and 1.23, respectively. The flow cytometry analysis showed that erlotinib induced G2/M phase arrest and increased the apoptosis rate in A549 cells. With the increase of exposure time, the effects were more significant. Erlotinib inhibits the A549 cell growth and enhances the radiosensitivity of A549 cells in vitro. The radiosensitizing mechanisms might be related to inhibiting repair of sublethal injury and inducing G2/M phase arrest and apoptosis.

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

    PubMed

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

    2018-02-01

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

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

    SciTech Connect

    Huang, Chunrong; Zheng, Haichong; He, Wanmei

    2016-05-20

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

  14. Fucoidan from Undaria pinnatifida induces apoptosis in A549 human lung carcinoma cells.

    PubMed

    Boo, Hye-Jin; Hyun, Jae-Hee; Kim, Sang-Cheol; Kang, Jung-Il; Kim, Min-Kyoung; Kim, Sun-Yeou; Cho, Heeyeong; Yoo, Eun-Sook; Kang, Hee-Kyoung

    2011-07-01

    Fucoidan, a sulfated polysaccharide, has various biological activities, such as anticancer, antiangiogenic and antiinflammatory effects; however, the mechanisms of action of fucoidan on anticancer activity have not been fully elucidated. The anticancer effects of fucoidan from Undaria pinnatifida on A549 human lung carcinoma cells were examined. Treatment of A549 cells with fucoidan resulted in potent antiproliferative activity. Also, some typical apoptotic characteristics, such as chromatin condensation and an increase in the population of sub-G1 hypodiploid cells, were observed. With respect to the mechanism underlying the induction of apoptosis, fucoidan reduced Bcl-2 expression, but the expression of Bax was increased in a dose-dependent manner compared with the controls. Furthermore, fucoidan induced caspase-9 activation, but decreased the level of procaspase-3. Cleavage of poly-ADP-ribose polymerase (PARP), a vital substrate of effector caspase, was found. The study further investigated the role of the MAPK and PI3K/Akt pathways with respect to the apoptotic effect of fucoidan, and showed that fucoidan activates ERK1/2 in A549 cells. Unlike ERK1/2, however, treatment with fucoidan resulted in the down-regulation of phospho-p38 expression. In addition, fucoidan resulted in the down-regulation of phospho-PI3K/Akt. Together, these results indicate that fucoidan induces apoptosis of A549 human lung cancer cells through down-regulation of p38, PI3K/Akt, and the activation of the ERK1/2 MAPK pathway. Copyright © 2011 John Wiley & Sons, Ltd.

  15. Tomatidine inhibits invasion of human lung adenocarcinoma cell A549 by reducing matrix metalloproteinases expression.

    PubMed

    Yan, Kun-Huang; Lee, Liang-Ming; Yan, Shao-Han; Huang, Hsiang-Ching; Li, Chia-Chen; Lin, Hui-Ting; Chen, Pin-Shern

    2013-05-25

    Tomatidine is an aglycone of glycoalkaloid tomatine in tomato. Tomatidine is found to possess anti-inflammatory properties and may serve as a chemosensitizer in multidrug-resistant tumor cells. However, the effect of tomatidine on cancer cell metastasis remains unclear. This study examines the effect of tomatidine on the migration and invasion of human lung adenocarcinoma A549 cell in vitro. The data demonstrates that tomatidine does not effectively inhibit the viability of A549 cells. When treated with non-toxic doses of tomatidine, cell invasion is markedly suppressed by Boyden chamber invasion assay, while cell migration is not affected. Tomatidine reduces the mRNA level of matrix metalloproteinase-2 (MMP-2), MMP-9 and increases the expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK), as well as tissue inhibitor of metalloproteinase-1 (TIMP-1). The immunoblotting assays indicate that tomatidine is very effective in suppressing the phosphorylation of Akt and extracellular signal regulating kinase (ERK). In addition, tomatidine significantly decreases the nuclear level of nuclear factor kappa B (NF-κB), which suggests that tomatidine inhibits NF-κB activity. Furthermore, the treatment of inhibitors specific for PI3K/Akt (LY294002), ERK (U0126), or NF-κB (pyrrolidine dithiocarbamate) to A549 cells reduced cell invasion and MMP-2/9 expression. The results suggest that tomatidine inhibits the invasion of A549 cells by reducing the expression of MMPs. It also inhibits ERK and Akt signaling pathways and NF-κB activity. These findings demonstrate a new therapeutic potential for tomatidine in anti-metastatic therapy. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  16. Ciclesonide uptake and metabolism in human alveolar type II epithelial cells (A549)

    PubMed Central

    Nonaka, Takashi; Nave, Rüdiger; McCracken, Nigel; Kawashimo, Atsuko; Katsuura, Yasuhiro

    2007-01-01

    Background Ciclesonide is a novel inhaled corticosteroid for the treatment of airway inflammation. In this study we investigated uptake and in vitro metabolism of ciclesonide in human alveolar type II epithelial cells (A549). Ciclesonide uptake was compared with fluticasone propionate, an inhaled corticosteroid that is not metabolized in lung tissue. A549 cells were incubated with 2 × 10-8 M ciclesonide or fluticasone propionate for 3 to 30 min to determine uptake; or with 2 × 10-8 M ciclesonide for 1 h, followed by incubation with drug-free buffer for 3, 6, and 24 h to analyze in vitro metabolism. High performance liquid chromatography with tandem mass spectrometry was used to measure the concentrations of both corticosteroids and metabolites. Results At all time points the mean intracellular concentration was higher for ciclesonide when compared with fluticasone propionate. Activation of ciclesonide to desisobutyryl-ciclesonide (des-CIC) was confirmed and conjugates of des-CIC with fatty acids were detected. The intracellular concentration of ciclesonide decreased over time, whereas the concentration of des-CIC remained relatively stable: 2.27 to 3.19 pmol/dish between 3 and 24 h. The concentration of des-CIC fatty acid conjugates increased over time, with des-CIC-oleate being the main metabolite. Conclusion Uptake of ciclesonide into A549 cells was more efficient than that of the less lipophilic fluticasone propionate. Intracellular concentrations of the pharmacologically active metabolite des-CIC were maintained for up to 24 h. The local anti-inflammatory activity of ciclesonide in the lung may be prolonged by the slow release of active drug from the depot of fatty acid esters. PMID:17900334

  17. Cimicifuga foetida L. inhibited human respiratory syncytial virus in HEp-2 and A549 cell lines.

    PubMed

    Wang, Kuo Chih; Chang, Jung San; Chiang, Lien Chai; Lin, Chun Ching

    2012-01-01

    Human respiratory syncytial virus (HRSV) causes serious pediatric infection of the lower respiratory tract without effective therapeutic modality. Sheng-Ma-Ge-Gen-Tang (SMGGT; Shoma-kakkon-to) has been proven to be effective at inhibiting HRSV-induced plaque formation, and Cimicifuga foetida is the major constituent of SMGGT. We tested the hypothesis that C. foetida effectively inhibited the cytopathic effects of HRSV by a plaque reduction assay in both human upper (HEp2) and lower (A549) respiratory tract cell lines. Its ability to stimulate anti-viral cytokines was evaluated by an enzyme-linked immunosorbent assay (ELISA). C. foetida dose-dependently inhibited HRSV-induced plaque formation (p < 0.0001) before and after viral inoculation, especially in A549 cells (p < 0.0001). C. foetida dose-dependently inhibited viral attachment (p < 0.0001) and could increase heparins effect on viral attachment. In addition, C. foetida time-dependently and dose-dependently (p < 0.0001) inhibited HRSV internalization. C. foetida could stimulate epithelial cells to secrete IFN-β to counteract viral infection. However, C. foetida did not stimulate TNF-α secretion. Therefore, C. foetida could be useful in managing HRSV infection. This is the first evidence to support that C. foetida possesses antiviral activity.

  18. [Effects of 17-AAG on the proliferation and apoptosis of human lung cancer A549 and H446 cells].

    PubMed

    Niu, Ben; Lin, Jingshuang; Feng, Tao

    2015-04-01

    To observe the effect of 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) on the apoptosis of human lung cancer cell lines A549 and H446, and to investigate the potential mechanisms. Proliferation inhibition and apoptosis assays, and the cell cycles were detected by MTT and flow cytometry respectively. Western blot was used to determine the expression level of proteins such as Hsp90, Hsp70, AKt, Her-2, Bcl-2 and Bax. After treated with 17-AAG, the proliferation of both A549 and H446 cells was inhibited significantly in a dose-dependent manner; as the concentration of 17-AAG was from 50 to 500 nmol/L, the IC₅₀ values to A549 and H446 cell lines were (222 ± 13) nmol/L and (189 ± 7) nmol/L respectively at 48 h. Cell cycle assays showed that 17-AAG was able to arrest cell cycles of A549 and H446 cell lines at the G₂/M phase. Apoptosis assay showed that 17-AAG was capable of inducing apoptosis in A549 and H446 cell lines. After treated with 17-AAG for 48 h, there were significant differences between the 400 nmol/L groups(46.3% for A549 cell line and 56.9% for H446 cell line) and the control group (11.9% for A549 cell line and 6.9% for H446 cell line, P < 0.01). Western blot results showed that the relative proteins of the Hsp90 signal pathway in both A549 and H446 cell lines underwent similar changes after 17-AAG treatment: Akt and Her-2 decreased significantly while the expression of Hsp70 increased. Meanwhile, the expression of Bcl-2 decreased but that of Bax increased, indicating that 17-AAG was able to promote apoptosis mode in A549 and H446 cells. 17-AAG can regulate the expression level of apoptosis-related proteins such as Bax and Bcl-2 by Hsp90 signaling pathway in A549 and H446 cells, and ultimately inhibit cell proliferation and induce apoptosis.

  19. 4-Nitroquinoline-1-oxide effects human lung adenocarcinoma A549 cells by regulating the expression of POLD4

    PubMed Central

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

    2016-01-01

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

  20. Combined toxic effect of airborne heavy metals on human lung cell line A549.

    PubMed

    Choi, Yeowool; Park, Kihong; Kim, Injeong; Kim, Sang D

    2018-02-01

    Many studies have demonstrated that heavy metals existing as a mixture in the atmospheric environment cause adverse effects on human health and are important key factors of cytotoxicity; however, little investigation has been conducted on a toxicological study of a metal mixture from atmospheric fine particulate matter. The objective of this study was to predict the combined effects of heavy metals in aerosol by using in vitro human cells and obtain a suitable mixture toxicity model. Arsenic, nickel, and lead were selected for mixtures exposed to A549 human lung cancer cells. Cell proliferation (WST-1), glutathione (GSH), and interleukin (IL)-8 inhibition were observed and applied to the prediction models of mixture toxicity, concentration addition (CA) and independent action (IA). The total mixture concentrations were set by an IC 10 -fixed ratio of individual toxicity to be more realistic for mortality and enzyme inhibition tests. The results showed that the IA model was statistically closer to the observed results than the CA model in mortality, indicating dissimilar modes of action. For the GSH inhibition, the results predicted by the IA and CA models were highly overestimated relative to mortality. Meanwhile, the IL-8 results were stable with no significant change in immune reaction related to inflammation. In conclusion, the IA model is a rapid prediction model in heavy metals mixtures; mortality, as a total outcome of cell response, is a good tool for demonstrating the combined toxicity rather than other biochemical responses.

  1. Overview on biological implications of metal oxide nanoparticle exposure to human alveolar A549 cell line.

    PubMed

    Martin, Ansie; Sarkar, Angshuman

    2017-08-01

    Metal oxides (MeOx) are exponentially being used in a wide range of applications and are the largest class of commercially produced nanomaterials. This presents unprecedented human exposure. Thus, understanding nanoparticle induced cellular stress can greatly help design strategies to combat them. Scores of studies have been carried out to understand the effects of MeOx nanoparticle exposure on human alveolar cells, which are highly susceptible to aerosolized matter. There is a huge redundancy of information generated, also, a lack of a comprehensive conglomeration of this information. We have built here in a sincere summary of the cellular responses reported till date as a direct consequence of MeOx nanoparticle exposure on human alveolar (A549) cells. Detailed accounts of cellular morphology modulation, generation of reactive oxygen species (ROS) and oxidative stress, inflammation and cytokine release, genotoxic and epi-genotoxic insults, toxicological trend, nanoparticle internalization, modes of cell death, protein synthesis, and membrane damage among others are discussed. Finally, to aid predictability of the highly dynamic and multifactorial nature of this toxicity, we have hypothesized models that describe the ensuing mechanisms based on common patterns discovered throughout our literature survey.

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

    NASA Astrophysics Data System (ADS)

    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.

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

    PubMed

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

    2010-12-01

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

  4. AMPK is required for PM2.5-induced autophagy in human lung epithelial A549 cells

    PubMed Central

    Wang, Yahong; Lin, Ziying; Huang, Haili; He, Huijuan; Yang, Lawei; Chen, Ting; Yang, Teng; Ren, Nina; Jiang, Yun; Xu, Wenya; Kamp, David W; Liu, Tie; Liu, Gang

    2015-01-01

    The aim is to investigate the molecular mechanisms underlying the PM2.5-induced autophagy in human lung cancer epithelial cells (A549). The effects of the PM2.5 on morphological and biochemical markers of autophagy in A549 were analyzed by electron microscopy, GFP-LC3 puncta was observed by confocal fluorescence microscope. The effects of phosphorylation of AMPK, mTOR, AKT, ERK, JNK, and p53 on LC3II in A549 were observed following PM2.5 exposure; the role of autophagy in PM2.5-induced apoptosis was examined using 3-methyladenine and rapamycin. PM2.5 induced morphological and biochemical markers of autophagy in A549. Phosphorylation of AMPK and dephosphorylation of mTOR were observed following PM2.5 treatment, and AMPK inhibitor blocked LC3B-II expression. In addition, we demonstrated that PM2.5-induced autophagy confers a pro-survival role in host defense. PMID:25784975

  5. Chloroquine inhibits cell growth in human A549 lung cancer cells by blocking autophagy and inducing mitochondrial‑mediated apoptosis.

    PubMed

    Liu, Likun; Han, Cuicui; Yu, Haitao; Zhu, Wenbin; Cui, Hongxia; Zheng, Lihong; Zhang, Chunjing; Yue, Liling

    2018-04-12

    Chloroquine (CQ) has been revealed to exhibit antitumor activity in several human tumors including lung cancer as mono‑ or add‑on therapy. The antitumor effect of CQ appears to depend on the tumor type, stage and genetic context. Few studies have focused on the mechanism concerning the antitumor effect of CQ monotherapy and the cause and effect relationship among autophagy, apoptosis and CQ in human lung A549 cells. Therefore, the present study aimed to identify the antitumor effect of CQ monotherapy and analyze the possible mechanism. In the present study, we demonstrated that CQ suppressed human A549 cell growth in a dose‑ and time‑dependent manner. CQ‑mediated growth inhibition in A549 cells was characterized by the targeting of the PI3K/AKT pathway, thus, inducing mitochondria‑mediated apoptosis at relatively higher concentrations by downregulating Bcl‑2 expression, increasing the expression level of Bax, decreasing mitochondrial membrane potential, releasing cytochrome c from the mitochondria into the cytosol, activating caspase‑3 and cleaving PARP. Collectively, these findings may offer a new rationale for using CQ as a lung cancer therapy drug in clinical practice.

  6. Oxidative stress and inflammatory response to printer toner particles in human epithelial A549 lung cells.

    PubMed

    Könczöl, Mathias; Weiß, Adilka; Gminski, Richard; Merfort, Irmgard; Mersch-Sundermann, Volker

    2013-02-04

    Reports on adverse health effects related to occupational exposure to toner powder are still inconclusive. Therefore, we have previously conducted an in vitro-study to characterize the genotoxic potential of three commercially available black printer toner powders in A549 lung cells. In these cell-based assays it was clearly demonstrated that the tested toner powders damage DNA and induce micronucleus (MN) formation. Here, we have studied the cytotoxic and proinflammatory potential of these three types of printer toner particles and the influence of ROS and NF-κB induction in order to unravel the underlying mechanisms. A549 cells were exposed to various concentrations of printer toner particle suspensions for 24 h. The toner particles were observed to exert significant cytotoxic effects in the WST-1 and neutral red (NR)-assays, although to a varying extent. Caspase 3/7 activity increased, while the mitochondrial membrane potential (MMP) was not affected. Particles of all three printer toner powders induced concentration-dependent formation of reactive oxygen species (ROS), as measured in the DCFH-DA assay. Furthermore, toner particle exposure enhanced interleukin-6 and interleukin-8 production, which is in agreement with activation of the transcription factor NF-κB in A549 cells shown by the electrophoretic mobility shift assay (EMSA). Therefore, it can be concluded that exposure of A549 lung cells to three selected printer toner powders caused oxidative stress through induction of ROS. Increased ROS formation may trigger genotoxic effects and activate proinflammatory pathways. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  7. Dexamethasone-(C21-phosphoramide)-[anti-EGFR]: molecular design, synthetic organic chemistry reactions, and antineoplastic cytotoxic potency against pulmonary adenocarcinoma (A549)

    PubMed Central

    Coyne, Cody P; Narayanan, Lakshmi

    2016-01-01

    Purpose Corticosteroids are effective in the management of a variety of disease states, such as several forms of neoplasia (leukemia and lymphoma), autoimmune conditions, and severe inflammatory responses. Molecular strategies that selectively “target” delivery of corticosteroids minimize or prevents large amounts of the pharmaceutical moiety from passively diffusing into normal healthy cell populations residing within tissues and organ systems. Materials and methods The covalent immunopharmaceutical, dexamethasone-(C21-phosphoramide)-[anti-EGFR] was synthesized by reacting dexamethasone-21-monophosphate with a carbodiimide reagent to form a dexamethasone phosphate carbodiimide ester that was subsequently reacted with imidazole to create an amine-reactive dexamethasone-(C21-phosphorylimidazolide) intermediate. Monoclonal anti-EGFR immunoglobulin was combined with the amine-reactive dexamethasone-(C21-phosphorylimidazolide) intermediate, resulting in the synthesis of the covalent immunopharmaceutical, dexamethasone-(C21-phosphoramide)-[anti-EGFR]. Following spectrophotometric analysis and validation of retained epidermal growth factor receptor type 1 (EGFR)-binding avidity by cell-ELISA, the selective anti-neoplasic cytotoxic potency of dexamethasone-(C21-phosphoramide)-[anti-EGFR] was established by MTT-based vitality stain methodology using adherent monolayer populations of human pulmonary adenocarcinoma (A549) known to overexpress the tropic membrane receptors EGFR and insulin-like growth factor receptor type 1. Results The dexamethasone:IgG molar-incorporation-index for dexamethasone-(C21-phosphoramide)-[anti-EGFR] was 6.95:1 following exhaustive serial microfiltration. Cytotoxicity analysis: covalent bonding of dexamethasone to monoclonal anti-EGFR immunoglobulin did not significantly modify the ex vivo antineoplastic cytotoxicity of dexamethasone against pulmonary adenocarcinoma at and between the standardized dexamethasone equivalent concentrations of 10

  8. Dexamethasone-(C21-phosphoramide)-[anti-EGFR]: molecular design, synthetic organic chemistry reactions, and antineoplastic cytotoxic potency against pulmonary adenocarcinoma (A549).

    PubMed

    Coyne, Cody P; Narayanan, Lakshmi

    2016-01-01

    Corticosteroids are effective in the management of a variety of disease states, such as several forms of neoplasia (leukemia and lymphoma), autoimmune conditions, and severe inflammatory responses. Molecular strategies that selectively "target" delivery of corticosteroids minimize or prevents large amounts of the pharmaceutical moiety from passively diffusing into normal healthy cell populations residing within tissues and organ systems. The covalent immunopharmaceutical, dexamethasone-(C21-phosphoramide)-[anti-EGFR] was synthesized by reacting dexamethasone-21-monophosphate with a carbodiimide reagent to form a dexamethasone phosphate carbodiimide ester that was subsequently reacted with imidazole to create an amine-reactive dexamethasone-(C21-phosphorylimidazolide) intermediate. Monoclonal anti-EGFR immunoglobulin was combined with the amine-reactive dexamethasone-(C21-phosphorylimidazolide) intermediate, resulting in the synthesis of the covalent immunopharmaceutical, dexamethasone-(C21-phosphoramide)-[anti-EGFR]. Following spectrophotometric analysis and validation of retained epidermal growth factor receptor type 1 (EGFR)-binding avidity by cell-ELISA, the selective anti-neoplasic cytotoxic potency of dexamethasone-(C21-phosphoramide)-[anti-EGFR] was established by MTT-based vitality stain methodology using adherent monolayer populations of human pulmonary adenocarcinoma (A549) known to overexpress the tropic membrane receptors EGFR and insulin-like growth factor receptor type 1. The dexamethasone:IgG molar-incorporation-index for dexamethasone-(C21-phosphoramide)-[anti-EGFR] was 6.95:1 following exhaustive serial microfiltration. Cytotoxicity analysis: covalent bonding of dexamethasone to monoclonal anti-EGFR immunoglobulin did not significantly modify the ex vivo antineoplastic cytotoxicity of dexamethasone against pulmonary adenocarcinoma at and between the standardized dexamethasone equivalent concentrations of 10(-9) M and 10(-5) M. Rapid increases in

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

    SciTech Connect

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

    2010-08-13

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

  10. A Comprehensive Proteomic View of Responses of A549 Type II Alveolar Epithelial Cells to Human Respiratory Syncytial Virus Infection*

    PubMed Central

    Dave, Keyur A.; Norris, Emma L.; Bukreyev, Alexander A.; Headlam, Madeleine J.; Buchholz, Ursula J.; Singh, Toshna; Collins, Peter L.; Gorman, Jeffrey J.

    2014-01-01

    Human respiratory syncytial virus is a major respiratory pathogen for which there are no suitable antivirals or vaccines. A better understanding of the host cell response to this virus may redress this problem. The present report concerns analysis of multiple independent biological replicates of control and 24 h infected lysates of A549 cells by two different proteomic workflows. One workflow involved fractionation of lysates by in-solution protein IEF and individual fractions were digested using trypsin prior to capillary HPLC-LTQ-OrbitrapXL-MS/MS. A second workflow involved digestion of whole cell lysates and analysis by nanoUltraHPLC-LTQ-OrbitrapElite-MS/MS. Both workflows resulted in the quantification of viral proteins exclusively in lysates of infected cells in the relative abundances anticipated from previous studies. Unprecedented numbers (3247 - 5010) of host cell protein groups were also quantified and the infection-specific regulation of a large number (191) of these protein groups was evident based on a stringent false discovery rate cut-off (<1%). Bioinformatic analyses revealed that most of the regulated proteins were potentially regulated by type I, II, and III interferon, TNF-α and noncanonical NF-κB2 mediated antiviral response pathways. Regulation of specific protein groups by infection was validated by quantitative Western blotting and the cytokine-/key regulator-specific nature of their regulation was confirmed by comparable analyses of cytokine treated A549 cells. Overall, it is evident that the workflows described herein have produced the most comprehensive proteomic characterization of host cell responses to human respiratory syncytial virus published to date. These workflows will form the basis for analysis of the impacts of specific genes of human respiratory syncytial virus responses of A549 and other cell lines using a gene-deleted version of the virus. They should also prove valuable for the analysis of the impact of other infectious

  11. Antimetastatic effects of Phyllanthus on human lung (A549) and breast (MCF-7) cancer cell lines.

    PubMed

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

    2011-01-01

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

  12. The effects of combined treatment with sevoflurane and cisplatin on growth and invasion of human adenocarcinoma cell line A549.

    PubMed

    Liang, Hua; Wang, Han Bing; Liu, Hong Zhen; Wen, Xian Jie; Zhou, Qiao Ling; Yang, Cheng Xiang

    2013-07-01

    Sevoflurane, an inhalational anesthetic, and cisplatin (DDP)-based chemotherapy have been widely used during lung cancer surgery. However, the effect of sevoflurane on the sensitivity of lung cancer cells to DDP chemotherapy remains unclear. In this study, the effects of combined treatment with sevoflurane and cisplatin on the growth and invasion of human lung adenocarcinoma A549 cell line have been investigated. The underlying mechanism has also been explored. In our experiment, A549 cells were treated with 2.5% sevoflurane, 10μmol/L DDP, or the co-treatment of sevoflurane and DDP for 4h, respectively. Cell proliferation was evaluated by the MTT assay and colony formation assay. Apoptosis was assessed by flow cytometry. Cell invasion was detected by Transwell assay. The expressions of X-linked inhibitor of apoptosis protein (XIAP), Survivin, matrix metalloproteinase (MMP)-2 and MMP-9 were determined by western blotting. Our results showed that sevoflurane combined with DDP resulted in a more pronounced inhibition of tumor cells growth and invasion as compared with either drug alone. Besides, XIAP, Survivin, MMP-2, and MMP-9 were downregulated more significantly by the co-treatment of the two drugs as compared to sevoflurane treatment or DDP treatment alone. Taken together, the growth-inhibitory and invasion-inhibitory synergy between sevoflurane and DDP in human adenocarcinoma A549 cell line was found in this study. Furthermore, we showed that the growth-inhibitory synergy between sevoflurane and DDP might be associated with the downregulation of XIAP and Survivin, and the invasion-inhibitory synergy between sevoflurane and DDP might be involved in the downregulation of MMP-2 and MMP-9. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

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

    PubMed

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

    2018-02-15

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

  14. Coenzyme Q0 from Antrodia cinnamomea in Submerged Cultures Induces Reactive Oxygen Species-Mediated Apoptosis in A549 Human Lung Cancer Cells

    PubMed Central

    Chung, Cheng-Han; Lee, Kung-Ta

    2014-01-01

    We investigated the anticancer effects of Antrodia cinnamomea, a medicinal mushroom from Taiwan, on A549 human lung cancer cells using the ethyl acetate extract from submerged culture filtrates. Our results showed that 2,3-dimethoxy-5-methyl-1,4-benzoquinone (coenzyme Q0; CoQ0) derived from A. cinnamomea submerged culture filtrates has anticancer activity. CoQ0 treatment reduced the viability of A549, HepG2, and SW480 cancer cell lines. Furthermore, CoQ0 induced reactive oxygen species (ROS) generation and apoptosis in A549 cells, which was inhibited by the antioxidant ascorbic acid. To our knowledge, these data demonstrate for the first time that CoQ0 derived from A. cinnamomea submerged culture filtrates exerts its anticancer effect through the induction of ROS-mediated apoptosis in A549 human lung cancer cells. PMID:25431605

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

    PubMed Central

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

    2011-01-01

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

  16. Screening of Stat3 inhibitory effects of Korean herbal medicines in the A549 human lung cancer cell line.

    PubMed

    Park, Jong-Shik; Bang, Ok-Sun; Kim, Jinhee

    2014-06-01

    The transcription factor signal transducer and activator of transcription 3 (Stat3) is constitutively activated in many human cancers. It promotes tumor cell proliferation, inhibits apoptosis, induces angiogenesis and metastasis, and suppresses antitumor host immune responses. Therefore, Stat3 has emerged as a promising molecular target for cancer therapies. In this study, we evaluated the Stat3-suppressive activity of 38 herbal medicines traditionally used in Korea. Medicinal herb extracts in 70% ethanol were screened for their ability to suppress Stat3 in the A549 human lung cancer cell line. A Stat3-responsive reporter assay system was used to detect intracellular Stat3 activity in extract-treated cells, and Western blot analyses were performed to measure the expression profiles of Stat3-regulated proteins. Fifty percent of the 38 extracts possessed at least mild Stat3-suppressive activities (i.e., activity less than 75% of the vehicle control). Ethanol extracts of Bupleurum falcatum L., Taraxacum officinale Weber, Solanum nigrum L., Ulmus macrocarpa Hance, Euonymus alatus Sieb., Artemisia capillaris Thunb., and Saururus chinensis (Lour.) Baill inhibited up to 75% of the vehicle control Stat3 activity level. A549 cells treated with these extracts also had reduced Bcl-xL, Survivin, c-Myc, and Mcl-1 expression. Many medicinal herbs traditionally used in Korea contain Stat3 activity-suppressing substances. Because of the therapeutic impact of Stat3 inhibition, these results could be useful when developing novel cancer therapeutics from medicinal herbs.

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

    NASA Astrophysics Data System (ADS)

    Syaefudin; Juniarti, A.; Rosiyana, L.; Setyani, A.; Khodijah, S.

    2016-01-01

    The aim of the present study is to evaluate cytotoxicity effect of nanoparticles of Selaginella doederleinii (S. doederleinii) leaves extract. S. doederleinii was extracted by maceration method using 70%(v/v) ethanol as solvent. Phytochemical content was analyzed qualitatively by using Harborne and Thin Layer Chromatography (TLC) methods. Nanoparticle extract was prepared by ionic gelation using chitosan as encapsulant agent. Anticancer activity was performed by using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The results showed that S. doederleinii contains of flavonoids. Nanoparticle of S. doederleinii leaves extract greatly inhibited A549 cells growth (cancer cells), with IC50 of 3% or 1020 μg/ml. These nanoparticles extract also inhibited the growth of Chang cells (normal cells), with IC50 of 4% or 1442 μg/ml. The effective concentration of nanoparticles extract which inhibits cancer cells without harming the normal cells is 0.5% or 167 μg/ml. Further studies are needed to obtain the concentration of nanoparticles extract which can selectively suppress cancer cells.

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

    SciTech Connect

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

    2005-08-26

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

  19. Cathepsin L is involved in cathepsin D processing and regulation of apoptosis in A549 human lung epithelial cells.

    PubMed

    Wille, Aline; Gerber, Annegret; Heimburg, Anke; Reisenauer, Anita; Peters, Christoph; Saftig, Paul; Reinheckel, Thomas; Welte, Tobias; Bühling, Frank

    2004-07-01

    Cathepsins are implicated in a multitude of physiological and pathophysiological processes. The aim of the present study was to investigate the function of cathepsin L (catL) in the proteolytic network of human lung epithelial cells and its role in the regulation of apoptosis. We found that catL-deficient A549 cells as well as lung tissue extracts of catL(-/-) mice express increased amounts of single-chain cathepsin D (catD). Degradation experiments indicate that catL specifically degrades the single-chain isoform of catD. Furthermore, we found that catL-deficient cells showed increased sensitivity to apoptosis. Finally, we demonstrate that the inhibition of catD activity by pepstatin A decreased the number of apoptotic cells in catL-deficient A549 cells after anti-Fas treatment. In conclusion, catL is involved in catD processing and the accumulation of catD isoforms in catL-deficient cells is associated with increased rates of spontaneous and anti-Fas-induced apoptosis.

  20. In vitro cytotoxicity and morphological assessment of smoke from polymer combustion in human lung derived cells (A549).

    PubMed

    Lestari, F; Hayes, A J; Green, A R; Chattopadhyay, G

    2012-04-01

    The application of polymer and composites in building and modern transport interiors raises concerns of potential health hazards during combustion. Cytotoxicity and morphological assessment of smoke from polymer combustion in human lung derived cells (A549) has been investigated. A laboratory scale vertical tube furnace was used for the generation of combustion products. A range of materials used in the building and transport industry including high density-polyethylene (HDPE), polypropylene (PP), polycarbonate (PC), and polyvinyl chloride (PVC), fiberglass reinforced polymers (FRPs), and melamine faced plywood (MFP) were studied. The exposure of combustion toxicants to human lung cells (A549) at the air/liquid interface was acquired using a Harvard Navicyte Chamber. Cytotoxic effects on human cells were assessed based on cell viability using a selected in vitro cytotoxicity assays, including NRU (neutral red uptake) and ATP (adenosine triphosphate). Morphological assessment on the effects of combustion products in human lung cells from selected materials including PVC, FRP and MFP was assessed using scanning electron microscopy (SEM). The volatile organic compounds from thermal decomposition products were identified using ATD-GCMS (Automatic Thermal Desorption Gas Chromatography Mass Spectrometry). NOAEC (No Observable Adverse Effect Concentration), IC(10) (10% inhibitory concentration), IC(50) (50% inhibitory concentration), and TLC (Total Lethal Concentration) values (mg/l) were generated. The following toxicity ranking was observed from the most toxic material to the least toxic using the NRU assay: PVC>PP>HDPE>PC >FRP-10>MFP>FRP-16; and the ATP assay: PVC>HDPE>PP>FRP-10>FRP-16>MFP>PC. The method described here could potentially be an alternative to current fire toxicity standards. Copyright © 2011 Elsevier GmbH. All rights reserved.

  1. Portulaca oleracea Seed Oil Exerts Cytotoxic Effects on Human Liver Cancer (HepG2) and Human Lung Cancer (A-549) Cell Lines.

    PubMed

    Al-Sheddi, Ebtesam Saad; Farshori, Nida Nayyar; Al-Oqail, Mai Mohammad; Musarrat, Javed; Al-Khedhairy, Abdulaziz Ali; Siddiqui, Maqsood Ahmed

    2015-01-01

    Portulaca oleracea (Family: Portulacaceae), is well known for its anti-inflammatory, antioxidative, anti- bacterial, and anti-tumor activities. However, cytotoxic effects of seed oil of Portulaca oleracea against human liver cancer (HepG2) and human lung cancer (A-549) cell lines have not been studied previously. Therefore, the present study was designed to investigate the cytotoxic effects of Portulaca oleracea seed oil on HepG2 and A-549 cell lines. Both cell lines were exposed to various concentrations of Portulaca oleracea seed oil for 24h. After the exposure, percentage cell viability was studied by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) (MTT), neutral red uptake (NRU) assays, and cellular morphology by phase contrast inverted microscopy. The results showed a concentration-dependent significant reduction in the percentage cell viability and an alteration in the cellular morphology of HepG2 and A-549 cells. The percentage cell viability was recorded as 73%, 63%, and 54% by MTT assay and 76%, 61%, and 50% by NRU assay at 250, 500, and 1000 μg/ml, respectively in HepG2 cells. Percentage cell viability was recorded as 82%, 72%, and 64% by MTT assay and 83%, 68%, and 56% by NRU assay at 250, 500, and 1000 μg/ml, respectively in A-549 cells. The 100 μg/ml and lower concentrations were found to be non cytotoxic to A-549 cells, whereas decrease of 14% and 12% were recorded by MTT and NRU assay, respectively in HepG2 cells. Both HepG2 and A-549 cell lines exposed to 250, 500, and 1000 μg/ ml of Portulaca oleracea seed oil lost their normal morphology, cell adhesion capacity, become rounded, and appeared smaller in size. The data from this study showed that exposure to seed oil of Portulaca oleracea resulted in significant cytotoxicity and inhibition of growth of the human liver cancer (HepG2) and human lung cancer (A-549) cell lines.

  2. On the cytotoxicity of some microbial volatile organic compounds as studied in the human lung cell line A549.

    PubMed

    Kreja, Ludwika; Seidel, Hans-Joachim

    2002-10-01

    The cytotoxicity of 13 microbial volatile organic compounds (MVOC) was studied using a human lung carcinoma epithelial cell line A549 in a colony formation assay and two colorimetric assays: the microculture tetrazolium assay (MTT assay) and the cellular protein assay (methylene blue-MB assay). For comparison, two known cytotoxic substances: the non-volatile mycotoxin gliotoxin and the mono-functional alkylating agent methyl methanesulfonate (MMS) were studied. Concentration-response curves for each agent were established and the IC50 value (concentration resulting in 50% inhibition of colony growth or absorbance) was estimated. There are differences in toxicity levels between the MVOC tested and gliotoxin and MMS. The most toxic MVOC was 1-decanol which was as effective as MMS in all test systems. 1-decanol was about 10-fold more toxic than the other MVOC. All MVOC tested were more than 1000-fold less toxic than gliotoxin.

  3. Epigallocatechin-3-Gallate Enhances the Therapeutic Effects of Leptomycin B on Human Lung Cancer A549 Cells

    PubMed Central

    Cromie, Meghan M.; Gao, Weimin

    2015-01-01

    Our previous studies have shown Leptomycin B (LMB) is a promising antilung cancer drug. Epigallocatechin-3-gallate (EGCG) has antitumor properties but a debatable clinical application. The objective of this study is to evaluate the combination therapeutic effect of LMB and EGCG and its molecular mechanisms in human lung cancer A549 cells. Increased cytotoxicity was observed in LMB+EGCG-treated cells compared to LMB-treated cells. Elevated ROS was maximized 2 h after treatment, and LMB+EGCG-treated cells had higher ROS levels compared to LMB. N-Acetyl-L-cysteine (NAC) studies confirmed the oxidative role of LMB and/or EGCG treatment. In comparison to the control, CYP3A4, SOD, GPX1, and p21 mRNA expression levels were increased 7.1-, 2.0-, 4.6-, and 13.1-fold in LMB-treated cells, respectively, while survivin was decreased 42.6-fold. Additionally, these increases of CYP3A4, SOD, and GPX1 were significantly reduced, while p21 was significantly increased in LMB+EGCG-treated cells compared to LMB-treated cells. The qRT-PCR results for p21 and survivin were further confirmed by Western blot. Our study first shows that LMB produces ROS and is possibly metabolized by CYP3A4, GPX1, and SOD in A549 cells, and combination treatment of LMB and EGCG augments LMB-induced cytotoxicity through enhanced ROS production and the modulation of drug metabolism and p21/survivin pathways. PMID:25922640

  4. Inhibitory effect of liquiritigenin on migration via downregulation proMMP-2 and PI3K/Akt signaling pathway in human lung adenocarcinoma A549 cells.

    PubMed

    Wang, Yu; Xie, Sirou; Liu, Changwei; Wu, Yuchen; Liu, Yuxin; Cai, Yunqing

    2012-01-01

    Liquiritigenin (LQ) is a flavanone extracted from Glycyrrhizae, which has multiple biological effects, such as antiinflammation and anticancer. This study is the first to investigate the effect of LQ on the migration of human lung adenocarcinoma A549 cells in vitro. First, LQ exhibited inhibitory effects on the adhesion and migration of A549 cells in the absence of cytotoxicity. Gelatin zymography and Western blot analysis showed that LQ significantly reduced the expression of promatrix metalloproteinase-2 (proMMP-2) in A549 cells in terms of both activity and protein level. Second, LQ inhibited the phosphorylation of Akt and activated the phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2). Furthermore, the treatment of inhibitors specific for Akt (LY294002) and ERK1/2 (U0126) to A549 cells resulted in reduced activity of proMMP-2. These results suggested that the inhibition on proMMP-2 expression by LQ may be through suppression on PI3K/Akt signaling pathway, which in turn led to the inhibition of lung adenocarcinoma A549 cells migration. However, activation of ERK might not be involved in the regulation of proMMP-2. Taken together, LQ may be considered as a potential interfering agent of cancer progression.

  5. Flavonoids isolated from Citrus platymamma induced G2/M cell cycle arrest and apoptosis in A549 human lung cancer cells

    PubMed Central

    Nagappan, Arulkumar; Lee, Ho Jeong; Saralamma, Venu Venkatarame Gowda; Park, Hyeon Soo; Hong, Gyeong Eun; Yumnam, Silvia; Raha, Suchismita; Charles, Shobana Nancy; Shin, Sung Chul; Kim, Eun Hee; Lee, Won Sup; Kim, Gon Sup

    2016-01-01

    Citrus platymamma hort. ex Tanaka belongs to the Rutaceae family and is widely used in folk medicines in Korea due to its anti-proliferative, anti-cancer, anti-oxidant, anti-inflammatory and anti-diabetic activities. However, the molecular mechanism of its anti-cancer effect is not well understood. The present study was conducted to elucidate the anti-cancer effect and molecular mechanism of flavonoids from Citrus platymamma (FCP) on A549 cells. FCP displayed concentration-dependent inhibition on A549 cells proliferation. Further, flow cytometry revealed that FCP significantly increased the sub-G1 (apoptotic cell population) and G2/M phase population, and the total number of apoptotic cells, in a dose-dependent manner. Nuclear condensation and fragmentation were also observed upon staining with Hoechst 33342 in FCP-treated A549 cells. Immunoblotting demonstrated a dose-dependent downregulation of cyclin B1, cyclin-dependent kinase 1, cell division cycle 25c, pro-caspases −3, −6, −8 and −9, and poly (adenosine diphosphate-ribose) polymerase (PARP) in FCP-treated A549 cells. In addition, FCP induced caspase-3 activation and subsequent PARP cleavage, and increased the B-cell lymphoma (Bcl)-2-associated X protein/Bcl-extra large ratio in A549 cells. These findings suggest that FCP induced G2/M arrest and apoptosis of A549 cells. The present study provides evidence that FCP may be useful in the treatment of human lung cancer. PMID:27446443

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

    PubMed

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

    2016-08-26

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

  7. 2,3,5,4-tetrahydroxy diphenylethylene-2-O-glucoside inhibits the adhesion and invasion of A549 human lung cancer cells

    PubMed Central

    Xu, Ming; Wang, Cong; Zhu, Minglin; Wang, Xianguo; Zhang, Li; Zhao, Jinping

    2017-01-01

    Lung cancer is considered to be a serious disease that poses a significant threat to human health. 2,3,5,4-tetrahydroxy diphenylethylene-2-O-glucoside (THSG) is a bioactive compound derived from Polygonum multiflorum Thunb. That has been demonstrated to possess antioxidative, anti-inflammatory and antitumor activities. However, little is currently known regarding the potential anticancer effects of this compound in lung cancer. Therefore, the present study aimed to investigate the effects of THSG on the adhesion and invasion of A549 human lung cancer cells in vitro, and to identify the putative mechanisms involved. Cell Counting kit-8 assay was performed to determine A549 cell viability following treatment with various doses (0, 5, 10, 25, 50, 100, 150 and 200 µM) of THSG for 12, 24 and 48 h. In addition, cell adhesion and invasion were determined following treatment of A549 cells with 0, 10, 25 or 50 µM THSG for 1, 2 or 3 h, respectively. Reverse transcription-quantitative polymerase chain reaction analysis was performed to examine the mRNA expression levels of Snail, E-cadherin, vimentin, matrix metalloproteinase (MMP) 2 and MMP9 following THSG treatment for 12 h. Western blot analysis was conducted to detect the protein expression levels of Snail, E-cadherin, vimentin, MMP2 and MMP9 following THSG treatment for 24 h. Treatment with THSG (10, 25 and 50 µM) significantly suppressed the adhesion and invasion of A549 human lung cancer cells in a dose-dependent manner. In addition, the mRNA and protein expression levels of adhesion and invasion-associated factors were decreased significantly in A549 cells treated with THSG. In conclusion, THSG effectively suppressed the adhesion and invasion of human lung cancer cells potentially by inhibiting the expression of adhesion and invasion-related genes. PMID:28990072

  8. SGK1 inhibits PM2.5-induced apoptosis and oxidative stress in human lung alveolar epithelial A549 cells.

    PubMed

    Li, Jin; Zhou, Qiulian; Yang, Tingting; Li, Yongqin; Zhang, Yuhui; Wang, Jinhua; Jiao, Zheng

    2018-02-19

    Emerging evidence demonstrated that particulate matter 2.5 (PM2.5) is an important environmental risk factor for lung diseases. Serum- and glucocorticoid-inducible kinase 1(SGK1) was reported to be a crucial factor for cell survival. However, the role of SGK1 in PM2.5-induced cell injury is still unclear. In this work, we firstly found that the expression of SGK1 was decreased in PM2.5-treated human lung alveolar epithelial (A549) cells by western blot. In addition, overexpression of SGK1 significantly attenuated A549 cell apoptosis and reduced the reactive oxygen species (ROS) generation induced by PM2.5. Moreover, we found that PM2.5 exposure significantly promoted the ERK1/2 activation and inhibited the AKT activation, whereas overexpression of SGK1 could reverse that. Finally, the results of the rescue experiment showed that MK2206 (AKT inhibitor) could rescue the impact of SGK1 on A549 cell apoptosis, while PD98059 (ERK1/2 inhibitor) could not further aggravate the impact. Taken together, our results suggest that SGK1 inhibits PM2.5-induced cell apoptosis and ROS generation via ERK1/2 and AKT signaling pathway in human lung alveolar epithelial A549 cells. Copyright © 2018 Elsevier Inc. All rights reserved.

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

    NASA Astrophysics Data System (ADS)

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

    2015-07-01

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

  10. Proteomic analysis of selective cytotoxic anticancer properties of flavonoids isolated from Citrus platymamma on A549 human lung cancer cells.

    PubMed

    Nagappan, Arulkumar; Venkatarame Gowda Saralamma, Venu; Hong, Gyeong Eun; Lee, Ho Jeong; Shin, Sung Chul; Kim, Eun Hee; Lee, Won Sup; Kim, Gon Sup

    2016-10-01

    Citrus platymamma Hort. ex Tanaka (Byungkyul in Korean) has been used in Korean folk medicine for the treatment of inflammatory disorders and cancer. However, the molecular mechanism underlying the anticancer properties of flavonoids isolated from C. platymamma (FCP) remains to be elucidated. Therefore, the present study attempted to identify the key proteins, which may be important in the anticancer effects of FCP on A549 cells using a proteomic approach. FCP showed a potent cytotoxic effect on the A549 human lung cancer cells, however, it had no effect on WI‑38 human fetal lung fibroblasts at the same concentrations. Furthermore, 15 differentially expressed protein spots (spot intensities ≥2‑fold change; P<0.05) were obtained from comparative proteome analysis of two‑dimensional gel electrophoresis maps of the control (untreated) and FCP‑treated A549 cells. Finally, eight differentially expressed proteins, one of which was upregulated and seven of which were downregulated, were successfully identified using matrix‑assisted laser desorption/ionization time‑of‑flight/time‑of‑flight tandem mass spectrometry and peptide mass fingerprinting analysis. Specifically, proteins involved in signal transduction were significantly downregulated, including annexin A1 (ANXA1) and ANXA4, whereas 14‑3‑3ε was upregulated. Cytoskeletal proteins, including cofilin‑1 (CFL1), cytokeratin 8 (KRT8) and KRT79, and molecular chaperones/heat shock proteins, including endoplasmin, were downregulated. Proteins involved in protein metabolism, namely elongation factor Ts were also downregulated. Consistent with results of the proteome analysis, the immunoblotting results showed that 14‑3‑3ε was upregulated, whereas CFL1, ANXA4 and KRT8 were downregulated in the FCP‑treated A549 cells. The majority of the proteins were involved in tumor growth, cell cycle, apoptosis, migration and signal transduction. These findings provide novel insights into the molecular

  11. Andrographolide antagonizes cigarette smoke extract-induced inflammatory response and oxidative stress in human alveolar epithelial A549 cells through induction of microRNA-218.

    PubMed

    Li, Ying-jie; Yu, Chang-hai; Li, Jing-bo; Wu, Xi-ya

    2013-12-01

    Andrographolide is a major bioactive labdane diterpenoid isolated from Andrographis paniculata and has protective effects against cigarette smoke (CS)-induced lung injury. This study was done to determine whether such protective effects were mediated through modulation of microRNA (miR)-218 expression. Therefore, we exposed human alveolar epithelial A549 cells to cigarette smoke extract (CSE) with or without andrographolide pretreatment and measured the level of glutathione, nuclear factor-kappaB (NF-κB) activation, proinflammatory cytokine production, and miR-218 expression. We found that andrographolide pretreatment significantly restored the glutathione level in CSE-exposed A549 cells, coupled with reduced inhibitor κB (IκB)-α phosphorylation and p65 nuclear translocation and interleukin (IL)-8 and IL-6 secretion. The miR-218 expression was significantly upregulated by andrographolide pretreatment. To determine the biological role of miR-218, we overexpressed and downregulated its expression using miR-218 mimic and anti-miR-218 inhibitor, respectively. We observed that miR-218 overexpression led to a marked reduction in IκB-α phosphorylation, p65 nuclear accumulation, and NF-κB-dependent transcriptional activity in CSE-treated A549 cells. In contrast, miR-218 silencing enhanced IκB-α phosphorylation and p65 nuclear accumulation in cells with andrographolide pretreatment and reversed andrographolide-mediated reduction of IL-6 and IL-8 production. In addition, depletion of miR-218 significantly reversed the upregulation of glutathione levels in A549 cells by andrographolide. Taken together, our results demonstrate that andrographolide mitigates CSE-induced inflammatory response in A549 cells, largely through inhibition of NF-κB activation via upregulation of miR-218, and thus has preventive benefits in CS-induced inflammatory lung diseases.

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

    PubMed Central

    2013-01-01

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

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

    PubMed

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

    2013-10-20

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

  14. Dual effects of human adipose tissue-derived mesenchymal stem cells in human lung adenocarcinoma A549 xenografts and colorectal adenocarcinoma HT-29 xenografts in mice.

    PubMed

    Rhyu, Jung Joo; Yun, Jun-Won; Kwon, Euna; Che, Jeong-Hwan; Kang, Byeong-Cheol

    2015-10-01

    Human adipose tissue-derived mesenchymal stem cells (hATMSCs) have great potential as a therapy for various diseases. However, emerging evidence shows that there are conflicting results concerning effects of hATMSCs on tumor progression. Our objective was to determine whether and how hATMSCs modulate tumor growth. After cancer cell lines were subcutaneously inoculated into BALB/c-nude and hairless severe combined immunodeficient mice, hATMSCs were intratumorally injected into the mice. The growth of the A549 tumors was inhibited by hATMSCs, yet that of the HT-29 tumors was significantly promoted by hATMSCs in the in vivo xenograft models. In vitro study using a co-culture system of cancer cells and hATMSCs was consistent with the in vivo experiments. To reveal the molecular events induced by hATMSCs in the xenograft models, global gene expression profiles of the A549 and HT-29 tumors in the absence or presence of hATMSCs were determined. Significant numbers of genes involved in biological processes were altered in the hATMSC-treated A549 tumors, whereas no biological process was regulated by treatment with hATMSCs in the HT-29 tumors, reflecting the different effects of hATMSCs in the different types of cancer. Notably, histone cluster 1, H2aj and neuropeptide Y receptor Y4 were found to be expressed in direct or inverse proportion to tumor size in both xenograft models. In addition, nuclear factor κB (NF-κB) p65 was differentially phosphorylated by the hATMSCs dependent on the source of the cancer cells. In conclusion, the identified gene profiling and NF-κB signaling provide molecular evidence to explain the conflicting findings in tumor‑MSC studies, although further study is needed to confirm these findings using various types of cancer.

  15. Inflammatory effects induced by selected limonene oxidation products: 4-OPA, IPOH, 4-AMCH in human bronchial (16HBE14o-) and alveolar (A549) epithelial cell lines.

    PubMed

    Lipsa, Dorelia; Leva, Paolo; Barrero-Moreno, Josefa; Coelhan, Mehmet

    2016-11-16

    Limonene, a monoterpene abundantly present in most of the consumer products (due to its pleasant citrus smell), easily undergoes ozonolysis leading to several limonene oxidation products (LOPs) such as 4-acetyl-1-methylcyclohexene (4-AMCH), 4-oxopentanal (4-OPA) and 3-isopropenyl-6-oxoheptanal (IPOH). Toxicological studies have indicated that human exposure to limonene and ozone can cause adverse airway effects. However, little attention has been paid to the potential health impact of specific LOPs, in particular of IPOH, 4-OPA and 4-AMCH. This study evaluates the cytotoxic effects of the selected LOPs on human bronchial epithelial (16HBE14o-) and alveolar epithelial (A549) cell lines by generating concentration-response curves using the neutral red uptake assay and analyzing the inflammatory response with a series of cytokines/chemokines. The cellular viability was mostly reduced by 4-OPA [IC 50 =1.6mM (A549) and 1.45mM (16HBE14o-)] when compared to IPOH [IC 50 =3.5mM (A549) and 3.4mM (16HBE14o-)] and 4-AMCH [IC 50 could not be calculated]. As a result from the inflammatory response, IPOH [50μM] induced an increase of both IL-6 and IL-8 secretion in A549 (1.5-fold change) and in 16HBE14o- (2.8- and 7-fold change respectively). 4-OPA [50μM] treatment of A549 increased IL-6 (1.4-times) and IL-8 (1.3-times) levels, while in 16HBE14o- had an opposite effect. A549 treated with 4-AMCH [50μM] elevate both IL-6 and IL-8 levels by 1.2-times, while in 16HBE14o- had an opposite effect. Based on our results, lung cellular injury characterized by inflammatory cytokine release was observed for both cell lines treated with the selected chemicals at concentrations that did not affect their cellular viability. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  16. Inhibition of cellular proliferation and induction of apoptosis in human lung adenocarcinoma A549 cells by T-type calcium channel antagonist.

    PubMed

    Choi, Doo Li; Jang, Sun Jeong; Cho, Sehyeon; Choi, Hye-Eun; Rim, Hong-Kun; Lee, Kyung-Tae; Lee, Jae Yeol

    2014-03-15

    The anti-proliferative and apoptotic activities of new T-type calcium channel antagonist, 6e (BK10040) on human lung adenocarcinoma A549 cells were investigated. The MTT assay results indicated that BK10040 was cytotoxic against human lung adenocarcinoma (A549) and pancreatic cancer (MiaPaCa2) cells in a dose-dependent manner with IC50 of 2.25 and 0.93μM, respectively, which is ca. 2-fold more potent than lead compound KYS05090 despite of its decreased T-type calcium channel blockade. As a mode of action for cytotoxic effect of BK10040 on lung cancer (A549) cells, this cancer cell death was found to have the typical features of apoptosis, as evidenced by the accumulation of positive cells for annexin V. In addition, BK10040 triggered the activations of caspases 3 and 9, and the cleavages of poly (ADP-ribose) polymerase (PARP). Moreover, the treatment with z-VAD-fmk (a broad spectrum caspase inhibitor) significantly prevented BK10040-induced apoptosis. Based on these results, BK10040 may be used as a potential therapeutic agent for human lung cancer via the potent apoptotic activity. Copyright © 2014 Elsevier Ltd. All rights reserved.

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

    PubMed

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

    2017-05-01

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

  18. Identification of side population cells in human lung adenocarcinoma A549 cell line and elucidation of the underlying roles in lung cancer.

    PubMed

    Xie, Tong; Mo, Lingzhao; Li, Li; Mao, Naiquan; Li, Danrong; Liu, Deseng; Zuo, Chuantian; Huang, Dingming; Pan, Qi; Yang, Li; Wang, Shoufeng

    2018-04-01

    The present study aimed to isolate and characterize side population (SP) cells in the human lung cancer A549 cell line, and elucidate the molecular mechanism of SP cells underlying lung cancer. The SP and non-SP (NSP) cells in A549 cells were isolated and their differentiation was analyzed by fluorescence-activated cell sorting. An in vitro plate clone assay, Matrigel ® Transwell assay and chemoresistance analysis of the sorted SP and NSP cells were performed. In addition, the sorted SP and NSP cells were injected into BALB/c nude mice to detect their tumorigenic potential in vivo . The expression of ATP-binding cassette sub-family G member 2 (ABCG2) in transplanted tumors was detected by immunohistochemistry. The SP and NSP cells were successfully isolated. The results demonstrated that SP cells accounted for 1.09% of live A549 cells. SP cells produced SP and NSP cells, while NSP cells only produced NSP cells. In addition, SP cells formed more colonies, exhibited improved invasive ability and increased levels of chemoresistance compared with NSP cells in vitro . SP cells demonstrated a higher tumorigenic potential in BALB/c nude mice, and the number of ABCG2-positive cells in the SP xenograft tumors were significantly increased compared with that in the NSP xenograft tumors. The present study indicated that SP cells isolated from the human lung cancer A549 cell line demonstrated increased tumorigenicity, and improved invasive ability and chemoresistance compared with NSP cells. In addition, detection of ABCG2 expression may assist in predicting the chemotherapeutic outcome of patients, and serve as a target for treating lung cancer.

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

    PubMed

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

    2014-09-01

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

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

    PubMed

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

    2017-08-01

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

  1. MiR-509-3-5p causes aberrant mitosis and anti-proliferative effect by suppression of PLK1 in human lung cancer A549 cells.

    PubMed

    Wang, Xian-Hui; Lu, Yao; Liang, Jing-Jing; Cao, Ji-Xiang; Jin, Ya-Qiong; An, Guo-Shun; Ni, Ju-Hua; Jia, Hong-Ti; Li, Shu-Yan

    2016-09-16

    MicroRNAs (miRNAs) are potent post-transcriptional regulators of gene expression and play roles in DNA damage response (DDR). PLK1 is identified as a modulator of DNA damage checkpoint. Although down-regulation of PLK1 by certain microRNAs has been reported, little is known about the interplay between PLK1 and miR-509-3-5p in DDR. Here we have demonstrated that miR-509-3-5p repressed PLK1 expression by targeting PLK1 3'-UTR, thereby causing mitotic aberration and growth arrest of human lung cancer A549 cells. Repression of PLK1 by miR-509-3-5p was further evidenced by over-expression of miR-509-3-5p in A549, HepG2 and HCT116p53(-/-) cancer cells, in which PLK1 protein was suppressed. Consistently, miR-509-3-5p was stimulated, while PLK1 protein was down-regulated in A549 cells exposed to CIS and ADR, suggesting that suppression of PLK1 by miR-509-3-5p is a component of CIS/ADR-induced DDR pathway. Flow cytometry and immunofluorescence labeling showed that over-expression of miR-509-3-5p in A549 induced G2/M arrest and aberrant mitosis characterized by abnormal bipolar mitotic spindles, condensed chromosomes, lagging DNA and chromosome bridges. In addition, over-expression of miR-509-3-5p markedly blocked A549 cell proliferation and sensitized the cells to CIS and ADR treatment. Taken together, miR-509-3-5p is a feasible suppressor for cancer by targeting PLK1. Our data may provide aid in potential design of combined chemotherapy and in our better understanding of the roles of microRNAs in response to DNA damage. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Combination therapy in A549 cells.

    PubMed

    Yuan, Menghui; Wang, Jing; Deng, Jinglan; Wang, Zhe; Yang, Weidong; Li, Guoquan; Ren, Bingxiu

    2010-04-01

    We investigated the anti-tumor effect induced by the combination of the radiotherapeutic agent (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). 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 (99m)Tc-RC-160 was assessed at different time points. The tumor inhibitory effects of (131)I-RC-160 and/or 5-FC were evaluated by measurement of tumor growth and immunohistochemical analysis. 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 (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. 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. Crown Copyright 2010. Published by Elsevier Inc. All rights reserved.

  3. Α-MMC and MAP30, two ribosome-inactivating proteins extracted from Momordica charantia, induce cell cycle arrest and apoptosis in A549 human lung carcinoma cells.

    PubMed

    Fan, Xiang; He, Lingli; Meng, Yao; Li, Gangrui; Li, Linli; Meng, Yanfa

    2015-05-01

    α‑Momorcharin (α‑MMC) and momordica anti‑human immunodeficiency virus protein (MAP30), produced by Momordica charantia, are ribosome‑inactivating proteins, which have been reported to exert inhibitory effects on cultured tumor cells. In order to further elucidate the functions of these agents, the present study aimed to investigate the effects of α‑MMC and MAP30 on cell viability, the induction of apoptosis, cell cycle arrest, DNA integrity and superoxide dismutase (SOD) activity. α‑MMC and MAP30 were purified from bitter melon seeds using ammonium sulfate precipitation in combination with sulfopropyl (SP)‑sepharose fast flow, sephacryl S‑100 and macro‑Cap‑SP chromatography. MTT, flow cytometric and DNA fragmentation analyses were then used to determine the effects of α‑MMC and MAP30 on human lung adenocarcinoma epithelial A549 cells. The results revealed that A549 cells were sensitive to α‑MMC and MAP30 cytotoxicity assays in vitro. Cell proliferation was significantly suppressed following α‑MMC and MAP30 treatment in a dose‑ and time‑dependent manner; in addition, the results indicated that MAP30 had a more potent anti‑tumor activity compared with that of α‑MMC. Cell cycle arrest in S phase and a significantly increased apoptotic rate were observed following treatment with α‑MMC and MAP30. Furthermore, DNA integrity analysis revealed that the DNA of A549 cells was degraded following treatment with α‑MMC and MAP30 for 48 h. The pyrogallol autoxidation method and nitrotetrazolium blue chloride staining were used to determine SOD activity, the results of which indicated that α‑MMC and MAP30 did not possess SOD activity. In conclusion, the results of the present study indicated that α‑MMC and MAP30 may have potential as novel therapeutic agents for the prophylaxis and treatment of cancer.

  4. Titanium dioxide nanoparticles-mediated in vitro cytotoxicity does not induce Hsp70 and Grp78 expression in human bronchial epithelial A549 cells.

    PubMed

    Aueviriyavit, Sasitorn; Phummiratch, Duangkamol; Kulthong, Kornphimol; Maniratanachote, Rawiwan

    2012-10-01

    Titanium dioxide nanoparticles (TiO(2)NPs) are increasingly being used in various industrial applications including the production of paper, plastics, cosmetics and paints. With the increasing number of nano-related products, the concern of governments and the general public about the health and environmental risks, especially with regard to occupational and other environmental exposure, are gradually increasing. However, there is insufficient knowledge about the actual affects upon human health and the environment, as well as a lack of suitable biomarkers for assessing TiO(2)NP-induced cytotoxicity. Since the respiratory tract is likely to be the main exposure route of industrial workers to TiO(2)NPs, we investigated the cytotoxicity of the anatase and rutile crystalline forms of TiO(2)NPs in A549 cells, a human alveolar type II-like epithelial cell line. In addition, we evaluated the transcript and protein expression levels of two heat shock protein (HSP) members, Grp78 and Hsp70, to ascertain their suitability as biomarkers of TiO(2)NP-induced toxicity in the respiratory system. Ultrastructural observations confirmed the presence of TiO(2)NPs inside cells. In vitro exposure of A549 cells to the anatase or rutile forms of TiO(2)NPs led to cell death and induced intracellular ROS generation in a dose-dependent manner, as determined by the MTS and dichlorofluorescein (DCF) assays, respectively. In contrast, the transcript and protein expression levels of Hsp70 and Grp78 did not change within the same TiO(2)NPs dose range (25-500 μg/ml). Thus, whilst TiO(2)NPs can cause cytotoxicity in A549 cells, and thus potentially in respiratory cells, Hsp70 and Grp78 are not suitable biomarkers for evaluating the acute toxicological effects of TiO(2)NPs in the respiratory system.

  5. Responses of A549 human lung epithelial cells to cristobalite and α-quartz exposures assessed by toxicoproteomics and gene expression analysis.

    PubMed

    Vuong, Ngoc Q; Goegan, Patrick; De Rose, Francesco; Breznan, Dalibor; Thomson, Errol M; O'Brien, Julie S; Karthikeyan, Subramanian; Williams, Andrew; Vincent, Renaud; Kumarathasan, Premkumari

    2017-06-01

    In this study, we used cytotoxicity assays, proteomic and gene expression analyses to examine the difference in response of A549 cells to two silica particles that differ in physical properties, namely cristobalite (CR) and α-quartz (Min-U-Sil 5, MI). Cytotoxicity assays such as lactate dehydrogenase release, 5-bromo-2'-deoxyuridine incorporation and cellular ATP showed that both silica particles could cause cell death, decreased cell proliferation and metabolism in the A549 human lung epithelial cells. While cytotoxicity assays revealed little difference between CR and MI exposures, proteomic and gene expression analyses unveiled both similar and unique molecular changes in A549 cells. For instance, two-dimensional gel electrophoresis data indicated that the expression of proteins in the cell death (e.g., ALDH1A1, HTRA2 and PRDX6) and cell proliferation (e.g., FSCN1, HNRNPAB and PGK1) pathways were significantly different between the two silica particles. Reverse transcription-polymerase chain reaction data provided additional evidence supporting the proteomic findings. Preliminary assessment of the physical differences between CR and MI suggested that the extent of surface interaction between particles and cells could explain some of the observed biological effects. However, the differential dose-response curves for some other genes and proteins suggest that other physical attributes of particulate matter can also contribute to particulate matter-related cellular toxicity. Our results demonstrated that toxicoproteomic and gene expression analyses are sensitive in distinguishing subtle toxicity differences associated with silica particles of varying physical properties compared to traditional cytotoxicity endpoints. Copyright © 2016 Her Majesty the Queen in Right of Canada. Journal of Applied Toxicology published by John Wiley & Sons, Ltd. Copyright © 2016 Her Majesty the Queen in Right of Canada. Journal of Applied Toxicology published by John Wiley & Sons

  6. ROS/Autophagy/Nrf2 Pathway Mediated Low-Dose Radiation Induced Radio-Resistance in Human Lung Adenocarcinoma A549 Cell

    PubMed Central

    Chen, Ni; Wu, Lijun; Yuan, Hang; Wang, Jun

    2015-01-01

    Low-dose ionizing radiation (LDIR) can induce radio-resistance to following high dose radiation in various mammalian cells. The protective role of LDIR has been thought to be associated with the overall outcomes of cancer radiotherapy. NF-E2 related factor 2 (Nrf2) is a transcription factor that plays pivotal roles in maintaining cellular oxidative equilibrium. Since oxidative stress has been indicated to be a mediator of LDIR induced radio-resistance, the role of Nrf2 in this process was investigated in this research. Our results showed that in human lung adenocarcinoma A549 cell, 5cGy alpha particle induced radio-resistance to following 75cGy alpha particle radiation. The expression level of Nrf2 and its target Heme Oxygenase-1(HO-1) increased after 5cGy radiation. Both the shRNA of Nrf2 and the chemical inhibitor of HO-1 suppressed the induced radio-resistance, indicating the involvement of Nrf2 antioxidant pathway in this process. Further, we found 5cGy radiation stimulated autophagy process in A549. Inhibition of the autophagy process resulted in suppression of the radio-resistance and the induced expression of Nrf2 and HO-1. ROS scavenger N-acetyl-L-cysteine (NAC) blocked the autophagy process induced by 5cGy alpha particle, the upregulation of Nrf2 and HO-1, as well as the induced radio-resistance. In conclusion, ROS elevation caused by LDIR promoted Autophagy/Nrf2-HO-1 and conferred radio-resistance in A549. PMID:26078725

  7. In vitro effects induced by diesel exhaust at an air-liquid interface in a human lung alveolar carcinoma cell line A549.

    PubMed

    Okubo, Tomoko; Hosaka, Mitsugu; Nakae, Dai

    2015-01-01

    The present study examined the effects induced in vitro in human adenocarcinoma-derived alveolar basal epithelial A549 cells by diesel particulate matter (DPM) administered into the culture medium or by diesel exhaust administered at an air-liquid interface. When A549 cells were exposed to DPM in the culture medium, cell proliferation was inhibited at doses of 10-100 μg/mL; generation of interleukin (IL)-8 and the antioxidant enzyme, heme oxygenase-1 (HO-1), were inhibited at a dose of 100 μg/mL, and hydroxyl radicals were produced, but could be inhibited by catalase or superoxide dismutase. In contrast, when A549 cells were exposed to diesel exhaust, cell proliferation was inhibited in the absence, but not in the presence, of a diesel particulate filter (DPF); in the absence of a DPF IL-8 was produced in the same amount as in the control cells but was suppressed in the presence of a DPF; HO-1 mRNA was transiently over-expressed in the presence of a DPF, and it was also increased slightly produced in the absence of a DPF but statistically not significant in the presence of a DPF, and it was also increased slightly produced in the absence of a DPF but statistically not significant; HO-1 was transiently produced independent of the absence or the presence of a DPF; and hydroxyl radicals were weakly produced, even in the presence of a DPF but could be inhibited by catalase or superoxide dismutase. It is thus suggested that oxidative stress may be induced by exposure to DPM or diesel exhaust and thereby exerts cytotoxic effect. The introduction of a DPF is effective to protect cells from the toxicity of diesel exhaust presumably by suppression of an oxidative stress. Copyright © 2015 Elsevier GmbH. All rights reserved.

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

    NASA Astrophysics Data System (ADS)

    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.

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

    PubMed

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

    2018-04-01

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

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

    PubMed

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

    2016-12-01

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

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

    PubMed Central

    Rossner, Pavel; Strapacova, Simona; Stolcpartova, Jitka; Schmuczerova, Jana; Milcova, Alena; Neca, Jiri; Vlkova, Veronika; Brzicova, Tana; Machala, Miroslav; Topinka, Jan

    2016-01-01

    We investigated the toxicity of benzo[a]pyrene (B[a]P), 1-nitropyrene (1-NP) and 3-nitrobenzanthrone (3-NBA) in A549 cells. Cells were treated for 4 h and 24 h with: B[a]P (0.1 and 1 μM), 1-NP (1 and 10 μM) and 3-NBA (0.5 and 5 μM). Bulky DNA adducts, lipid peroxidation, DNA and protein oxidation and mRNA expression of CYP1A1, CYP1B1, NQO1, POR, AKR1C2 and COX2 were analyzed. Bulky DNA adducts were induced after both treatment periods; the effect of 1-NP was weak. 3-NBA induced high levels of bulky DNA adducts even after 4-h treatment, suggesting rapid metabolic activation. Oxidative DNA damage was not affected. 1-NP caused protein oxidation and weak induction of lipid peroxidation after 4-h incubation. 3-NBA induced lipid peroxidation after 24-h treatment. Unlike B[a]P, induction of the aryl hydrocarbon receptor, measured as mRNA expression levels of CYP1A1 and CYP1B1, was low after treatment with polycyclic aromatic hydrocarbon (PAH) nitro-derivatives. All test compounds induced mRNA expression of NQO1, POR, and AKR1C2 after 24-h treatment. AKR1C2 expression indicates involvement of processes associated with reactive oxygen species generation. This was supported further by COX2 expression induced by 24-h treatment with 1-NP. In summary, 3-NBA was the most potent genotoxicant, whereas 1-NP exhibited the strongest oxidative properties. PMID:27571070

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

    PubMed

    Rossner, Pavel; Strapacova, Simona; Stolcpartova, Jitka; Schmuczerova, Jana; Milcova, Alena; Neca, Jiri; Vlkova, Veronika; Brzicova, Tana; Machala, Miroslav; Topinka, Jan

    2016-08-26

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

  13. Evaluation of anticancer activity of water and juice extracts of young Hordeum vulgare in human cancer cell lines HT-29 and A549.

    PubMed

    Czerwonka, Arkadiusz; Kawka, Katarzyna; Cykier, Klaudia; Lemieszek, Marta Kinga; Rzeski, Wojciech

    2017-06-12

    Introduction and objective. Barley (Hordeum vulgare L.) is known as a rich source of different bioactive compounds. At present, considerable attention of researchers is focused on young barley grass. It can be a good source of dietary minerals, vitamins, carbohydrates, amino acids, phenolic compounds and proteins. It is possible that the composition of chemical ingredients beneficial for health may induce an anticancer potential of young barley in human colon adenocarcinoma (HT-29) and human lung adenocarcinoma (A549) cell lines. Materials and method. Hordeum vulgare water extract (HWE) and Hordeum vulgare juice extract (HJE) were prepared. Cell proliferation and viability were examined with the use of MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and NR (neutral red) methods. Induction of necrosis was assessed by propidium iodide/Hoechst staining. Progress of the cell cycle involved in cell proliferation, apoptosis, and regulation of transcription was estimated using flow cytometry analysis. Additionally, the capability of free radical scavenging was evaluated with the DPPH assay. Results. The study revealed that extracts inhibited the proliferation of cancer cells. The NR study confirmed the low cytotoxic activity of the tested extracts to normal human colon epithelial cells (CCD 841 CoTr) and human skin fibroblasts (HSF). Furthermore, a dose-dependent cytotoxicity against HT-29 cells, but not A549 cells, has been reported. The free radical scavenging activity was observed in the case of the HWE but not the HJE. Conclusions. The obtained results indicate a cancer chemopreventive potential of young barley as a safe dietary agent in colon carcinoma.

  14. Baicalin, a Chinese Herbal Medicine, Inhibits the Proliferation and Migration of Human Non-Small Cell Lung Carcinoma (NSCLC) Cells, A549 and H1299, by Activating the SIRT1/AMPK Signaling Pathway.

    PubMed

    You, Jiawen; Cheng, Jun; Yu, Bing; Duan, Changhua; Peng, Jinghua

    2018-04-10

    BACKGROUND Baicalin is a flavonoid derived from Scutellaria baicalensis, used in Chinese herbal medicine. Activation of the sirtuin 1 gene (SIRT1) and adenosine monophosphate (AMP)-activated protein kinase gene (AMPK), the SIRT1/AMPK signaling pathway, is associated with human malignant tumors. The aim of this study was to investigate the effects of baicalin on the cell viability, apoptosis, proliferation, and migration of human non-small cell lung cancer (NSCLC) cells, A549 and H1299, in vitro. MATERIAL AND METHODS Human NSCLC cells, A549 and H1299, were treated with serial doses of baicalin. Small interfering RNA (siRNA) silencing of the SIRT1 and AMPK genes was performed using cell transfection. The MTT assay was used to determine cell viability, flow cytometry was used to measure cell apoptosis, wound healing and transwell assays were used to assess cell migration of A549 and H1299 cells. Western blotting was used to measure protein expression and phosphorylation levels in untreated A549 and H1299 cells, and cells treated with increasing doses of baicalin. RESULTS Baicalin inhibited the viability, migration, and invasion of A549 and H1299 cells, and increased cell apoptosis in a dose-dependent manner. Baicalin activated the SIRT1/AMPK and mechanistic target of rapamycin (mTOR), and SIRT1/AMPK and matrix metalloproteinase (MMP) signaling in A549 and H1299 cells in a dose-dependent manner. siRNA silencing of SIRT1 and AMPK reduced the effects of baicalin on cell proliferation and migration. CONCLUSIONS Baicalin, a flavonoid used in Chinese herbal medicine, inhibited the proliferation and migration of human NSCLC cells, A549 and H1299, by activating the SIRT1/AMPK signaling pathway.

  15. Cytotoxicity and genotoxicity in human lung epithelial A549 cells caused by airborne volatile organic compounds emitted from pine wood and oriented strand boards.

    PubMed

    Gminski, Richard; Tang, Tao; Mersch-Sundermann, Volker

    2010-06-16

    Due to the massive reduction of air-change rates in modern, energy-saving houses and dwellings, the contribution of volatile organic compound (VOCs) emissions from wood-based materials to indoor air quality has become increasingly important. To evaluate toxicity of VOC mixtures typically emitted from pine wood and oriented strand boards (OSB) and their main constituents (selected terpenes and aldehydes), cytotoxicity and genotoxicity were investigated in human A549 lung cells. To facilitate exposure directly via gas phase, a 250 L emission chamber was combined with a Vitrocell exposure system. VOC exposure concentrations were measured by GC/MSD. Biological effects were determined after an exposure time of 1h by measuring cytotoxicity (erythrosine B staining) and genotoxicity (comet assay). Neither cytotoxic nor genotoxic effects were observed for VOC mixtures emitted from pine wood or OSB at loading factors of approximately 13 m(2)/m(3) (worst case conditions) of the panels (with maximum VOC levels of about 80 mg/m(3)) in comparison to clean air. While alpha-pinene and Delta(3)-carene did not induce toxic effects even at exposure concentrations of up to 1800 mg/m(3) and 600 mg/m(3), respectively, hexanal showed a cytotoxic effect at 2000 mg/m(3). The alpha,beta-unsaturated aldehydes 2-heptenal and 2-octenal caused genotoxic effects in concentrations exceeding 100mg/m(3) and 40 mg/m(3), respectively. In conclusion, high concentrations of VOCs and VOC mixtures emitted from pine wood and OSB did not lead to adverse effects in A549 human lung cells even at concentrations 10(2) to 10(5)-fold higher than those found in normal indoor air. Attention must be paid to mutagenic and possibly carcinogenic alpha,beta-unsaturated aldehydes. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

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

    NASA Astrophysics Data System (ADS)

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

    2015-02-01

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

  17. Preclinical PK/PD model for combined administration of erlotinib and sunitinib in the treatment of A549 human NSCLC xenograft mice.

    PubMed

    Li, Jing-Yun; Ren, Yu-Peng; Yuan, Yin; Ji, Shuang-Min; Zhou, Shu-Pei; Wang, Li-Jie; Mou, Zhen-Zhen; Li, Liang; Lu, Wei; Zhou, Tian-Yan

    2016-07-01

    Combined therapy of EGFR TKI and VEGFR TKI may produce a greater therapeutic benefit and overcome EGFR TKI-induced resistance. However, a previous study shows that a combination of EGFR TKI erlotinib (ER) with VEGFR TKI sunitinib (SU) did not improve the overall survival in patients with non-small-cell lung cancer (NSCLC). In this study we examined the anticancer effect of ER, SU and their combination in the treatment of A549 human NSCLC xenograft mice, and conducted PK/PD modeling and simulations to optimize the dose regimen. ER (20, 50 mg·kg(-1)·d(-1)) or SU (5, 10, 20 mg·kg(-1)·d(-1)) alone, or their combination were administered to BALB/c nude mice bearing A549 tumors for 22 days. The tumor size and body weight were recorded daily. The experimental data were used to develop PK/PD models describing the quantitative relationship between the plasma concentrations and tumor suppression in different dose regimens. The models were further evaluated and validated, and used to predict the efficacy of different combination regimens and to select the optimal regimen. The in vivo anticancer efficacy of the combination groups was much stronger than that of either drug administered alone. A PK/PD model was developed with a combination index (φ) of 4.4, revealing a strong synergistic effect between ER and SU. The model simulation predicted the tumor growth in different dosage regimens, and showed that the dose of SU played a decisive role in the combination treatment, and suggested that a lower dose of ER (≤5 mg·kg(-1)·d(-1)) and adjusting the dose of SU might yield a better dosage regimen for clinical research. The experimental data and modeling confirm synergistic anticancer effect of ER and SU in the treatment of A549 xenograft mice. The optimal dosage regimen determined by the PK/PD modeling and simulation can be used in future preclinical study and provide a reference for clinical application.

  18. Comparative physicochemical and biological characterization of NIST Interim Reference Material PM2.5 and SRM 1648 in human A549 and mouse RAW264.7 cells.

    PubMed

    Mitkus, Robert J; Powell, Jan L; Zeisler, Rolf; Squibb, Katherine S

    2013-12-01

    The epidemiological association between exposure to fine particulate matter (PM2.5) and adverse health effects is well-known. Here we report the size distribution, metals content, endotoxin content, and biological activity of National Institute of Standards and Technology (NIST) Interim Reference Material (RM) PM2.5. Biological activity was measured in vitro by effects on cell viability and the release of four inflammatory immune mediators, from human A549 alveolar epithelial cells or murine RAW264.7 monocytes. A dose range covering three orders of magnitude (1-1000μg/mL) was tested, and biological activity was compared to an existing Standard Reference Material (SRM) for urban PM (NIST SRM 1648). Robust release of IL-8 and MCP-1 from A549 cells was observed in response to IRM PM2.5 exposures. Significant TNF-α, but not IL-6, secretion from RAW264.7 cells was observed in response to both IRM PM2.5 and SRM 1648 particle types. Cytokine or chemokine release at high doses often occurred in the presence of cytotoxicity, likely as a result of externalization of preformed mediator. Our results are consistent with a local cytotoxic and pro-inflammatory mechanism of response to exposure to inhaled ambient PM2.5 and reinforce the continued relevance of in vitro assays for mechanistic research in PM toxicology. Our study furthers the goal of developing reference samples of environmentally relevant particulate matter of various sizes that can be used for hypothesis testing by multiple investigators. Published by Elsevier Ltd.

  19. Suppression on metastasis by rhubarb through modulation on MMP-2 and uPA in human A549 lung adenocarcinoma: an ex vivo approach.

    PubMed

    Shia, Chi-Sheng; Suresh, Govindan; Hou, Yu-Chi; Lin, Yu-Chin; Chao, Pei-Dawn Lee; Juang, Shin-Hun

    2011-01-27

    The aim of this study is to determine and identify the possible molecular mechanisms of anti-cancer effect of rhubarb under the physiologically achievable concentrations by using an ex vivo approach. Rats were orally administered rhubarb decoction and then serum metabolites were extracted, prepared and characterized to assay for the following in vitro study. The MTT assay, zymography analysis, wound healing assay, RT-PCR, and Western blot analysis were used to reveal molecular events of rhubarb metabolites in this study. Experimental metastasis model was used to investigate the in vivo anti-metastatic efficacy of rhubarb. Our results demonstrated that cell line mobility was strongly inhibited and the enzymatic activity of MMP-2 decreased following culture with the rhubarb serum metabolite in human lung adenocarcinoma A549 cells. Further experiments demonstrated that the downregulation of MMP-2 enzymatic activity act through both transcriptional and post-translational mechanisms. NF-κB/c-Jun and uPA were observed involving in the inhibition of MMP-2 transcription and post-translational modification, respectively, in A549 cells treated with rhubarb serum metabolite. Further animal experiments demonstrated a significant reduction in lung metastatic colonies in rhubarb-treated mice, suggesting that rhubarb contain enriched active components that block cancer metastasis. Our studies, both in vitro and in vivo, clearly demonstrated the anti-tumor effect of rhubarb in an experimental setting of achievable physiological concentrations and also provide possible molecular mechanisms of anti-metastatic mechanisms by rhubarb treatment. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  20. Therapeutic effects of gold nanoparticles synthesized using Musa paradisiaca peel extract against multiple antibiotic resistant Enterococcus faecalis biofilms and human lung cancer cells (A549).

    PubMed

    Vijayakumar, S; Vaseeharan, B; Malaikozhundan, B; Gopi, N; Ekambaram, P; Pachaiappan, R; Velusamy, P; Murugan, K; Benelli, G; Suresh Kumar, R; Suriyanarayanamoorthy, M

    2017-01-01

    Botanical-mediated synthesis of nanomaterials is currently emerging as a cheap and eco-friendly nanotechnology, since it does not involve the use of toxic chemicals. In the present study, we focused on the synthesis of gold nanoparticles using the aqueous peel extract of Musa paradisiaca (MPPE-AuNPs) following a facile and cheap fabrication process. The green synthesized MPPE-AuNPs were bio-physically characterized by UV-Vis spectroscopy, FTIR, XRD, TEM, Zeta potential analysis and EDX. MPPE-AuNPs were crystalline in nature, spherical to triangular in shape, with particle size ranging within 50 nm. The biofilm inhibition activity of MPPE-AuNPs was higher against multiple antibiotic resistant (MARS) Gram-positive Enterococcus faecalis. Light and confocal laser scanning microscopic observations evidenced that the MPPE-AuNPs effectively inhibited the biofilm of E. faecalis when tested at 100 μg mL -1 . Cytotoxicity studies demonstrated that MPPE-AuNPs were effective in inhibiting the viability of human A549 lung cancer cells at higher concentrations of 100 μg mL -1 . The morphological changes in the MPPE-AuNPs treated A549 lung cancer cells were visualized under phase-contrast microscopy. Furthermore, the ecotoxicity of MPPE-AuNPs on the freshwater micro crustacean Ceriodaphnia cornuta were evaluated. Notably, no mortality was recorded in MPPE-AuNPs treated C. cornuta at 250 μg mL -1 . This study concludes that MPPE-AuNPs are non-toxic, eco-friendly and act as a multipurpose potential biomaterial for biomedical applications. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Induction of apoptosis and autophagy by sodium selenite in A549 human lung carcinoma cells through generation of reactive oxygen species.

    PubMed

    Park, Shin-Hyung; Kim, Jeong-Hwan; Chi, Gyoo Yong; Kim, Gi-Young; Chang, Young-Chae; Moon, Sung-Kwon; Nam, Soo-Wan; Kim, Wun-Jae; Yoo, Young Hyun; Choi, Yung Hyun

    2012-08-03

    Selenium in the form of sodium selenite has been reported to exert anti-tumor effects in several cancer cell types by inducing autophagic cell death and apoptosis mediated by reactive oxygen species (ROS). However, the exact molecular pathways underlying these effects have not been fully established. The present study used A549 human lung carcinoma cells for further investigation of the anti-cancer mechanism of sodium selenite. We showed that sodium selenite modulated both the extrinsic and intrinsic apoptotic pathways, which were interconnected by Bid truncation. We used z-VAD-fmk, a pan-caspase inhibitor, to demonstrate that sodium selenite-induced apoptosis was dependent on the activation of caspases. Sodium selenite also increased autophagy, as indicated by an increase in microtubule-associated protein light chain-3 (LC3) puncta, accumulation of LC3II, and elevation of autophagic flux. Pretreatment with bafilomycin A1 enhanced sodium selenite-induced apoptosis, indicating that sodium selenite-induced autophagy functioned as a survival mechanism. Sodium selenite treatment also resulted in generation of ROS, which abrogated mitochondrial membrane potential (MMP) and regulated both apoptosis and autophagy. Phospho-nuclear factor erythroid 2-related factor 2 (p-Nrf2) showed a ROS-dependent translocation to the nucleus, which suggested that Nrf2 might increase cell survival by suppressing ROS accumulation and apoptosis mediated by oxidative stress. Sodium selenite treatment of A549 cells therefore appeared to trigger both apoptosis and cytoprotective autophagy, which were both mediated by ROS. The data suggest that regulation of ROS generation and autophagy can be a potential strategy for treating lung cancer that is resistant to pro-apoptotic therapeutics. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

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

    PubMed

    Hevel, Joan M; Olson-Buelow, Laura C; Ganesan, Balasubramanian; Stevens, John R; Hardman, Jared P; Aust, Ann E

    2008-08-07

    Although exposure to asbestos is now regulated, patients continue to be diagnosed with mesothelioma, asbestosis, fibrosis and lung carcinoma because of the long latent period between exposure and clinical disease. Asbestosis is observed in approximately 200,000 patients annually and asbestos-related deaths are estimated at 4,000 annually. Although advances have been made using single gene/gene product or pathway studies, the complexity of the response to asbestos and the many unanswered questions suggested the need for a systems biology approach. The objective of this study was to generate a comprehensive view of the transcriptional changes induced by crocidolite asbestos in A549 human lung epithelial cells. A statistically robust, comprehensive data set documenting the crocidolite-induced changes in the A549 transcriptome was collected. A systems biology approach involving global observations from gene ontological analyses coupled with functional network analyses was used to explore the effects of crocidolite in the context of known molecular interactions. The analyses uniquely document a transcriptome with function-based networks in cell death, cancer, cell cycle, cellular growth, proliferation, and gene expression. These functional modules show signs of a complex interplay between signaling pathways consisting of both novel and previously described asbestos-related genes/gene products. These networks allowed for the identification of novel, putative crocidolite-related genes, leading to several new hypotheses regarding genes that are important for the asbestos response. The global analysis revealed a transcriptome that bears signatures of both apoptosis/cell death and cell survival/proliferation. Our analyses demonstrate the power of combining a statistically robust, comprehensive dataset and a functional network genomics approach to 1) identify and explore relationships between genes of known importance 2) identify novel candidate genes, and 3) observe the

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

    SciTech Connect

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

    2006-01-15

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

  4. Physicochemical characterization of ambient PM2.5in Tehran air and its potential cytotoxicity in human lung epithelial cells (A549).

    PubMed

    MohseniBandpi, Anoushiravan; Eslami, Akbar; Shahsavani, Abbas; Khodagholi, Fariba; Alinejad, Abdolazim

    2017-09-01

    As air pollution is a major problem in Tehran, this study aimed to investigate the physicochemical characterization of the water-soluble and organic contents of ambient PM 2.5 in Tehran and determine its in vitro toxicological impact on human lung epithelial cells (A549). A total of 11 sampling stations were selected, including three categories: traffic, urban, and suburban. All sampling was carried out in the spring and summer of 2015. Ion chromatography (IC), inductively coupled plasma atomic emission spectroscopy (ICP-AES), and GC-MS were used to analyze ionic compounds, heavy metals, and polycyclic aromatic hydrocarbons (PAHs), respectively, and an ELISA reader was used for cytotoxicity analysis. The most prevalent ionic species found for all three categories was SO 4 2- . PAH concentrations were 43.45±32.71, 50.51±37.27, and 29.13±33.29ng/m 3 for traffic, urban, and suburban stations, respectively. For all sampling stations, Al and Fe had the highest values among the investigated heavy metals. Cell viability measurements, carried out using the MTT assay, showed that all three categories of samples cause cytotoxicity, although the urban station samples showed higher cytotoxicity than those from the other stations (p˂0.05). Based on the results of the present study, organic compounds and insoluble particles could be the main causes of cytotoxicity. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    PubMed

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

    2017-07-14

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

  6. Fabrication of nano-silver particles using Cymodocea serrulata and its cytotoxicity effect against human lung cancer A549 cells line

    NASA Astrophysics Data System (ADS)

    Palaniappan, P.; Sathishkumar, G.; Sankar, R.

    2015-03-01

    The present study reports, green synthesis of bioactive silver nanoparticles (AgNPs) under different temperature (60 °C, room temperature and 4° refrigerator) using the aqueous extract of sea grass Cymodocea serrulata as a potential bioreductant. Increased temperature fabricates more AgNPs compare to room temperature and refrigerator condition. At first the reduction of Ag+ ions were confirmed through color change which produces an absorbance spectra at 420 nm in UV-Visible spectrophotometer. Additionally various exclusive instrumentations such as X-ray diffraction (XRD), Dynamic light scattering (DLS), scanning electron microscope (SEM) analysis and Transmission electron microscope (TEM) were authorizes the biosynthesis and physio-chemical characterization of AgNPs. From Fourier transform infrared spectroscopy (FTIR) analysis, it was identified that the water soluble fractions of the sea grass mainly responsible for reduction of ionic silver (Ag+) into (Ag0) nano-ranged particles and also they act as stabilizing agent to sustain the durability of NPs for long period of time. Further, synthesized AgNPs shows potential cytotoxicity against human lung cancer A549 cells (LD50-100 μg/ml). The overall results suggest that C. serrulata is a valuable bioresource to generate rapid and eco-friendly bioactive AgNPs towards cancer therapy.

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

    PubMed

    Liu, Betty R; Huang, Yue-Wern; Aronstam, Robert S; Lee, Han-Jung

    2016-01-01

    Cell-penetrating peptides (CPPs) have been shown to deliver cargos, including protein, DNA, RNA, and nanomaterials, in fully active forms into live cells. Most of the CPP sequences in use today are based on non-native proteins that may be immunogenic. Here we demonstrate that the L5a CPP (RRWQW) from bovine lactoferricin (LFcin), stably and noncovalently complexed with plasmid DNA and prepared at an optimal nitrogen/phosphate ratio of 12, is able to efficiently enter into human lung cancer A549 cells. The L5a CPP delivered a plasmid containing the enhanced green fluorescent protein (EGFP) coding sequence that was subsequently expressed in cells, as revealed by real-time PCR and fluorescent microscopy at the mRNA and protein levels, respectively. Treatment with calcium chloride increased the level of gene expression, without affecting CPP-mediated transfection efficiency. Zeta-potential analysis revealed that positively electrostatic interactions of CPP/DNA complexes correlated with CPP-mediated transport. The L5a and L5a/DNA complexes were not cytotoxic. This biomimetic LFcin L5a represents one of the shortest effective CPPs and could be a promising lead peptide with less immunogenic for DNA delivery in gene therapy.

  8. The environmental carcinogen 3-nitrobenzanthrone and its main metabolite 3-aminobenzanthrone enhance formation of reactive oxygen intermediates in human A549 lung epithelial cells

    SciTech Connect

    Hansen, Tanja; Seidel, Albrecht; Borlak, Juergen

    2007-06-01

    The environmental contaminant 3-nitrobenzanthrone (3-NBA) is highly mutagenic and a suspected human carcinogen. We aimed to evaluate whether 3-NBA is able to deregulate critical steps in cell cycle control and apoptosis in human lung epithelial A549 cells. Increased intracellular Ca{sup 2+} and caspase activities were detected upon 3-NBA exposure. As shown by cell cycle analysis, an increased number of S-phase cells was observed after 24 h of treatment with 3-NBA. Furthermore, 3-NBA was shown to inhibit cell proliferation when added to subconfluent cell cultures. The main metabolite of 3-NBA, 3-ABA, induced statistically significant increases in tail moment as judged bymore » alkaline comet assay. The potential of 3-NBA and 3-ABA to enhance the production of reactive oxygen species (ROS) was demonstrated by flow cytometry using 2',7'-dichlorofluorescein-diacetate (DCFH-DA). The enzyme inhibitors allopurinol, dicumarol, resveratrol and SKF525A were used to assess the impact of metabolic conversion on 3-NBA-mediated ROS production. Resveratrol decreased dichlorofluorescein (DCF) fluorescence by 50%, suggesting a role for CYP1A1 in 3-NBA-mediated ROS production. Mitochondrial ROS production was significantly attenuated (20% reduction) by addition of rotenone (complex I inhibition) and thenoyltrifluoroacetone (TTFA, complex II inhibition). Taken together, the results of the present study provide evidence for a genotoxic potential of 3-ABA in human epithelial lung cells. Moreover, both compounds lead to increased intracellular ROS and create an environment favorable to DNA damage and the promotion of cancer.« less

  9. The environmental carcinogen 3-nitrobenzanthrone and its main metabolite 3-aminobenzanthrone enhance formation of reactive oxygen intermediates in human A549 lung epithelial cells.

    PubMed

    Hansen, Tanja; Seidel, Albrecht; Borlak, Jürgen

    2007-06-01

    The environmental contaminant 3-nitrobenzanthrone (3-NBA) is highly mutagenic and a suspected human carcinogen. We aimed to evaluate whether 3-NBA is able to deregulate critical steps in cell cycle control and apoptosis in human lung epithelial A549 cells. Increased intracellular Ca(2+) and caspase activities were detected upon 3-NBA exposure. As shown by cell cycle analysis, an increased number of S-phase cells was observed after 24 h of treatment with 3-NBA. Furthermore, 3-NBA was shown to inhibit cell proliferation when added to subconfluent cell cultures. The main metabolite of 3-NBA, 3-ABA, induced statistically significant increases in tail moment as judged by alkaline comet assay. The potential of 3-NBA and 3-ABA to enhance the production of reactive oxygen species (ROS) was demonstrated by flow cytometry using 2',7'-dichlorofluorescein-diacetate (DCFH-DA). The enzyme inhibitors allopurinol, dicumarol, resveratrol and SKF525A were used to assess the impact of metabolic conversion on 3-NBA-mediated ROS production. Resveratrol decreased dichlorofluorescein (DCF) fluorescence by 50%, suggesting a role for CYP1A1 in 3-NBA-mediated ROS production. Mitochondrial ROS production was significantly attenuated (20% reduction) by addition of rotenone (complex I inhibition) and thenoyltrifluoroacetone (TTFA, complex II inhibition). Taken together, the results of the present study provide evidence for a genotoxic potential of 3-ABA in human epithelial lung cells. Moreover, both compounds lead to increased intracellular ROS and create an environment favorable to DNA damage and the promotion of cancer.

  10. Genotoxic effects of three selected black toner powders and their dimethyl sulfoxide extracts in cultured human epithelial A549 lung cells in vitro.

    PubMed

    Gminski, Richard; Decker, Katharina; Heinz, Christina; Seidel, Albrecht; Könczöl, Mathias; Goldenberg, Ella; Grobéty, Bernard; Ebner, Winfried; Gieré, Reto; Mersch-Sundermann, Volker

    2011-05-01

    Until now, the adverse effects of toner powders on humans have been considered to be minimal. However, several recent reports have suggested possible significant adverse health effects from toner dust inhalation. The aim of this study was to evaluate the genotoxic potential of black toner powders in vitro. For the study of DNA damage, A549 cells were exposed to toner-powder suspensions and to their DMSO extracts, and then subjected to the comet assay and to the in-vitro cytokinesis block micronucleus test (CB-MNvit). Cytotoxic effects of the toner samples were assessed by the erythrosin B assay. Furthermore, size, shape, and composition of the toner powders were investigated. None of the three toner powders or their DMSO extracts reduced cell viability; however, they did induce DNA damage and formed micronuclei at concentrations from 80 to 400 μg cm(-2) , although to a varying extent. All toner powders contain considerable amounts of the pigments carbon black and magnetite (Fe(3) O(4) ) as well as small amounts of polycyclic aromatic hydrocarbons (PAHs). The overall results of our in-vitro study suggest that the investigated toner-powder samples are not cytotoxic but genotoxic. From the results of the physical and chemical characterization, we conclude that metals and metalloids as components of magnetite, or PAHs as components of the carbon-bearing material, are responsible for the genotoxic effects. Further research is necessary to determine the relevance of these in-vitro observations for private and occupational toner powder exposure. Copyright © 2010 Wiley-Liss, Inc.

  11. Investigations on cytotoxic and genotoxic effects of laser printer emissions in human epithelial A549 lung cells using an air/liquid exposure system.

    PubMed

    Tang, Tao; Gminski, Richard; Könczöl, Mathias; Modest, Christoph; Armbruster, Benedikt; Mersch-Sundermann, Volker

    2012-03-01

    Exposure to emissions from laser printers during the printing process is commonplace worldwide, both in the home and workplace environment. In the present study, cytotoxic and genotoxic effects of the emission from five low to medium-throughput laser printers were investigated with respect to the release of ozone (O(3) ), volatile organic compounds (VOC), particulate matter (PM), and submicrometer particles (SMP) during standby and operation. Experiments were conducted in a 1 m(3) emission chamber connected to a Vitrocell® exposure system. Cytotoxicity was determined by the WST-1 assay and genotoxicity by the micronucleus test in human A549 lung cells. The five laser printers emitted varying but generally small amounts of O(3) , VOC, and PM. VOC emissions included 13 compounds with total VOC concentrations ranging from 95 to 280 μg/m(3) (e.g., 2-butanone, hexanal, m,p-xylene, and o-xylene). Mean PM concentrations were below 2.4 μg/m(3). SMP number concentration levels during standby ranged from 9 to 26 particles/cm(3). However, three of the printers generated a 90 to 16 × 10(3) -fold increase of SMP during the printing process (maximum 294,460 particles/cm(3)). Whereas none of the printer emissions were found to cause cytotoxicity, emissions from two printers induced formation of micronuclei (P < 0.001), thus providing evidence for genotoxicity. As yet, differences in biological activity cannot be explained on the basis of the specific emission characteristics of the different printers. Because laser printing technology is widely used, studies with additional cytogenetic endpoints are necessary to confirm the DNA-damaging potency and to identify emission components responsible for genotoxicity. Copyright © 2011 Wiley-Liss, Inc.

  12. LW6, a hypoxia-inducible factor 1 inhibitor, selectively induces apoptosis in hypoxic cells through depolarization of mitochondria in A549 human lung cancer cells.

    PubMed

    Sato, Mariko; Hirose, Katsumi; Kashiwakura, Ikuo; Aoki, Masahiko; Kawaguchi, Hideo; Hatayama, Yoshiomi; Akimoto, Hiroyoshi; Narita, Yuichiro; Takai, Yoshihiro

    2015-09-01

    Hypoxia‑inducible factor 1 (HIF‑1) activates the transcription of genes that act upon the adaptation of cancer cells to hypoxia. LW6, an HIF‑1 inhibitor, was hypothesized to improve resistance to cancer therapy in hypoxic tumors by inhibiting the accumulation of HIF‑1α. A clear anti‑tumor effect under low oxygen conditions would indicate that LW6 may be an improved treatment strategy for cancer in hypoxia. In the present study, the HIF‑1 inhibition potential of LW6 on the growth and apoptosis of A549 lung cancer cells in association with oxygen availability was evaluated. LW6 was observed to inhibit the expression of HIF‑1α induced by hypoxia in A549 cells at 20 mM, independently of the von Hippel‑Lindau protein. In addition, at this concentration, LW6 induced hypoxia‑selective apoptosis together with a reduction in the mitochondrial membrane potential. The intracellular reactive oxygen species levels increased in LW6‑treated hypoxic A549 cells and LW6 induced a hypoxia‑selective increase of mitochondrial O2•‑. In conclusion, LW6 inhibited the growth of hypoxic A549 cells by affecting the mitochondria. The inhibition of the mitochondrial respiratory chain is suggested as a potentially effective strategy to target apoptosis in cancer cells.

  13. Radiosensitizing effect of schinifoline from Zanthoxylum schinifolium Sieb et Zucc on human non-small cell lung cancer A549 cells: a preliminary in vitro investigation.

    PubMed

    Wang, Cheng-Fang; Fan, Li; Tian, Mei; Qi, Xue-Song; Liu, Jian-Xiang; Feng, Jiang-Bin; Du, Shu-Shan; Su, Xu; Wang, Yong-Yan

    2014-12-01

    Schinifoline (SF), a 4-quinolinone derivative, was found in Zanthoxylum schinifolium for the first time. 4-Quinolinone moieties are thought to have cytotoxic activity and are often used as a tubulin polymerization inhibitors, heterogeneous enzyme inhibitors and antiplatelet agents. However, very little information respect to radiosensitization has focused on SF. This work aimed to investigate the radiosensitizing effect of SF on A549 cells. The cell viability results indicated cytotoxicity of SF on A549 cells, with IC50 values of 33.7 ± 2.4, 21.9 ± 1.9 and 16.8 ± 2.2 μg/mL, respectively, after 6, 12, 24 h treatment with different concentrations, and the 10% or 20% IC50 concentration during 12 h was applied in later experiments. The results of cell proliferative inhibition and clonogenic assay showed that SF enhanced the radiosensitivity of A549 cells when applied before 60Co γ-irradiation and this effect was mainly time and concentration dependent. The flow cytometric data indicated that SF treatment before the irradiation increased the G2/M phase, thus improving the radiosensitivity of A549, leading to cell apoptosis. This paper is the first study that describes the in vitro radiosensitising, cell cycle and apoptotic-inducing effects of schinifoline.

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

    PubMed

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

    2014-12-05

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

  15. Comparison of wood smoke PM2.5 obtained from the combustion of FIR and beech pellets on inflammation and DNA damage in A549 and THP-1 human cell lines.

    PubMed

    Corsini, Emanuela; Budello, Silvia; Marabini, Laura; Galbiati, Valentina; Piazzalunga, Andrea; Barbieri, Pierluigi; Cozzutto, Sergio; Marinovich, Marina; Pitea, Demetrio; Galli, Corrado L

    2013-12-01

    The aim of this study was to investigate the effect on the induction of interleukin-8 of particulate matter (PM) from fir and beech pellets burnt in domestic appliances on two human cells lines, namely the lung epithelial cell line A549 and the promyelocytic cell line THP-1. The effects of PM2.5 obtained from combustion of beech and fir pellets were compared to reference diesel exhaust particulates (DEP). In parallel, wood smoke PM-induced genotoxicity and oxidative stress were also investigated in A549 cells. Cells were treated for different times (3-72 h) with increasing concentrations of PM2.5 obtained from sequential combustions of fir and beech pellets or reference DEP. Cell viability was assessed by lactate dehydrogenase leakage, and the release of interleukin-8 or CXCL8 (IL-8) was measured to evaluate the pro-inflammatory effect. Oxidative stress was evaluated by the 5(6)-carboxy-2',7'dichlorofluorescein diacetate (DCFH-DA) assay and DNA damage by the alkaline comet assay and micronucleus frequency by flow cytometry. Both A549 and THP-1 cells responded in a dose- and time-related manner to wood smoke PM2.5 with IL-8 release, particles obtained from late combustions being the most active. THP-1 cells were more sensitive than A549 cells. On a mass base, similar effects were observed for both fir and beech PM2.5. However, the combustion of beech pellets generated approximately three times more PM2.5 than fir pellets. Regarding the mechanism of PM2.5 uptake, in both THP-1 and A549 cells, cytochalasin D prevented PM2.5-induced IL-8 mRNA expression and cytokine release, indicating a key role for actin polymerization in particles uptake and that the production of IL-8 correlated with particle phagocytosis. As signal transduction pathway involvement, in both THP-1 and A549 cells, PM2.5-induced IL-8 release could be completely blocked by the selective inhibitor SB203580, indicating a role of p38 MAPK activation. PM2.5 from both fir and beech pellets also induced

  16. Application of a lipid-coated hollow calcium phosphate nanoparticle in synergistic co-delivery of doxorubicin and paclitaxel for the treatment of human lung cancer A549 cells.

    PubMed

    Wu, Chao; Xu, Jie; Hao, Yanna; Zhao, Ying; Qiu, Yang; Jiang, Jie; Yu, Tong; Ji, Peng; Liu, Ying

    2017-01-01

    In this study, we developed a lipid-coated hollow calcium phosphate (LCP) nanoparticle for the combined application of two chemotherapeutic drugs to human lung cancer A549 cells. Hydrophilic doxorubicin (DOX) was incorporated into the hollow structure of hollow calcium phosphate (HCP), and a lipid bilayer containing hydrophobic paclitaxel (PTX) was subsequently coated on the surface of HCP. The study on combinational effects demonstrated that the combination of DOX and PTX at a mass ratio of 12:1 showed a synergistic effect against A549 cells. The particle size, zeta potential, and encapsulation efficiency were measured to obtain optimal values: particle size was 335.0 3.2 nm, zeta potential -41.1 mV, and encapsulation efficiency 80.40%±2.24%. An in vitro release study indicated that LCP produced a sustained drug release. A549 cells had a better uptake of LCP with good biocompatibility. Furthermore, in vitro cytotoxicity experiment, apoptosis analysis, in vivo anti-tumor efficacy and protein expression analysis of Bax, Bcl-2, and Caspase-3 demonstrated that the co-delivery system based on LCP had significant synergistic anti-tumor activity. All conclusions suggested that LCP is a promising platform for co-delivery of multiple anti-tumor drugs.

  17. Application of a lipid-coated hollow calcium phosphate nanoparticle in synergistic co-delivery of doxorubicin and paclitaxel for the treatment of human lung cancer A549 cells

    PubMed Central

    Wu, Chao; Xu, Jie; Hao, Yanna; Zhao, Ying; Qiu, Yang; Jiang, Jie; Yu, Tong; Ji, Peng; Liu, Ying

    2017-01-01

    In this study, we developed a lipid-coated hollow calcium phosphate (LCP) nanoparticle for the combined application of two chemotherapeutic drugs to human lung cancer A549 cells. Hydrophilic doxorubicin (DOX) was incorporated into the hollow structure of hollow calcium phosphate (HCP), and a lipid bilayer containing hydrophobic paclitaxel (PTX) was subsequently coated on the surface of HCP. The study on combinational effects demonstrated that the combination of DOX and PTX at a mass ratio of 12:1 showed a synergistic effect against A549 cells. The particle size, zeta potential, and encapsulation efficiency were measured to obtain optimal values: particle size was 335.0 3.2 nm, zeta potential −41.1 mV, and encapsulation efficiency 80.40%±2.24%. An in vitro release study indicated that LCP produced a sustained drug release. A549 cells had a better uptake of LCP with good biocompatibility. Furthermore, in vitro cytotoxicity experiment, apoptosis analysis, in vivo anti-tumor efficacy and protein expression analysis of Bax, Bcl-2, and Caspase-3 demonstrated that the co-delivery system based on LCP had significant synergistic anti-tumor activity. All conclusions suggested that LCP is a promising platform for co-delivery of multiple anti-tumor drugs. PMID:29184399

  18. DNA damage and endoplasmic reticulum stress mediated curcumin-induced cell cycle arrest and apoptosis in human lung carcinoma A-549 cells through the activation caspases cascade- and mitochondrial-dependent pathway.

    PubMed

    Lin, Song-Shei; Huang, Hsuan-Pang; Yang, Jai-Sing; Wu, Jeng-Yuan; Hsia, Te-Chun; Hsai, Te-Chun; Lin, Chin-Chung; Lin, Cheng-Wen; Kuo, Chao-Lin; Gibson Wood, W; Chung, Jing-Gung

    2008-12-08

    Curcumin, a major component of the Curcuma species, is known to have antioxidant, anti-inflammatory properties and induce apoptosis of cancer cells, however, the precise molecular mechanisms of apoptosis in vitro are unclear. In this study, we showed that curcumin, a plant product containing the phenolic phytochemical, caused DNA damage and endoplasmic reticulum (ER) stress and mitochondrial-dependent-induced apoptosis through the activation of caspase-3 at a treatment concentration of 30 microM in human lung cancer A-549 cells. In contrast, treatment with 5-10 microM of curcumin did not induce significant apoptosis, but rather induced G2/M-phase arrest in A-549 cells. Flow cytometric analysis indicated that curcumin directly increased intracellular oxidative stress based on the cell permeable dye, 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) acting as an indicator of reactive oxygen species (ROS) generation. GADD153 and GRP78 were increased by curcumin which was indicative of ER stress. Curcumin increased Ca(2+) levels and the mitochondrial membrane potential (DeltaPsi(m)), was decreased in A-549 cells. Overall, our results demonstrated that curcumin treatment causes cell death by activating pathways inducing G2/M-phase arrest and apoptosis.

  19. K9(C4H4FN2O2)2Nd(PW11O39)2·25H2O induces apoptosis in human lung cancer A549 cells

    PubMed Central

    Xia, Rong-Yao; Zhang, Ran-Ran; Jiang, Zhe; Sun, Ya-Jiao; Liu, Jing; Chen, Fu-Hui

    2017-01-01

    Lung cancer is the leading cause of cancer-associated mortality worldwide. The present study investigated the effects of K9(C4H4FN2O2)2Nd(PW11O39)2·25H2O (FNdPW), a chemically synthesized polyoxometalate that contains rare earth elements, on lung cancer growth, and explored the mechanism underlying its actions. The effects of FNdPW on the cell viability and apoptosis of human lung cancer A549 cells were measured using MTT assay, acridine orange/ethidium bromide staining and electron microscopy. The expression of apoptosis-related proteins, including B-cell lymphoma (Bcl)-2-associated death promoter (Bad), phosphorylated (p)-Bad, X-linked inhibitor of apoptosis (XIAP), apoptosis-inducing factor (AIF), Bcl-2-associated X protein (Bax) and Bcl-2, was determined by western blotting. Caspase-3 activity was measured using a caspase-3 activity kit. After 72 h of incubation, FNdPW reduced cell viability and induced apoptosis in A549 cells in a concentration- and time-dependent manner. FNdPW upregulated the pro-apoptotic Bad and Bax proteins, and downregulated the anti-apoptotic p-Bad, Bcl-2 and XIAP proteins. Furthermore, FNdPW also enhanced caspase-3 activity and increased the protein level of AIF in A549 cells, which was independent of the caspase-3 pathway. These events were associated with the regulation exerted by FNdPW on multiple targets involved in A549 cell proliferation. Therefore, FNdPW may be a novel drug for the treatment of lung cancer. PMID:28454260

  20. K9(C4H4FN2O2)2Nd(PW11O39)2·25H2O induces apoptosis in human lung cancer A549 cells.

    PubMed

    Xia, Rong-Yao; Zhang, Ran-Ran; Jiang, Zhe; Sun, Ya-Jiao; Liu, Jing; Chen, Fu-Hui

    2017-03-01

    Lung cancer is the leading cause of cancer-associated mortality worldwide. The present study investigated the effects of K 9 (C 4 H 4 FN 2 O 2 ) 2 Nd(PW 11 O 39 ) 2 ·25H 2 O (FNdPW), a chemically synthesized polyoxometalate that contains rare earth elements, on lung cancer growth, and explored the mechanism underlying its actions. The effects of FNdPW on the cell viability and apoptosis of human lung cancer A549 cells were measured using MTT assay, acridine orange/ethidium bromide staining and electron microscopy. The expression of apoptosis-related proteins, including B-cell lymphoma (Bcl)-2-associated death promoter (Bad), phosphorylated (p)-Bad, X-linked inhibitor of apoptosis (XIAP), apoptosis-inducing factor (AIF), Bcl-2-associated X protein (Bax) and Bcl-2, was determined by western blotting. Caspase-3 activity was measured using a caspase-3 activity kit. After 72 h of incubation, FNdPW reduced cell viability and induced apoptosis in A549 cells in a concentration- and time-dependent manner. FNdPW upregulated the pro-apoptotic Bad and Bax proteins, and downregulated the anti-apoptotic p-Bad, Bcl-2 and XIAP proteins. Furthermore, FNdPW also enhanced caspase-3 activity and increased the protein level of AIF in A549 cells, which was independent of the caspase-3 pathway. These events were associated with the regulation exerted by FNdPW on multiple targets involved in A549 cell proliferation. Therefore, FNdPW may be a novel drug for the treatment of lung cancer.

  1. Drug Transporter Protein Quantification of Immortalized Human Lung Cell Lines Derived from Tracheobronchial Epithelial Cells (Calu-3 and BEAS2-B), Bronchiolar-Alveolar Cells (NCI-H292 and NCI-H441), and Alveolar Type II-like Cells (A549) by Liquid Chromatography-Tandem Mass Spectrometry.

    PubMed

    Sakamoto, Atsushi; Matsumaru, Takehisa; Yamamura, Norio; Suzuki, Shinobu; Uchida, Yasuo; Tachikawa, Masanori; Terasaki, Tetsuya

    2015-09-01

    Understanding the mechanisms of drug transport in the human lung is an important issue in pulmonary drug discovery and development. For this purpose, there is an increasing interest in immortalized lung cell lines as alternatives to primary cultured lung cells. We recently reported the protein expression in human lung tissues and pulmonary epithelial cells in primary culture, (Sakamoto A, Matsumaru T, Yamamura N, Uchida Y, Tachikawa M, Ohtsuki S, Terasaki T. 2013. J Pharm Sci 102(9):3395-3406) whereas comprehensive quantification of protein expressions in immortalized lung cell lines is sparse. Therefore, the aim of the present study was to clarify the drug transporter protein expression of five commercially available immortalized lung cell lines derived from tracheobronchial cells (Calu-3 and BEAS2-B), bronchiolar-alveolar cells (NCI-H292 and NCI-H441), and alveolar type II cells (A549), by liquid chromatography-tandem mass spectrometry-based approaches. Among transporters detected, breast cancer-resistance protein in Calu-3, NCI-H292, NCI-H441, and A549 and OCTN2 in BEAS2-B showed the highest protein expression. Compared with data from our previous study,(Sakamoto A, Matsumaru T, Yamamura N, Uchida Y, Tachikawa M, Ohtsuki S, Terasaki T. 2013. J Pharm Sci 102(9):3395-3406) NCI-H441 was the most similar with primary lung cells from all regions in terms of protein expression of organic cation/carnitine transporter 1 (OCTN1). In conclusion, the protein expression profiles of transporters in five immortalized lung cell lines were determined, and these findings may contribute to a better understanding of drug transport in immortalized lung cell lines. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

  2. Anti-metastatic effects of isolinderalactone via the inhibition of MMP-2 and up regulation of NM23-H1 expression in human lung cancer A549 cells

    PubMed Central

    Chuang, Cheng-Hung; Wang, Li-Yu; Wong, Yuen Man; Lin, En-Shyh

    2018-01-01

    Metastatic lung cancer is a leading cause of mortality and has a mortality rate of ≥90%. Isolinderalactone (ILL) is a sesquiterpene lactone compound that has been used in traditional Chinese medicine. Research has demonstrated that ILL has anti-inflammatory and anti-proliferative properties; however, to the best of our knowledge, studies investigating whether ILL can inhibit lung cancer cell metastasis have not been conducted. In the present study, 1–10 µM ILL was applied in the culturing of the A549 lung cancer cell line to investigate the effects of ILL on the invasion and migration of lung cancer cells, including whether the possible mechanisms of ILL are associated with the expression of matrix metalloproteinase (MMP)-2 and NME/NM23 nucleoside diphosphate kinase 1 (NM23-H1) genes. The results of the present study indicated that ILL inhibited the invasion and migration of the A549 cancer cells and exhibited a dose-response association. ILL also significantly inhibited the protein expression and activity of MMP-2 (P<0.05), exhibiting a trend similar to that of its invasion- and migration-associated properties. Further research revealed that ILL significantly increased the expression of NM23-H1 protein and inhibited the expression of β-catenin protein (P<0.05). The results of the present study is, to the best of our knowledge, the first to confirm that ILL can inhibit the invasion and migration of A549 cancer cells, with the possible mechanisms potentially involving the inhibition of MMP-2 and β-catenin protein expression resulting from the up regulation of NM23-H1 expression. PMID:29541242

  3. Andrographolide down-regulates hypoxia-inducible factor-1{alpha} in human non-small cell lung cancer A549 cells

    SciTech Connect

    Lin, Hui-Hsuan; Department of Medical Research, Chung Shan Medical University Hospital, Taichung, Taiwan; Tsai, Chia-Wen

    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-1{alpha} (HIF-1{alpha}) in A549 cells. HIF-1{alpha} plays an important role in tumor growth, angiogenesis and lymph node metastasis of NSCLC. The Andro-induced decrease of cellular protein level of HIF-1{alpha} was correlated with a rapid ubiquitin-dependent degradation of HIF-1{alpha}, and was accompanied bymore » increased expressions of hydroxyl-HIF-1{alpha} 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-1{alpha} inactivation, because its DNA binding activity was suppressed by Andro. Molecular data showed that all these effects of Andro might be mediated via TGF{beta}1/PHD2/HIF-1{alpha} pathway, as demonstrated by the transfection of TGF{beta}1 overexpression vector and PHD2 siRNA, and the usage of a pharmacological MG132 inhibitor. Furthermore, we elucidated the involvement of Andro in HIF-1{alpha} 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.« less

  4. Microcystin-LR induces a wide variety of biochemical changes in the A549 human non-small cell lung cancer cell line: Roles for protein phosphatase 2A and its substrates.

    PubMed

    Wang, Hanying; Xu, Kailun; Wang, Beilei; Liu, Jinghui; Wang, Xiaofeng; Xing, Mingluan; Huang, Pu; Guo, Zonglou; Xu, Lihong

    2017-03-01

    Our previous studies have described the toxic effects of microcystin-LR (MC-LR) in various normal cell lines and human hepatoma SMMC-7721 cells, but the specific effects of MC-LR in other types of cancer cells with respect to protein phosphatase 2A (PP2A) have not been fully elaborated. A549 human lung adenocarcinoma cells have been identified to express organic anion-transporting polypeptides (OATP) involved in cellular uptake of MC-LR, and thus probably make an appropriate in vitro model to assess MC-LR's cytotoxicity. Hence, in our present study, A549 cells were treated with various concentrations of MC-LR for 24 h. The presence of MC-LR in A549 cells was confirmed, and PP2A activity, PP2A substrates, cytoskeleton, apoptosis, and proliferation were subsequently explored. The results showed that 5-10 μM MC-LR inhibited PP2A activity significantly but 0.5-1 μM MC-LR did not change PP2A activity dramatically. The inhibition could result from the hyperphosphorylation of PP2A/C at Tyr307, an elevation in the total PP2A/C expression and the dissociation of α4/PP2A/C complexes. Moreover, MC-LR led to rearrangements of filamentous actin and microtubules, which might be correlated with the hyperphosphorylation of Ezrin, VASP and HSP27 due to PP2A inhibition and mitogen-activated protein kinase (MAPK) activation. However, exposure to MC-LR for 24 h failed to trigger either apoptosis or proliferation, which might be related to PP2A-inhibition-induced hyperphosphorylation of Bcl-2 and Bad and the activation status of Akt. In conclusion, our data indicated that MC-LR induced extensive molecular and cellular alterations in A549 cells through a PP2A-centered pathway, which differed in some respects from our previous study in SMMC-7721 cells. To our knowledge, this is the first report comprehensively demonstrating the effects of MC-LR in A549 cells, and our findings provide insights into the mechanism of MC-LR toxicity in cancer cells. © 2016 Wiley Periodicals, Inc. Environ

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

    PubMed

    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.

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

    SciTech Connect

    Yu, Zhenhai, E-mail: tomsyu@163.com; Huang, Liangqian; Qiao, Pengyun

    2016-05-13

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

  7. Vitamin A (retinol) downregulates the receptor for advanced glycation endproducts (RAGE) by oxidant-dependent activation of p38 MAPK and NF-kB in human lung cancer A549 cells.

    PubMed

    de Bittencourt Pasquali, Matheus Augusto; Gelain, Daniel Pens; Zeidán-Chuliá, Fares; Pires, André Simões; Gasparotto, Juciano; Terra, Silvia Resende; Moreira, José Cláudio Fonseca

    2013-04-01

    As an essential component of the diet, retinol supplementation is often considered harmless and its application is poorly controlled. However, recent works demonstrated that retinol may induce a wide array of deleterious effects, especially when doses used are elevated. Controlled clinical trials have demonstrated that retinol supplementation increased the incidence of lung cancer and mortality in smokers. Experimental works in cell cultures and animal models showed that retinol may induce free radical production, oxidative stress and extensive biomolecular damage. Here, we evaluated the effect of retinol on the regulation of the receptor for advanced glycation end-products (RAGE) in the human lung cancer cell line A549. RAGE is constitutively expressed in lungs and was observed to be down-regulated in lung cancer patients. A549 cells were treated with retinol doses reported as physiologic (2 μM) or therapeutic (5, 10 or 20 μM). Retinol at 10 and 20 μM increased free radical production, oxidative damage and antioxidant enzyme activity in A549 cells. These doses also downregulated RAGE expression. Antioxidant co-treatment with Trolox®, a hydrophilic analog of α-tocopherol, reversed the effects of retinol on oxidative parameters and RAGE downregulation. The effect of retinol on RAGE was mediated by p38 MAPK activation, as blockade of p38 with PD169316 (10 μM), SB203580 (10 μM) or siRNA to either p38α (MAPK14) or p38β (MAPK11) reversed the effect of retinol on RAGE. Trolox also inhibited p38 phosphorylation, indicating that retinol induced a redox-dependent activation of this MAPK. Besides, we observed that NF-kB acted as a downstream effector of p38 in RAGE downregulation by retinol, as NF-kB inhibition by SN50 (100 μg/mL) and siRNA to p65 blocked the effect of retinol on RAGE, and p38 inhibitors reversed NF-kB activation. Taken together, our results indicate a pro-oxidant effect of retinol on A549 cells, and suggest that modulation of RAGE expression by

  8. Oxygen Enhancement Ratio in Radiation-Induced Initial DSBs by an Optimized Flow Cytometry-based Gamma-H2AX Analysis in A549 Human Cancer Cells.

    PubMed

    Sunada, Shigeaki; Hirakawa, Hirokazu; Fujimori, Akira; Uesaka, Mitsuru; Okayasu, Ryuichi

    2017-11-01

    High-linear energy transfer (LET) heavy ions cause higher therapeutic effects than low-LET radiation due to lower dependency on oxygen concentration in tumor cell killing. The lethality after irradiation largely depends on DNA double-strand breaks (DSBs), however the detailed LET dependency for DSB induction under oxic and hypoxic conditions has not been reported. Therefore, we evaluated the oxygen enhancement ratio (OER) of heavy ion-induced DSB induction using a highly-optimized flow cytometry-based method of γ-H2AX detection. Non-small cell lung cancer (NSCLC) A549 cells were exposed to X-ray, carbon-ion and iron-ion radiations under oxic or hypoxic condition. As a DSB marker, the γ-H2AX signal was measured 1 h postirradiation and analyzed by flow cytometry. DSB slope values were calculated as DSB induction per Gy. Our method was able to detect high-LET radiation-induced DSBs even from clustered DNA damage sites. We also showed a decrease in OER value in an LET-dependent manner regardless of radiation type. In summary, we demonstrated a simple, quick and highly-optimized flow cytometry-based method of DSB analysis that detects DSBs induced by heavy-ion radiation for hypoxic and nonhypoxic cancer cells. Our study may provide a useful biological basis for heavy-ion radiotherapy.

  9. Adverse effects of industrial multiwalled carbon nanotubes on human pulmonary cells

    PubMed Central

    Tabet, Lyes; Bussy, Cyrill; Amara, Nadia; Setyan, Ari; Grodet, Alain; Rossi, Michel J.; Pairon, Jean-Claude; Boczkowski, Jorge; Lanone, Sophie

    2009-01-01

    The aim of this study was to evaluate adverse effects of multi-walled carbon nanotubes (MWCNT) produced for industrial purposes, on the human epithelial cell line A549. MWCNT were dispersed in dipalmitoyl lecithin (DPL), a component of pulmonary surfactant, and the effects of dispersion in DPL were compared to those in 2 other media: ethanol (EtOH) and phosphate buffer saline (PBS). Effects of MWCNT were also compared to those of 2 asbestos fibers (chrysotile and crocidolite) and carbon black (CB) nanoparticles, not only in A549 cells, but also on mesothelial cells (MeT5A human cell line), used as an asbestos-sensitive cell type. MWCNT formed agglomerates on top of both cell lines (surface area 15–35 μm2), that were significantly larger and more numerous in PBS than in EtOH and DPL. Whatever the dispersion media, incubation with 100 μg/ml MWCNT induced a similar decrease in metabolic activity without changing cell membrane permeability or apoptosis. Neither MWCNT cellular internalization nor oxidative stress were observed. In contrast, asbestos fibers penetrated into the cells, decreased metabolic activity but not cell membrane permeability and increased apoptosis, without decreasing cell number. CB was internalized without any adverse effects. In conclusion, this study demonstrates that MWCNT produced for industrial purposes exert adverse effects without being internalized by human epithelial and mesothelial pulmonary cell lines. PMID:19034795

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

    PubMed

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

    2017-01-01

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

  11. A platycoside-rich fraction from the root of Platycodon grandiflorum enhances cell death in A549 human lung carcinoma cells via mainly AMPK/mTOR/AKT signal-mediated autophagy induction.

    PubMed

    Yim, Nam-Hui; Hwang, Youn-Hwan; Liang, Chun; Ma, Jin Yeul

    2016-12-24

    The root of Platycodon grandiflorum (PG), commonly known as Kilkyong in Korea, Jiegeng in China, and Kikyo in Japan, has been extensively used as a traditional anti-inflammatory medicine in Asia for the treatment of respiratory conditions, such as bronchitis, asthma, and tonsillitis. Platycosides isolated from PG are especially well-known for their anti-cancer effects. We investigated the involvement of autophagic cell death and other potential molecular mechanisms induced by the platycoside-containing butanol fraction of PG (PGB) in human lung carcinoma cells. PGB-induced growth inhibition and cell death were measured using a 5-diphenyl-tetrazolium bromide (MTT) assay. The effects of PGB on autophagy were determined by observing microtubule-associated protein 1 light chain 3 (LC3) redistribution with confocal microscopy. The PGB-mediated regulation of autophagy-associated proteins was investigated using Western blotting analysis. Furthermore, the anti-cancer mechanism of PGB was confirmed using chemical inhibitors. A high-performance liquid chromatography (HPLC)-DAD system was used to analyze the platycosides in PGB. In A549 cells, PGB induced significant autophagic cell death. Specifically, PGB upregulated LC3-II in a time- and dose-dependent manner, and it redistributed LC3 via autophagosome formation in the cytoplasm. PGB treatment increased the phosphorylation of AMP-activated protein kinase (AMPK) and subsequently suppressed the AKT/mammalian target of the rapamycin (mTOR) pathway. Furthermore, PGB inhibited cell proliferation by regulating the mitogen-activated protein kinase (MAPK) pathways. In this study, six types of platycosides were identified in the PGB using HPLC. PGB efficiently induced cancer cell death via autophagy and the modulation of the AMPK/mTOR/AKT and MAPK signaling pathways in A549 cells. Therefore, PGB may be an efficacious herbal anti-cancer therapy. Copyright © 2016. Published by Elsevier Ireland Ltd.

  12. The impact of anticancer activity upon Beta vulgaris extract mediated biosynthesized silver nanoparticles (ag-NPs) against human breast (MCF-7), lung (A549) and pharynx (Hep-2) cancer cell lines.

    PubMed

    Venugopal, K; Ahmad, H; Manikandan, E; Thanigai Arul, K; Kavitha, K; Moodley, M K; Rajagopal, K; Balabhaskar, R; Bhaskar, M

    2017-08-01

    The present study tried for a phyto-synthetic method of producing silver nanoparticles (Ag-NPs) with size controlled as and eco-friendly route that can lead to their advanced production with decorative tranquil morphology. By inducing temperature fluctuation of the reaction mixture from 25 to 80°C the plasmon resonance band raised slowly which had an ultimate effect on size and shape of Ag-NPs as shown by UV-visible spectroscopy and TEM results. The biosynthesized nanoparticles showed good cytotoxic impact against MCF-7, A549 and Hep2 cells compared to normal cell lines. Compared to control plates, the percentage of cell growth inhibition was found to be high with as concentrations of Ag-NPs becomes more as determined by MTT assay. The AO/EtBr staining observations demonstrated that the mechanism of cell death induced by Ag-NPs was due to apoptosis in cancer cells. These present results propose that the silver nanoparticles (Ag-NPs) may be utilized as anticancer agents for the treatment of various cancer types. However, there is a need for study of in vivo examination of these nanoparticles to find their role and mechanism inside human body. Further, studies we plan to do biomarker fabrication from the green synthesized plant extract nanoparticles like silver, gold and copper nanoparticles with optimized shape and sizes and their enhancement of these noble nanoparticles. Copyright © 2017. Published by Elsevier B.V.

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

    PubMed

    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. Copyright © 2015 John Wiley & Sons, Ltd.

  14. Suppression of Cell Growth, Migration and Drug Resistance by Ethanolic Extract of Antrodia cinnamomea in Human Lung Cancer A549 Cells and C57BL/6J Allograft Tumor Model

    PubMed Central

    Wu, Chi-Han; Liu, Fon-Chang; Lai, Ming-Tsung; Lan, Shou-Jen; Wu, Chieh-Hsi; Sheu, Ming-Jyh

    2018-01-01

    The purpose of this study was to investigate the inhibitory activities of ethanolic extracts from Antrodia cinnamomea (EEAC) on lung cancer. Cell proliferation and cell cycle distribution were analyzed using (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) (MTT) assay and flow cytometry, respectively. Wound-healing assay, Western blotting, and a murine tumor model were separately used to examine cell migration, protein expression, and tumor repression. Our results showed that EEAC induced cell cycle arrest at the G0/G1 phase resulting decreased cell viability in A549 cells. Moreover, EEAC up-regulated the growth-suppressing proteins, adenosine 5′-monophosphate-activated protein kinase (AMPK), p21 and p27, but down-regulated the growth-promoting proteins, protein kinase B (Akt), mammalian tarfet of rapamycin (mTOR), extracellular signal-regulating kinase 1/2 (ERK1/2), retinoblastoma protein (Rb), cyclin E, and cyclin D1. EEAC also inhibited A549 cell migration and reduced expression of gelatinases. In addition, our data showed that tumor growth was suppressed after treatment with EEAC in a murine allograft tumor model. Some bioactive compounds from EEAC, such as cordycepin and zhankuic acid A, were demonstrated to reduce the protein expressions of matrix metalloproteinase (MMP)-9 and cyclin D1 in A549 cells. Furthermore, EEAC enhanced chemosensitivity of A549 to paclitaxel by reducing the protein levels of caveolin-1. Our data suggests that EEAC has the potential to be an adjuvant medicine for the treatment of lung cancer. PMID:29522490

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

    PubMed

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

    2015-05-01

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

  16. Effects of EP4 solution and LPD solution vs Euro-Collins solution on Na(+)/K(+)-ATPase activity in rat alveolar type II cells and human alveolar epithelial cell line A549 cells.

    PubMed

    Suzuki, S; Inoue, K; Sugita, M; Tsubochi, H; Kondo, T; Fujimura, S

    2000-09-01

    Intact alveolar epithelial Na(+)/K(+)- adenosinetriphosphatase (ATPase) function is important in preventing alveolar fluid accumulation after lung transplantation. We examined whether the type of preservation solution used influences Na(+)/K(+)-ATPase activity in alveolar epithelial cells. Rat alveolar type II cells were preserved with EP4, low-potassium dextran (LPD), or Euro-Collins solution at 7 degrees C for 5 and 20 hours. To assess cell toxicity, we measured cell viability and lactate dehydrogenase release. Na(+)/K(+)-ATPase activity was measured as ouabain-sensitive ATPase hydrolysis. We also examined the effect of terbutaline (10(-3) mol/liter) and dibutyryl cyclic adenosine monophosphate (dbcAMP) (10(-3) mol/liter) on Na(+)/K(+)-ATPase activity in A549 cells preserved for 5 hours. All solutions caused significant damage of rat alveolar type II cells at 20 hours. However, Na(+)/K(+)-ATPase activity was preserved at normal levels with EP4 and LPD over 20 hours. Terbutaline and dbcAMP significantly increased Na(+)/K(+)-ATPase activity in A549 cells preserved with EP4 and LPD solutions for 5 hours. However, we observed no activation in the cells preserved with Euro-Collins solution. We found no significant difference in intracellular cAMP levels after terbutaline challenge among the types of preservation solution. We conclude that extracellular-type solutions such as EP4 and LPD may be preferable for maintaining not only the basal activity but also the ability to activate Na(+)/K(+)-ATPase in response to beta-adrenergic agonists, in alveolar epithelial cells.

  17. Design, synthesis, and characterization of α, β-unsaturated carboxylic acid, and its urea based derivatives that explores novel epigenetic modulators in human non-small cell lung cancer A549 cell line.

    PubMed

    Chidambaram, Anusha; Sundararaju, Kavya; Chidambaram, Ramesh K; Subbiah, Rajasekaran; Jayaraj, John M; Muthusamy, Karthikeyan; Vilwanathan, Ravikumar

    2018-07-01

    Histone deacetylase inhibitors (HDACi) are a small molecule chemotherapeutics that target the chromatin remodeling through the regulation of histone and non-histone proteins. These inhibitors directed against histone deacetylase (HDAC) enzymes have become an important therapeutic tool in oncology; consequently, scientific efforts have fortified the quest for newer and novel HDACi, which forces the design of structurally innovative HDACi. Various urea containing compounds exhibited admirable anticancer activity. On the basis of these observations, we design and synthesize HDAC specific blocker molecules which are specifically besieged towards class I, class II, and class IV HDAC isoforms to enhance the structural assortment for HDACi. Through docking experiments, we identified that the compounds were tightly bound to the isoforms of the HDAC enzymes at their receptor regions. These derivatives potently inhibited the different isoforms, namely, class I, II, and IV of HDACs, by hyperacetylation of lysine residues in A549 cells. The mechanism of apoptosis is evident, regulating tumor suppressor genes and proteins, thereby facilitating the activation of the death receptor pathway by the tumor necrosis factor (TNF) receptor. These derivative facilitated the induction of reactive oxygen species (ROS) generation leading to downregulation of Bcl 2 , and upregulation of Bax expression, thereby dysregulating mitochondrial membrane potential (ΔΨ m ) to release cytochrome c, and activation of intrinsic pathway. These compounds downregulate the extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway to inhibit cell growth, proliferation, and metastasis through the matrix metalloproteinases (MMPs) MMP2 and MMP9 in A549 cells. These results suggest that our designed urea based derivatives act as epigenetic targeting agents through HDAC inhibition. © 2017 Wiley Periodicals, Inc.

  18. [Pulmonary complications in children with human immunodeficiency virus infection].

    PubMed

    Brockmann V, Pablo; Viviani S, Támara; Peña D, Anamaría

    2007-08-01

    Pulmonary complications in children infected by human immunodeficiency virus (HIV) are common and may be the first manifestation of acquired immunodeficiency syndrome (AIDS). The aim of our study was to review pulmonary diseases and complications in pediatric patients with HIV infection in a large tertiary hospital in Santiago, Chile. We performed a retrospective, descriptive analysis of 17 patients with HIV infection controlled at the Hospital Dr. Sótero del Rio. Respiratory complications/diseases were: overall pneumonia (n: 14), recurrent pneumonia (n: 10), citomegalovirus associated pneumonia (n: 4), Pneumocystis jiroveci associated pneumonia (n: 1) pulmonary tuberculosis (n: 1), lymphoid interstitial pneumonia (n: 3) and chronic pulmonary disease (n: 7). Microorganisms isolated were mostly atypical and frequently associated with severe and chronic pulmonary damage. A high degree of suspicion is required to detect atypical microorganisms promptly, in order to rapidly implement pathogen targeted therapy that could potentially decrease the possibility of sequelae.

  19. Sulforaphane epigenetically demethylates the CpG sites of the miR-9-3 promoter and reactivates miR-9-3 expression in human lung cancer A549 cells.

    PubMed

    Gao, Linbo; Cheng, David; Yang, Jie; Wu, Renyi; Li, Wenji; Kong, Ah-Ng

    2018-02-09

    Increasing evidence suggests that epigenetic aberrations contribute to the development and progression of cancers such as lung cancer. The promoter region of miR-9-3 was recently found to be hypermethylated in lung cancer, resulting in down-regulation of miR-9-3 and poor patient prognosis. Sulforaphane (SFN), a natural compound that is obtained from cruciferous vegetables, has potent anticancer activities. In this study, we aimed to investigate the effect of SFN on restoring the miR-9-3 level in lung cancer A549 cells through epigenetic regulation. DNA methylation of the miR-9-3 promoter was examined using bisulfite genomic sequencing and methylated DNA immunoprecipitation analysis. The expression levels of miR-9-3 and several epigenetic modifying enzymes were measured using quantitative real-time polymerase chain reaction and Western blotting, respectively. The transcriptional activity of the miR-9-3 promoter was evaluated by patch methylation, and histone modifications were analyzed using chromatin immunoprecipitation (ChIP) assays. We found that CpG methylation was reduced in the miR-9-3 promoter and that miR-9-3 expression was increased after 5 days of treatment with SFN. In vitro methylation analysis showed that the methylated recombinant construct exhibited lower luciferase reporter activity than the unmethylated counterpart. ChIP assays revealed that SFN treatment increased H3K4me1 enrichment at the miR-9-3 promoter. Furthermore, SFN treatment attenuated enzymatic DNMT activity and DNMT3a, HDAC1, HDAC3, HDAC6 and CDH1 protein expression. Taken together, these findings indicate that SFN may exert its chemopreventive effects partly through epigenetic demethylation and restoration of miR-9-3. Copyright © 2017. Published by Elsevier Inc.

  20. Cell cycle regulation by glucosamine in human pulmonary epithelial cells.

    PubMed

    Chuang, Kun-Han; Lu, Chih-Shen; Kou, Yu Ru; Wu, Yuh-Lin

    2013-04-01

    Airway epithelial cells play an important role against intruding pathogens. Glucosamine, a commonly used supplemental compound, has recently begun to be regarded as a potential anti-inflammatory molecule. This study aimed to uncover how glucosamine impacts on cellular proliferation in human alveolar epithelial cells (A549) and bronchial epithelial cells (HBECs). With trypan blue-exclusion assay, we observed that glucosamine (10, 20, 50 mM) caused a decrease in cell number at 24 and 48 h; with a flow cytometric analysis, we also noted an enhanced cell accumulation within the G(0)/G(1) phase at 24 h and induction of late apoptosis at 24 and 48 h by glucosamine (10, 20, 50 mM) in A549 cells and HBECs. Examination of phosphorylation in retinoblastoma (Rb) protein, we found an inhibitory effect by glucosamine at 20 and 50 mM. Glucosamine at 50 mM was demonstrated to elevate both the mRNA and protein expression of p53 and heme oxygenase-1 (HO-1), but also caused a reduction in p21 protein expression. In addition, glucosamine attenuated p21 protein stability via the proteasomal proteolytic pathway, as well as inducing p21 nuclear accumulation. Altogether, our results suggest that a high dose of glucosamine may inhibit cell proliferation through apoptosis and disturb cell cycle progression with a halt at G(0)/G(1) phase, and that this occurs, at least in part, by a reduction in Rb phosphorylation together with modulation of p21, p53 and HO-1 expression, and nuclear p21 accumulation. Copyright © 2012 Elsevier Ltd. All rights reserved.

  1. HUMAN PULMONARY DISTOMIASIS CAUSED BY PARAGONIMUS WESTERMANNI.

    PubMed

    Nakagawa, K

    1917-09-01

    1. The morbidity of pulmonary distomiasis among the school children in the plains of the Prefecture of Shinchiku is 4.3 per cent, while in the mountainous regions among the savages it reaches in some districts 50 per cent. 2. Seventeen species of cercariae were discovered in fresh water mollusks in the Prefecture of Shinchiku, Formosa. But it was impossible to ascertain from morphological characteristics alone which of them developed into the pulmonary fluke. Consequently, the eggs of the pulmonary fluke after hatching into miracidia were allowed to come into contact with several species of fresh water mollusks, of which they infected two. But as it was difficult to keep the two spedes alive in the aquarium long enough to get cercariae, the second intermediate hosts of the pulmonary distomas were looked for in the severely infected villages of the savage tribes. 3. The miracidia of the pulmonary distomas leave the egg about 4 weeks after they are first set free in the water, and if they do not reach mollusks they soon die. 4. Three species of fresh water mollusks were found to act as the first intermediate host of the pulmonary distomas; viz., Melania libertina Gould, Melania tuberculata Mueller, and Melania obliquegranosa Smith. 5. The cercariae of the pulmonary distoma may be identified by their small size and a spine in the oral sucker. They develop in the liver of the three spedes of Melania mentioned above. 6. The second intermediate hosts of the pulmonary distoma,detected in the Prefecture of Shinchiku, are the following three species of fresh water crabs: Potamon (Geothelphusa) obtusipes Stimpson (native name, red crab), Potamon (Geothelphusa) dehaanii White (native name, dung crab), and Eriocheir japonicus De Haan (native name, hairy crab). In addition it was discovered that the following two species might act as intermediate hosts: Sesarma dehaanii Milne-Edwards and Potamon (Parathelphusa) sinensis Milne-Edwards. In Formosa four of the five species are the

  2. HUMAN PULMONARY DISTOMIASIS CAUSED BY PARAGONIMUS WESTERMANNI

    PubMed Central

    Nakagawa, Koan

    1917-01-01

    1. The morbidity of pulmonary distomiasis among the school children in the plains of the Prefecture of Shinchiku is 4.3 per cent, while in the mountainous regions among the savages it reaches in some districts 50 per cent. 2. Seventeen species of cercariæ were discovered in fresh water mollusks in the Prefecture of Shinchiku, Formosa. But it was impossible to ascertain from morphological characteristics alone which of them developed into the pulmonary fluke. Consequently, the eggs of the pulmonary fluke after hatching into miracidia were allowed to come into contact with several species of fresh water mollusks, of which they infected two. But as it was difficult to keep the two spedes alive in the aquarium long enough to get cercariæ, the second intermediate hosts of the pulmonary distomas were looked for in the severely infected villages of the savage tribes. 3. The miracidia of the pulmonary distomas leave the egg about 4 weeks after they are first set free in the water, and if they do not reach mollusks they soon die. 4. Three species of fresh water mollusks were found to act as the first intermediate host of the pulmonary distomas; viz., Melania libertina Gould, Melania tuberculata Mueller, and Melania obliquegranosa Smith. 5. The cercariæ of the pulmonary distoma may be identified by their small size and a spine in the oral sucker. They develop in the liver of the three spedes of Melania mentioned above. 6. The second intermediate hosts of the pulmonary distoma,detected in the Prefecture of Shinchiku, are the following three species of fresh water crabs: Potamon (Geothelphusa) obtusipes Stimpson (native name, red crab), Potamon (Geothelphusa) dehaanii White (native name, dung crab), and Eriocheir japonicus De Haan (native name, hairy crab). In addition it was discovered that the following two species might act as intermediate hosts: Sesarma dehaanii Milne-Edwards and Potamon (Parathelphusa) sinensis Milne-Edwards. In Formosa four of the five species are the

  3. High pemetrexed sensitivity of docetaxel-resistant A549 cells is mediated by TP53 status and downregulated thymidylate synthase

    PubMed Central

    Kuo, Wei-Ting; Tu, Dom-Gene; Chiu, Ling-Yen; Sheu, Gwo-Tarng; Wu, Ming-Fang

    2017-01-01

    The chemoresistance of non-small cell lung cancer (NSCLC) that occurs in docetaxel (DOC) chemotherapy substantially decreases the survival of patients. To overcome DOC-induced chemoresistance, we established DOC-selected A549 lung cancer sublines (A549/D16 and A549/D32) and revealed that both sublines were cross-resistant to vincristine (VCR) and doxorubicin (DXR). Notably, both sublines were more sensitive to pemetrexed (PEM) than parental cells according to MTT and clonogenic assays. The expression levels of thymidylate synthase (TS) and γ-glutamyl hydrolase (GGH) were downregulated in DOC-resistant sublines. When exogenous TS was overexpressed in A549/D16 cells, PEM sensitivity was significantly decreased, however it was not decreased by overexpression of exogenous GGH. PEM treatment induced more apoptotic sub-G1 cells in both DOC-resistant sublines and in the in vivo PEM sensitivities of A549/D16 cells. These findings were further confirmed by a xenografted tumor model. To unmask the mediator of TS downregulation, we investigated human lung cancer cell lines that have various TP53 statuses using DOC treatment. The level of TS protein was significantly decreased in wild-type TP53-containing cells with DOC treatment; TS expression levels were not affected in mutant-TP53 and TP53-null cells under the same conditions. Furthermore, when the expression of TP53 was inhibited in A549 cells, the expression level of TS was increased. Our data indicated that DOC activated wild-type TP53 and suppressed TS expression under continuous DOC exposure. Therefore, the expression of TS remained at low levels in DOC-resistant A549 cancer cells. Our data revealed that for lung cancer with DOC resistance and wild-type TP53 status, the administration of PEM as a second-line agent to overcome DOC-resistance may benefit patients. PMID:28901493

  4. A549 Lung Epithelial Cells Grown as Three-Dimensional Aggregates: Alternative Tissue Culture Model for Pseudomonas aeruginosa Pathogenesis

    PubMed Central

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

    2005-01-01

    A three-dimensional (3-D) lung aggregate model was developed from A549 human lung epithelial cells by using a rotating-wall vessel bioreactor to study the interactions between Pseudomonas aeruginosa and lung epithelial cells. The suitability of the 3-D aggregates as an infection model was examined by immunohistochemistry, adherence and invasion assays, scanning electron microscopy, and cytokine and mucoglycoprotein production. Immunohistochemical characterization of the 3-D A549 aggregates showed increased expression of epithelial cell-specific markers and decreased expression of cancer-specific markers compared to their monolayer counterparts. Immunohistochemistry of junctional markers on A549 3-D cells revealed that these cells formed tight junctions and polarity, in contrast to the cells grown as monolayers. Additionally, the 3-D aggregates stained positively for the production of mucoglycoprotein while the monolayers showed no indication of staining. Moreover, mucin-specific antibodies to MUC1 and MUC5A bound with greater affinity to 3-D aggregates than to the monolayers. P. aeruginosa attached to and penetrated A549 monolayers significantly more than the same cells grown as 3-D aggregates. Scanning electron microscopy of A549 cells grown as monolayers and 3-D aggregates infected with P. aeruginosa showed that monolayers detached from the surface of the culture plate postinfection, in contrast to the 3-D aggregates, which remained attached to the microcarrier beads. In response to infection, proinflammatory cytokine levels were elevated for the 3-D A549 aggregates compared to monolayer controls. These findings suggest that A549 lung cells grown as 3-D aggregates may represent a more physiologically relevant model to examine the interactions between P. aeruginosa and the lung epithelium during infection. PMID:15664956

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

    NASA Astrophysics Data System (ADS)

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

    2016-10-01

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

  6. Comparative analysis of microRNA expression profiles between A549, A549/DDP and their respective exosomes.

    PubMed

    Qin, Xiaobing; Yu, Shaorong; Xu, Xiaoyue; Shen, Bo; Feng, Jifeng

    2017-06-27

    Exosomes were reported to transport bioactive molecules and influence the biology behavior of recipient cells. In order to study the role of exosomal microRNAs in the mechanism of cisplatin resistance to lung cancer cells, we analyzed the expression profiles of microRNAs in A549, A549/DDP cells and their exosomes by microarray. The results showed that a certain proportion of microRNAs were co-expressed in the cells and exosomes. Linear regression analysis showed that the expression of microRNAs in A549 and A549/DDP cells were strongly correlated with those in their respective exosomes. The expression level of 5 microRNAs (miR-197-5p, miR-4443, miR-642a-3p, miR-27b-3p and miR-100-5p) with the most differential expression were verified by qRT-PCR. The results were consistent with those of the microarray. Target gene prediction and pathway analysis discovered that the microRNAs in the intersections may participate in drug resistance. And the prediction of their association with diseases found that most of these microRNAs was associated with lung cancer. We could draw a preliminary conclusion that microRNAs in exosomes may be involved in the drug resistance of lung cancer cells to cisplatin.

  7. Transforming growth factor-β impairs glucocorticoid activity in the A549 lung adenocarcinoma cell line.

    PubMed

    Salem, S; Harris, T; Mok, J S L; Li, M Y S; Keenan, C R; Schuliga, M J; Stewart, A G

    2012-08-01

    The lung adenocarcinoma cell line, A549, undergoes epithelial-mesenchymal cell transition (EMT) in response to TGF-β. Glucocorticoids do not prevent the EMT response, but TGF-β induced resistance to the cytokine-regulatory action of glucocorticoids. We sought to characterize the impairment of glucocorticoid response in A549 cells. A549 cells were exposed to TGF-β for up to 96 h before glucocorticoid treatment and challenge with IL-1α to assess glucocorticoid regulation of IL-6 and CXCL8 production. Nuclear localization of the glucocorticoid receptor α (GRα) was ascertained by immunofluorescence and Western blotting. Transactivation of the glucocorticoid response element (GRE) was measured with a transfected GRE-secreted human placental alkaline phosphatase reporter. TGF-β (40-400 pM) reduced the maximum inhibitory effect of dexamethasone on IL-1α-induced IL-6 and CXCL8 production. The impaired glucocorticoid response was detected with 4 h of TGF-β (40 pM) exposure (and 4 h IL-1α to induce CXCL8 expression) and therefore was not secondary to EMT, a process that requires longer incubation periods and higher concentrations of TGF-β. TGF-β also impaired dexamethasone regulation of granulocyte-macrophage colony-stimulating factor in thrombin-stimulated BEAS-2B epithelial cells. Impaired regulation of CXCL8 was associated with markedly reduced GRE transactivation and reduced induction of mRNA for IκBα, the glucocorticoid-inducible leucine zipper and the epithelial sodium channel (SCNN1A). The expression, cellular levels and nuclear localization of GRα were reduced by TGF-β. We have identified mechanisms underlying the impairment of responses to glucocorticoids by TGF-β in the A549 and BEAS-2B cell lines. © 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.

  8. Exogenous p53 upregulated modulator of apoptosis (PUMA) decreases growth of lung cancer A549 cells.

    PubMed

    Liu, Chun-Ju; Zhang, Xia-Li; Luo, Da-Ya; Zhu, Wei-Feng; Wan, Hui-Fang; Yang, Jun-Ping; Yang, Xiao-Jun; Wan, Fu-Sheng

    2015-01-01

    To investigate the influence of exogenous p53 upregulated modulator of apoptosis (PUMA) expression on cell proliferation and apoptosis in human non-small cell lung cancer A549 cells and transplanted tumor cell growth in nude mice. A549 cells were divided into the following groups: control, non- carrier (NC), PUMA (transfected with pCEP4- (HA) 2-PUMA plasmid), DDP (10 μg/mL cisplatin treatment) and PUMA+DDP (transfected with pCEP4-(HA)2-PUMA plasmid and 10 μg/mL cisplatin treatment). The MTT method was used to detect the cell survival rate. Cell apoptosis rates were measured by flow cytometry, and PUMA, Bax and Bcl-2 protein expression levels were measured by Western blotting. Compared to the control group, the PUMA, DDP and PUMA+DDP groups all had significantly decreased A549 cell proliferation (p<0.01), with the largest reduction in the PUMA+DDP group. Conversely, the apoptosis rates of the three groups were significantly increased (P<0.01), and the PUMA and DDP treatments were synergistic. Moreover, Bax protein levels significantly increased (p<0.01), while Bcl-2 protein levels significantly decreased (p<0.01). Finally, both the volume and the weights of transplanted tumors were significantly reduced (p<0.01), and the inhibition ratio of the PUMA+DDP group was significantly higher than in the single DDP or PUMA groups. Exogenous PUMA effectively inhibited lung cancer A549 cell proliferation and transplanted tumor growth by increasing Bax protein levels and reducing Bcl-2 protein levels.

  9. Adiponectin down-regulates CREB and inhibits proliferation of A549 lung cancer cells.

    PubMed

    Illiano, Michela; Nigro, Ersilia; Sapio, Luigi; Caiafa, Ilaria; Spina, Annamaria; Scudiero, Olga; Bianco, Andrea; Esposito, Sabrina; Mazzeo, Filomena; Pedone, Paolo Vincenzo; Daniele, Aurora; Naviglio, Silvio

    2017-08-01

    Adipokines are known to play a relevant role in a number of cancer related molecular pathways. Adiponectin is a major adipokine with anti-inflammatory and beneficial metabolic actions. Furthermore, it has been shown to exert anti-carcinogenic effects in various tumor models and some clinical studies suggested an inverse relationship between circulating levels of adiponectin and an increased risk for development of malignancies. On the other hand, the cyclic AMP response element binding (CREB) transcription factor has been clearly linked to lung cancer. we analyzed cell proliferation, cell cycle of A549 cells treated with adiponectin as well as CREB activation status in human lung adenocarcinoma A549 cells and in non-small cell lung cancer (NSCLC) samples. adiponectin treatment, at concentrations ranging between 5 and 50 μg/ml mimicking human serum levels, has a significant effect on reducing tumor cell proliferation of A549 cells, mainly by altering cell cycle progression. Importantly, we provide evidence that adiponectin clearly inhibits in a dose- and time-dependent manner CREB phosphorylation (activation) and, at least in part, also the level of CREB protein itself, preceding and accompanying the anti-proliferative effects in response to adiponectin. Moreover, in agreement with previous studies demonstrating that CREB over-expression occurs in many tumors, we also show by western-blotting from lung specimen that CREB is significantly up-regulated in NSCLC samples compared to adjacent normal tissues from six patients. Overall, our results represent the first evidence of CREB inhibition by adiponectin and may provide new insight into therapeutic strategies for lung cancer. Copyright © 2017. Published by Elsevier Ltd.

  10. Apoptosis inducing ability of silver decorated highly reduced graphene oxide nanocomposites in A549 lung cancer

    PubMed Central

    Khan, Merajuddin; Khan, Mujeeb; Al-Marri, Abdulhadi H; Al-Warthan, Abdulrahman; Alkhathlan, Hamad Z; Siddiqui, Mohammed Rafiq H; Nayak, Vadithe Lakshma; Kamal, Ahmed; Adil, Syed F

    2016-01-01

    Recently, graphene and graphene-based materials have been increasingly used for various biological applications due to their extraordinary physicochemical properties. Here, we demonstrate the anticancer properties and apoptosis-inducing ability of silver doped highly reduced graphene oxide nanocomposites synthesized by employing green approach. These nano composites (PGE-HRG-Ag) were synthesized by using Pulicaria glutinosa extract (PGE) as a reducing agent and were evaluated for their anticancer properties against various human cancer cell lines with tamoxifen as the reference drug. A correlation between the amount of Ag nanoparticles on the surface of highly reduced graphene oxide (HRG) and the anticancer activity of nanocomposite was observed, wherein an increase in the concentration of Ag nanoparticles on the surface of HRG led to the enhanced anticancer activity of the nanocomposite. The nanocomposite PGE-HRG-Ag-2 exhibited more potent cytotoxicity than standard drug in A549 cells, a human lung cancer cell line. A detailed investigation was undertaken and Fluorescence activated cell sorting (FACS) analysis demonstrated that the nanocomposite PGE-HRG-Ag-2 showed G0/G1 phase cell cycle arrest and induced apoptosis in A549 cells. Studies such as, measurement of mitochondrial membrane potential, generation of reactive oxygen species (ROS) and Annexin V-FITC staining assay suggested that this compound induced apoptosis in human lung cancer cells. PMID:27022256

  11. [Pulmonary arterial hypertension associated to human immunodeficiency virus].

    PubMed

    Sandoval-Gutiérrez, José Luis; Santos-Martínez, Luis Efren; Rodríguez-Silverio, Juan; Baranda-Tovar, Francisco Martín; Rivera-Rosales, Rosa María; Flores-Murrieta, Francisco Javier

    2015-01-01

    From the advent of the highly effective antiretroviral treatment, the life expectancy of patients with human immunodeficiency virus has increased significantly. At present, the causes of death are non-infectious complications. Between them, the pulmonary arterial hypertension has a special importance. It is important early detection to establish the therapeutic, with the objective of preventing a fatal outcome to future. Copyright © 2013 Instituto Nacional de Cardiología Ignacio Chávez. Published by Masson Doyma México S.A. All rights reserved.

  12. [Nickel exposure to A549 cell damage and L-ascorbic acid interference effect].

    PubMed

    Fu, Yao; Wang, Yue; Dan, Han; Zhang, Lin; Ma, Wenhan; Pan, Yulin; Wu, Yonghui

    2015-05-01

    Studying different concentrations of nickel smelting smoke subjects of human lung adenocarcinoma cells (A549) carcinogenic effects, discusses the influence of L-ascorbic acid protection. The A549 cells were divided into experimental and L-ascorbic acid in the intervention group. Plus exposure group concentration of nickel refining dusts were formulated 0.00, 6.25, 12.50, 25.00, 50.00, 100.00 µg/ml suspension, the intervention group on the basis of the added exposure group containing L-ascorbic acid (100 mmol/L), contact 24 h. Detection of cell viability by MTT assay. When the test substance concentration select 0.00, 25.00, 50.00, 100.00 µg/ml experiment for internal Flou-3 fluorescent probe to detect cell Ca²⁺ concentration, within DCFH-DA detect intracellular reactive oxygen (ROS) content, real-time quantitative PCR (real time, in the RT-PCR) was used to detect cell HIF-1α gene expression. With the increase of concentration, subjects increased cell growth inhibition rate, intracellular Ca²⁺ concentration increases, ROS content increased, HIF-1α gene expression increased, differences were statistically significant (P < 0.05). After L-ascorbic acid intervention treatment, the results of the intervention group were lower than that of the experimental group, and the difference was statistically significant (P < 0.05), so L-ascorbic acid can effectively protect the nickel exposure damage to cells. With subjects following exposure to nickel concentration increased, its effect on A549 cell damage increases, L-ascorbic acid cell damage caused by nickel has certain protective effect.

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

    PubMed

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

    2017-09-01

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

  14. Accumulation of surfactant protein D in human pulmonary alveolar proteinosis.

    PubMed Central

    Crouch, E.; Persson, A.; Chang, D.

    1993-01-01

    Surfactant protein D (SP-D) is a collagenous calcium-dependent carbohydrate-binding protein that is structurally related to the serum mannose-binding proteins and pulmonary surfactant protein A. SP-D was initially characterized as a biosynthetic product of freshly isolated rat type II cells and first purified in chemical amounts from bronchoalveolar lavage of rats with silica-induced alveolar lipoproteinosis. The present studies describe the characterization of human SP-D isolated from therapeutic bronchoalveolar lavage of patients with pulmonary alveolar proteinosis. Human proteinosis SP-D was extracted from the 10,000 x g pellet of bronchoalveolar lavage with 100 mmol/L glucose or ethylenediamine tetraacetic acid, and specifically bound to and eluted from maltosyl-agarose. The protein cross-reacted with monospecific antibodies to rat SP-D by enzyme-linked immunosorbent assay and immunoblot and eluted near the position of rat SP-D on reverse-phase high performance liquid chromatography. When chromatographed on 4% agarose (A-15M) in the presence of ethylenediamine tetraacetic acid, the solubilized human proteinosis SP-D eluted near the void volume and earlier than rat SP-D dodecamers or human SP-D multimers in the lavage supernatant. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting of proteins in the lavage pellet with antibodies to the carbohydrate-binding domain of proteinosis human SP-D demonstrated covalently cross-linked multimers of SP-D monomers (43 kd, reduced) and multimers of trimeric components stabilized by disulfide and non-disulfide bonds. These studies describe the isolation and biochemical characterization of human SP-D and demonstrate the abnormal accumulation of this protein in the air spaces of patients with alveolar proteinosis. Images Figure 1 Figure 2 Figure 3 Figure 5 Figure 6 Figure 7 PMID:8424457

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

    PubMed Central

    Chang, Hong-Bin; Chen, Bing-Huei

    2015-01-01

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

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

    PubMed

    Chang, Hong-Bin; Chen, Bing-Huei

    2015-01-01

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

  17. Cytotoxicity of semiconductor nanoparticles in A549 cells is attributable to their intrinsic oxidant activity

    NASA Astrophysics Data System (ADS)

    Escamilla-Rivera, Vicente; Uribe-Ramirez, Marisela; Gonzalez-Pozos, Sirenia; Velumani, Subramaniam; Arreola-Mendoza, Laura; De Vizcaya-Ruiz, Andrea

    2016-04-01

    Copper indium gallium diselenide (CIGS) and cadmium sulfide (CdS) nanoparticles (NP) are next generation semiconductors used in photovoltaic cells (PV). They possess high quantum efficiency, absorption coefficient, and cheaper manufacturing costs compared to silicon. Due to their potential for an industrial development and the lack of information about the risk associated in their use, we investigated the influence of the physicochemical characteristics of CIGS (9 nm) and CdS (20 nm) in relation to the induction of cytotoxicity in human alveolar A549 cells through ROS generation and mitochondrial dysfunction. CIGS induced cytotoxicity in a dose dependent manner in lower concentrations than CdS; both NP were able to induce ROS in A549. Moreover, CIGS interact directly with mitochondria inducing depolarization that leads to the induction of apoptosis compared to CdS. Antioxidant pretreatment significantly prevented the loss of mitochondrial membrane potential and cytotoxicity, suggesting ROS generation as the main cytotoxic mechanism. These results demonstrate that semiconductor characteristics of NP are crucial for the type and intensity of the cytotoxic effects. Our work provides relevant information that may help guide the production of a safer NP-based PV technologies, and would be a valuable resource on future risk assessment for a safer use of nanotechnology in the development of clean sources of renewable energy.

  18. α5-nAChR modulates nicotine-induced cell migration and invasion in A549 lung cancer cells.

    PubMed

    Sun, Haiji; Ma, Xiaoli

    2015-09-01

    Cigarette smoking is the most important risk factor in the development of human lung cancer. Nicotine, the major component in tobacco, not only contributes to carcinogenesis but also promotes tumor metastasis. By binding to nicotinic acetylcholine receptors (nAChRs), nicotine induces the proliferation and migration of non-small cell lung cancer. Recently studies have indicated that α5-nAChR is highly associated with lung cancer risk and nicotine dependence. Nevertheless, it is unclear whether nicotine promotes the migration and invasion through activation of α5-nAChR in lung cancer. In the present study, A549 cell was exposed to 1μN nicotine for 8, 24 or 48h. Wound-healing assay and transwell assay were used to evaluate the capability of A549 cell migration and cell invasion, respectively. Silencing of α5-nAChR was done by siRNA. Western blotting and PCR were used to detect α5-nAChR expression. Nicotine can induce activation of α5-nAChR in association with increased migration and invasion of human lung cancer A549 cell. Treatment of cells with α5-nAChR specific siRNA blocks nicotine-stimulated activation of α5-nAChR and suppresses A549 cell migration and invasion. Reduction of α5-nAChR resulted in upregulation of E-cadherin, consistent with E-cadherin being inhibitive of cancer cell invasion. These findings suggest that nicotine-induced migration and invasion may occur in a mechanism through activation of α5-nAChR, which can contribute to metastasis or development of human lung cancer. Copyright © 2015 Elsevier GmbH. All rights reserved.

  19. Indole-3-carbinol induces apoptosis through p53 and activation of caspase-8 pathway in lung cancer A549 cells.

    PubMed

    Choi, Hee-Sook; Cho, Min-Chul; Lee, Hee Gu; Yoon, Do-Young

    2010-03-01

    Indole-3-carbinol (I3C) has anti-tumor effects in various cancer cell lines. However, the anti-tumor effect of I3C on human lung cancers has been rarely reported. We investigated the anti-tumor effects and its mechanism of I3C on human lung carcinoma A549 cell line. Treatment of the A549 cells with I3C significantly reduced cell proliferation, increased formations of fragmented DNA and apoptotic body, and induced cell cycle arrest at G0/G1 phase. I3C increased not only the protein levels of cyclin D1, phosphorylated p53, and p21 but also the expression of Fas mRNA. Cleavage of caspase-9, -8, -3 and PARP also was increased by I3C. Treatment with wortmannin significantly suppressed both I3C-induced Ser15 phosphorylation and accumulation of p53 protein. The inhibition of caspase-8 by z-IETD-FMK significantly decreased cleavage of procaspase-8,-3 and PARP in I3C-treated A549 cells. Taken together, these results demonstrate that I3C induces cell cycle arrest at G0/G1 through the activation of p-p53 at Ser 15 and induces caspase-8 mediated apoptosis via the Fas death receptor. This molecular mechanism for apoptotic effect of I3C on A549 lung carcinoma cells may be a first report and suggest that I3C may be a preventive and therapeutic agent against lung cancer. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  20. No associations of human pulmonary tuberculosis with Sp110 variants.

    PubMed

    Thye, T; Browne, E N; Chinbuah, M A; Gyapong, J; Osei, I; Owusu-Dabo, E; Niemann, S; Rüsch-Gerdes, S; Horstmann, R D; Meyer, C G

    2006-07-01

    After a recent report on the role of the Ipr1 gene in mediating innate immunity in a mouse model of Mycobacterium tuberculosis infection, the human Ipr1 homologue, Sp110, was considered a promising candidate for an association study in human tuberculosis. In a sample of >1000 sputum positive, HIV negative West African patients with pulmonary tuberculosis and >1000 exposed, apparently healthy controls, we have genotyped 21 Sp110 gene variants that were either available from public databases, including HapMap data, or identified by DNA re-sequencing. No significant differences in the frequencies of any of the 21 variants were observed between patients and controls. This applied also for HapMap tagging variants and the corresponding haplotypes, when including sliding window analyses with three adjacent variants, and when stratifying controls for positivity and negativity according to the results of intradermal tuberculin (purified protein derivative, PPD) skin tests. DNA re-sequencing revealed 13 novel Sp110 variants in the 5'-UTR, exons, and adjacent intronic regions. Based on the results obtained in this case-control study, the hypothesis that Sp110 variants and haplotypes might be associated with distinct phenotypes of human M tuberculosis infection is doubtful.

  1. Applying cybernetic technology to diagnose human pulmonary sounds.

    PubMed

    Chen, Mei-Yung; Chou, Cheng-Han

    2014-06-01

    Chest auscultation is a crucial and efficient method for diagnosing lung disease; however, it is a subjective process that relies on physician experience and the ability to differentiate between various sound patterns. Because the physiological signals composed of heart sounds and pulmonary sounds (PSs) are greater than 120 Hz and the human ear is not sensitive to low frequencies, successfully making diagnostic classifications is difficult. To solve this problem, we constructed various PS recognition systems for classifying six PS classes: vesicular breath sounds, bronchial breath sounds, tracheal breath sounds, crackles, wheezes, and stridor sounds. First, we used a piezoelectric microphone and data acquisition card to acquire PS signals and perform signal preprocessing. A wavelet transform was used for feature extraction, and the PS signals were decomposed into frequency subbands. Using a statistical method, we extracted 17 features that were used as the input vectors of a neural network. We proposed a 2-stage classifier combined with a back-propagation (BP) neural network and learning vector quantization (LVQ) neural network, which improves classification accuracy by using a haploid neural network. The receiver operating characteristic (ROC) curve verifies the high performance level of the neural network. To expand traditional auscultation methods, we constructed various PS diagnostic systems that can correctly classify the six common PSs. The proposed device overcomes the lack of human sensitivity to low-frequency sounds and various PS waves, characteristic values, and a spectral analysis charts are provided to elucidate the design of the human-machine interface.

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

    PubMed

    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 < .0001). None of the compounds tested had any significant effect on GSH. Resveratrol at concentrations > 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.

  3. Human Pulmonary Infection by the Zoonotic Metastrongylus salmi Nematode. The First Reported Case in the Americas.

    PubMed

    Calvopina, Manuel; Caballero, Henry; Morita, Tatsushi; Korenaga, Masataka

    2016-10-05

    Pulmonary metastrongylosis, a zoonotic disease found primarily in pigs, is caused by eight different species of the cosmopolitan nematode Metastrongylus genus. To date, only four human cases have been reported, all from Europe. Herein, a severe case of pulmonary infection caused by Metastrongylus salmi in an Ecuadorian man, with successful treatment with ivermectin, is described. © The American Society of Tropical Medicine and Hygiene.

  4. Human Pulmonary Infection by the Zoonotic Metastrongylus salmi Nematode. The First Reported Case in the Americas

    PubMed Central

    Calvopina, Manuel; Caballero, Henry; Morita, Tatsushi; Korenaga, Masataka

    2016-01-01

    Pulmonary metastrongylosis, a zoonotic disease found primarily in pigs, is caused by eight different species of the cosmopolitan nematode Metastrongylus genus. To date, only four human cases have been reported, all from Europe. Herein, a severe case of pulmonary infection caused by Metastrongylus salmi in an Ecuadorian man, with successful treatment with ivermectin, is described. PMID:27382078

  5. Cell uptake of paclitaxel solid lipid nanoparticles modified by cell-penetrating peptides in A549 cells.

    PubMed

    Zhang, Yin-Long; Zhang, Zhen-Hai; Jiang, Tian-Yue; Ayman-Waddad; Jing-Li; Lv, Hui-Xia; Zhou, Jian-Ping

    2013-01-01

    The aim of this study was to investigate the cytotoxicity of paclitaxel solid lipid nanoparticles (SLN) modified with stearic acid octaarginine (SA-R8-PTX-SLN) as well as the cellular uptake of coumarin-6-loaded SLN modified with SA-R8 (SA-R8-C6-SLN) in human lung cancer cells, A549. SLN were prepared using a film dispersion method; and then their particle size, zeta potential, morphology, bound efficiency of SAR8, drug loading efficiency, and in vitro release were characterized. SA-R8-PTX-SLN and SA-R8-C6-SLN were incubated with A549 cells to measure their cytotoxicity and cellular uptake, respectively. The results indicated that the cytotoxicity of SA-R8-PTX-SLN was enhanced significantly with the increasing amount of SA-R8 and the cellular uptakes of SLN increased with the incubated concentrations and the incubated time of SLN. In contrast, SA-R8-SLN could significantly enhance the cellular uptake of SLN and the cytotoxicity of PTX in A549 cells. These in vitro results suggest that SA-R8-SLN could be proposed as alternative drug delivery system.

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

    PubMed

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

    2016-12-01

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

  7. Uptake and efflux kinetics, and intracellular activity of voriconazole against Aspergillus fumigatus in human pulmonary epithelial cells: a new application for the prophylaxis and early treatment of invasive pulmonary aspergillosis.

    PubMed

    Wang, Taotao; Yang, Qianting; Chen, Lu; Li, Ying; Meng, Ti; Wang, Yan; Zhang, Tao; Lei, Jin'e; Xing, Jianfeng; Dong, Yalin

    2017-06-01

    Invasive pulmonary aspergillosis (IPA), most caused by Aspergillus fumigatus, is a serious life-threatening infection in immunocompromised patients. Voriconazole is used to prevent and treat IPA. However, little is known about the pharmacological characteristics of voriconazole in pulmonary epithelial cells, which are the target site for the prophylaxis and early treatment of IPA. The aim of the study was to evaluate the kinetics and activity of voriconazole against A. fumigatus in A549 cells. High-performance liquid chromatography/tandem mass spectrometry and time-kill method were used to study the cellular pharmacokinetic and pharmacodynamics of voriconazole. Voriconazole exerted a concentration-dependent toxic effect on A549 cells and could penetrate into cells, reaching plateau concentrations of 1.14 ± 0.64, 3.72 ± 1.38 and 6.36 ± 0.95 ng/mg protein after A549 cells were exposed to voriconazole at extracellular concentrations of 2, 8 and 16 mg/L for 2 h, respectively. The efflux of voriconazole was rapid, with a half-life of 10.2 min. Voriconazole can decrease the A. fumigatus conidia invade cells, and the number of viable A. fumigatus conidia in cells can be decreased 2.1- to 20.6-fold when A549 cells were cultured in medium containing voriconazole. After 24-h incubation, 75.6% and 80.5% of intracellular A. fumigatus were killed when extracellular voriconazole concentration was 8 and 16 mg/L, respectively. This study illustrated a new application for the prophylaxis and early treatment of IPA from the cellular pharmacokinetics and pharmacodynamics and emphasized the importance of monitoring concentrations of voriconazole in epithelial lining fluid in immunocompromised patients receiving voriconazole therapy. © 2016 Société Française de Pharmacologie et de Thérapeutique.

  8. Adhesion of MRC-5 and A549 cells on poly(dimethylsiloxane) surface modified by proteins.

    PubMed

    Zuchowska, Agnieszka; Kwiatkowski, Piotr; Jastrzebska, Elzbieta; Chudy, Michal; Dybko, Artur; Brzozka, Zbigniew

    2016-02-01

    PDMS is a very popular material used for fabrication of Lab-on-a-Chip systems for biological applications. Although PDMS has numerous advantages, it is a highly hydrophobic material, which inhibits adhesion and proliferation of the cells. PDMS surface modifications are used to enrich growth of the cells. However, due to the fact that each cell type has specific adhesion, it is necessary to optimize the parameters of these modifications. In this paper, we present an investigation of normal (MRC-5) and carcinoma (A549) human lung cell adhesion and proliferation on modified PDMS surfaces. We have chosen these cell types because often they are used as models for basic cancer research. To the best of our knowledge, this is the first presentation of this type of investigation. The combination of a gas-phase processing (oxygen plasma or ultraviolet irradiation) and wet chemical methods based on proteins' adsorption was used in our experiments. Different proteins such as poly-l-lysine, fibronectin, laminin, gelatin, and collagen were incubated with the activated PDMS samples. To compare with other works, here, we also examined how ratio of prepolymer to curing agent (5:1, 10:1, and 20:1) influences PDMS hydrophilicity during further modifications. The highest adhesion of the tested cells was observed for the usage of collagen, regardless of PDMS ratio. However, the MRC-5 cell line demonstrated better adhesion than A549 cells. This is probably due to the difference in their morphology and type (normal/cancer). © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Alterations of A549 lung cell gene expression in response to biochemical toxins.

    PubMed

    Boesewetter, D E; Collier, J L; Kim, A M; Riley, M R

    2006-03-01

    Health risks associated with the inhalation of potentially toxic materials have been a topic of great public concern. In vitro cellular analyses can provide mechanistic information on the molecular-level responses of lung-derived cell lines to a variety of these hazards. This understanding may be used to develop methods to reduce the damage from such toxins or to detect early stages of their effects. Here we describe an evaluation of the alterations in gene expression of an immortalized lung cell line (A549, human type II epithelia) to a variety of inhalation health hazards including etoposide, gliotoxin, streptolysin O, methyl methansesulfonate (MMS), and Triton X-100. The A549 cells display a dose-response relationship to each toxin with initial responses including alterations in metabolic activity, increases in membrane permeability, and initiation of response genes. In general, membrane-damaging agents (streptolysin O and Triton X-100) induce production of new ion channel proteins, structural proteins, and metabolic enzymes. Gliotoxin impacted the metabolic machinery, but also altered ion channels. Etoposide and MMS caused alterations in the cell cycle, induced DNA repair enzymes, and initiated apoptotic pathways, but MMS also induced immune response cascades. The mechanism of cell response to each toxin is supported by physiological analyses that indicated a fairly slow initiation of cell response to all compounds tested, except for Triton, which caused rapid decline in cell function due to solubilization of the cell membrane. However, Triton does induce production of a number of cell membrane-associated proteins and so its effects at low concentrations are likely translated throughout the cell. Together these results indicate a broader array of cellular responses to each of the test toxins than have previously been reported.

  10. The soybean isoflavonoid equol blocks ritonavir-induced endothelial dysfunction in porcine pulmonary arteries and human pulmonary artery endothelial cells.

    PubMed

    Cheng, Charlie; Wang, Xinwen; Weakley, Sarah M; Kougias, Panagiotis; Lin, Peter H; Yao, Qizhi; Chen, Changyi

    2010-01-01

    HIV protease inhibitor (PI) ritonavir (RTV) may cause vascular injury through oxidative stress. Our purpose in this study was to determine whether equol, a soy isoflavone, could prevent RTV-induced endothelial dysfunction in porcine pulmonary arteries and in human pulmonary artery endothelial cells (HPAEC). Fresh porcine pulmonary artery rings were treated with 15 micromol/L of RTV and/or equol in concentrations of 0.1, 1, and 10 micromol/L for 24 h. A control was set with no amount of equol or RTV administered. Based on myograph tension analysis, RTV significantly reduced endothelium-dependent relaxation in response to bradykinin in the artery rings compared with untreated vessels, whereas the antioxidant equol effectively reversed the RTV effect in a concentration-dependent manner. RTV also reduced the contraction of artery rings in response to thromboxane A(2) analogue U46619 and this reduction was blocked by equol. In addition, RTV treatment significantly reduced endothelial nitric oxide synthase (eNOS) expression in both porcine pulmonary arteries and HPAEC, whereas equol effectively blocked RTV-induced eNOS downregulation. Furthermore, RTV significantly increased superoxide anion production, whereas equol reversed this effect of RTV in porcine pulmonary arteries. Thus, the antioxidant equol effectively protects vascular function from the detrimental effects of HIV PI RTV in both porcine pulmonary arteries and HPAEC via a reduction in the vasomotor dysfunction, eNOS downregulation, and oxidative stress induced by RTV. These novel data suggest that equol may have a clinical application in preventing HIV-associated cardiovascular complications.

  11. Human Regional Pulmonary Gas Exchange with Xenon Polarization Transfer (XTC)

    NASA Astrophysics Data System (ADS)

    Muradian, Iga; Butler, James; Hrovat, Mirko; Topulos, George; Hersman, Elizabeth; Ruset, Iulian; Covrig, Silviu; Frederick, Eric; Ketel, Stephen; Hersman, F. W.; Patz, Samuel

    2007-03-01

    Xenon Transfer Contrast (XTC) is an existing imaging method (Ruppert et al, Magn Reson Med, 51:676-687, 2004) that measures the fraction F of ^129Xe magnetization that diffuses from alveolar gas spaces to septal parenchymal tissue in lungs in a specified exchange time. As previously implemented, XTC is a 2-breath method and has been demonstrated in anesthetized animals. To use XTC in humans and to avoid issues associated with obtaining identical gas volumes on subsequent breath-hold experiments as well as precise image registration in post-processing, a single breath XTC method was developed that acquires three consecutive gradient echo images in an 8s acquisition. We report here initial measurements of the mean and variance of F for 5 normal healthy subjects as well as 7 asymptomatic smokers. The experiments were performed at two lung volumes (˜45 and 65% of TLC). We found that both the mean and variance of F increased with smoking history. In comparison, standard pulmonary function tests such as DLCO FEV1 showed no correlation with smoking history.

  12. PD-1/PD-L1 Pathway Mediates the Alleviation of Pulmonary Fibrosis by Human Mesenchymal Stem Cells in Humanized Mice.

    PubMed

    Ni, Ke; Liu, Ming; Zheng, Jian; Wen, Liyan; Chen, Qingyun; Xiang, Zheng; Lam, Kowk-Tai; Liu, Yinping; Chan, Godfrey Chi-Fung; Lau, Yu-Lung; Tu, Wenwei

    2017-12-08

    Pulmonary fibrosis is a chronic progressive lung disease with few treatments. Human mesenchymal stem cells (MSC) have been shown to be beneficial to pulmonary fibrosis as they have the immunomodulatory capacity. However, there is no reliable model to test the therapeutic effect of human MSC in vivo. Here, to mimic pulmonary fibrosis in humans, we established a novel bleomycin-induced pulmonary fibrosis model in humanized mice. Based on this model, the benefit of human MSC to pulmonary fibrosis and underlying mechanism were investigated. In addition, the relevant parameters in the patients with pulmonary fibrosis were also examined. We demonstrated that human CD8+ T cells were critical for the induction of pulmonary fibrosis in humanized mice. Human MSC could alleviate pulmonary fibrosis and improve lung function by suppressing bleomycin-induced human T cell infiltration and pro-inflammatory cytokine productions in the lungs from humanized mice. Importantly, the alleviation of pulmonary fibrosis by human MSC was mediated by the PD-1/PD-L1 pathway. Moreover, the abnormal PD-1 expression was found in circulating T cells and lung tissues of the patients with pulmonary fibrosis. Our study supports the potential benefit of targeting PD-1/PD-L1 pathway for the treatment of pulmonary fibrosis.

  13. Supramolecular assembly of human pulmonary surfactant protein SP-D.

    PubMed

    Arroyo, R; Martin-González, A; Echaide, M; Jain, A; Brondyk, W H; Rosenbaum, J; Moreno-Herrero, F; Perez-Gil, J

    2018-04-04

    Pulmonary surfactant protein D (SP-D) is a glycoprotein from the collectin family that is a component of the lung surfactant system. It exhibits host defense and immune regulatory functions in addition to contributing to the homeostasis of the surfactant pool within the alveolar airspaces. It is known that the SP-D monomer forms trimers, which further associate into higher order oligomers. However, the pathway and the interactions involved in the assembly of SP-D oligomers are not clearly understood. In the current study, a recombinant form of full-length human SP-D (rhSP-D) has been qualitatively and quantitatively studied by Atomic Force Microscopy (AFM) and electrophoresis, with the aim to understand the conformational diversity and the determinants defining the oligomerization of the protein. The rhSP-D preparation studied is a mixture of trimers, hexamers, dodecamers and higher order oligomeric species, with dodecamers accounting for more than 50% of the protein by mass. Similar structures were also found in hSP-D obtained from proteinosis patients, with the largest fuzzy-ball-like oligomers being more abundant in these samples. The proportion of dodecamer is increased under acidic conditions, accompanied by a conformational change into more compact configurations. Two hexamers appear to be the minimal necessary unit for dodecamer formation, with stabilization of the dodecamer occurring via non-covalent ionic and hydrophobic interactions between the individual N-terminal domains and the proximal area of the SP-D collagen stems. Copyright © 2018. Published by Elsevier Ltd.

  14. MiR-9 enhances the sensitivity of A549 cells to cisplatin by inhibiting autophagy.

    PubMed

    Zhang, Yan; Meng, Xia; Li, Cheng; Tan, Zhoulin; Guo, Xinwei; Zhang, Zhiting; Xi, Tao

    2017-07-01

    To demonstrate that miR-9 inhibits autophagy by down-regulating Beclin1 and thus enhances the sensitivity of A549 cells to cisplatin. MiR-9 inhibited Beclin1 expression by binding to its 3'UTR. The inhibition decreased the cisplatin-induced autophagy in A549 cells, evidenced by the decreased expression of LC3II and GFP-LC3 puncta and the increased expression of P62. Upregulation of miR-9 level enhanced the sensibility of A549 cells to cisplatin and increased the cisplatin-induced apoptosis. Overexpression of Beclin1 reversed above effects of miR-9 mimics, cisplatin-induced autophagy was increased and apoptosis was decreased. MiR-9 inhibits autophagy via targeting Beclin1 3'UTR and thus enhances cisplatin sensitivity in A549 cells.

  15. Radiation-Induced Bystander Effects in A549 Cells Exposed to 6 MV X-rays.

    PubMed

    Yang, Shuning; Xu, Jing; Shao, Weixian; Geng, Chong; Li, Jia; Guo, Feng; Miao, Hui; Shen, Wenbin; Ye, Tao; Liu, Yazhou; Xu, Haiting; Zhang, Xuguang

    2015-07-01

    The aim of the study is to explore the bystander effects in A549 cells that have been exposed to 6MV X-ray. Control group, irradiated group, irradiated conditioned medium (ICM)-received group, and fresh medium group were designed in this study. A549 cells in the logarithmic growth phase were irradiated with 6MV X-ray at 0, 0.5, 1, 1.5, and 2. In ICM-received group, post-irradiation A549 cells were cultured for 3 h and were transferred into non-irradiated A549 cells for further cultivation. Clone forming test was applied to detect the survival fraction of cells. Annexin V-FITC/PI double-staining assay was used to detect the apoptosis of A549 cells 24, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation, and the curves of apoptosis were drawn. The changes in the cell cycles 4, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation were detected using PI staining flow cytometry. With the increase of irradiation dose, the survival fraction of A549 cells after the application of 0.5 Gy irradiation was decreasing continuously. In comparison to the control group, the apoptosis rate of the ICM-received group was increased in a time-dependent pattern, with the highest apoptosis rate observed at 72 h (p < 0.05). Cell count in G2/M stages was obviously increased compared with that of the control group (p < 0.05), with the highest count observed at 72 h, after which G2/M stage arrest was diminished. ICM can cause apparent A549 cell damage, indicating that 6MV X-ray irradiation can induce bystander effect on A549 cells, which reaches a peak at 72 h.

  16. Copper doping enhanced the oxidative stress-mediated cytotoxicity of TiO2nanoparticles in A549 cells.

    PubMed

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

    2018-05-01

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

  17. Grape Seed Proanthocyanidin Inhibits Mucin Synthesis and Viral Replication by Suppression of AP-1 and NF-κB via p38 MAPKs/JNK Signaling Pathways in Respiratory Syncytial Virus-Infected A549 Cells.

    PubMed

    Lee, Jin-Woo; Kim, Young Il; Im, Chang-Nim; Kim, Sung Wan; Kim, Su Jin; Min, Seoyeon; Joo, Yong Hoon; Yim, Sung-Vin; Chung, Namhyun

    2017-06-07

    Airway epithelial cells are often infected by respiratory syncytial virus (RSV), one of the most common causes of asthma, bronchiolitis, chronic obstructive pulmonary disease, and pneumonia. During the infection process, excessive mucins instigate airway inflammation. However, the mechanism underlying RSV-induced airway hyper-responsiveness and inflammation is poorly understood. Furthermore, no reliable vaccines or drugs for antiviral therapy are available. In this study, the effect of the natural compound grape seed proanthocyanidin (GSP) on RSV-infected human airway epithelial cells A549 was evaluated. After pretreatment of the cells with or without exposure to RSV with 5-10 μg GSP/mL, the expression of various mucins (MUC1, MUC2, MUC5AC, MUC5B, and MUC8) was evaluated by real-time polymerase chain reaction, enzyme-linked immunosorbent assay, and Western blotting, as well as confocal microscopy. We found that GSP significantly decreased RSV-induced mucin synthesis at the mRNA and protein levels. In addition, GSP suppressed the RSV-induced signaling pathways, including extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38, together with nuclear factor kappa B (NF-κB) and activating protein-1 family members (c-Jun and c-Fos). Concomitantly, GSP inhibited the replication of RSV within A549 cells. Taken together, all our results suggest that GSP could be a potent therapeutic agent to suppress excessive mucus production and viral replication in RSV-induced airway inflammatory disorders.

  18. AMP-activated protein kinase (AMPK) activation is involved in chrysin-induced growth inhibition and apoptosis in cultured A549 lung cancer cells.

    PubMed

    Shao, Jun-jie; Zhang, Ai-ping; Qin, Wei; Zheng, Lin; Zhu, Yi-fan; Chen, Xin

    2012-07-06

    Here we show that chrysin induces growth inhibition and apoptosis in cultured lung cancer A549 cells, and activation of AMP-activated protein kinase (AMPK) may contribute to this process. Our Western-blots results demonstrated a significant AMPK activation after chrysin treatment in A549 cells. Inhibition of AMPK by shRNA-mediated gene silencing, or by its inhibitor, diminished chrysin-induced A549 cell growth inhibition and apoptosis. Forced activation of AMPK by introducing a constitutively active form of AMPKα (CA-AMPKα), or by its activators, mimicked chrysin's effect. For mechanism analysis, we found chrysin inhibited Akt/mammalian target of rapamycin (mTOR) activation, and knocking-down of AMPK by shRNA almost reversed this effect. Finally, we observed that a relative low dose of chrysin enhanced doxorubicin-induced AMPK activation to promote A549 cell apoptosis. Our study suggests that activation of AMPK by chrysin contributes to Akt suppression, growth inhibition and apoptosis in human lung cancer cells, and agents that could activate AMPK may serve as useful adjuvants for traditional chemotherapy against lung cancer. Copyright © 2012 Elsevier Inc. All rights reserved.

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

    PubMed

    Qin, Xia; Qiu, Feng; Zou, Zhen

    2017-11-04

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

  20. Pulmonary surfactant mitigates silver nanoparticle toxicity in human alveolar type-I-like epithelial cells.

    PubMed

    Sweeney, Sinbad; Leo, Bey Fen; Chen, Shu; Abraham-Thomas, Nisha; Thorley, Andrew J; Gow, Andrew; Schwander, Stephan; Zhang, Junfeng Jim; Shaffer, Milo S P; Chung, Kian Fan; Ryan, Mary P; Porter, Alexandra E; Tetley, Teresa D

    2016-09-01

    Accompanying increased commercial applications and production of silver nanomaterials is an increased probability of human exposure, with inhalation a key route. Nanomaterials that deposit in the pulmonary alveolar region following inhalation will interact firstly with pulmonary surfactant before they interact with the alveolar epithelium. It is therefore critical to understand the effects of human pulmonary surfactant when evaluating the inhalation toxicity of silver nanoparticles. In this study, we evaluated the toxicity of AgNPs on human alveolar type-I-like epithelial (TT1) cells in the absence and presence of Curosurf(®) (a natural pulmonary surfactant substitute), hypothesising that the pulmonary surfactant would act to modify toxicity. We demonstrated that 20nm citrate-capped AgNPs induce toxicity in human alveolar type I-like epithelial cells and, in agreement with our hypothesis, that pulmonary surfactant acts to mitigate this toxicity, possibly through reducing AgNP dissolution into cytotoxic Ag(+) ions. For example, IL-6 and IL-8 release by TT1 cells significantly increased 10.7- and 35-fold, respectively (P<0.01), 24h after treatment with 25μg/ml AgNPs. In contrast, following pre-incubation of AgNPs with Curosurf(®), this effect was almost completely abolished. We further determined that the mechanism of this toxicity is likely associated with Ag(+) ion release and lysosomal disruption, but not with increased reactive oxygen species generation. This study provides a critical understanding of the toxicity of AgNPs in target human alveolar type-I-like epithelial cells and the role of pulmonary surfactant in mitigating this toxicity. The observations reported have important implications for the manufacture and application of AgNPs, in particular for applications involving use of aerosolised AgNPs. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Activation of paracrine growth factors by heparan sulphate induced by glucocorticoid in A549 lung carcinoma cells.

    PubMed Central

    Yevdokimova, N.; Freshney, R. I.

    1997-01-01

    Alkaline phosphatase, a marker of differentiation in the human alveolar adenocarcinoma cell line A549, is inducible by conditioned medium from lung fibroblasts and by cytokines including oncostatin M and interleukin 6, but only in the presence of a glucocorticoid, dexamethasone. Dexamethasone was shown to induce incorporation of [3H]glucosamine into three fractions of medium and cell trypsinate from subconfluent A549 cells, eluting from DEAE ion-exchange chromatography. The first peak did not correspond to any of the unlabelled glycosaminoglycans and was not characterized further. Induction was seen in two other peaks, corresponding to hyaluronic acid and heparan sulphate. Of these, heparan sulphate, eluting as one well-defined peak (referred to as HS1) and another of lower activity and less well defined (HS2), was selected as the most likely to interact with growth factors and cytokines and was isolated from the eluate, concentrated and desalted, and used in alkaline phosphatase induction experiments in place of dexamethasone. HS1 isolated from the medium (HS1m) of subconfluent A549 cells was shown to replace dexamethasone in induction experiments with fibroblast-conditioned medium, oncostatin M and interleukin 6. HS1 from the cell trypsinate and HS2 from the medium and trypsinate were inactive. As the activity of HS1m could be abolished by heparinase and heparitinase but not by chondroitinase ABC, it was concluded that HS1m was a fraction of heparan sulphate involved in the regulation of paracrine growth factor activity in lung fibroblast-conditioned medium, and in the regulation of other growth factors with potential roles in the paracrine control of cell differentiation. PMID:9252193

  2. Activation of paracrine growth factors by heparan sulphate induced by glucocorticoid in A549 lung carcinoma cells.

    PubMed

    Yevdokimova, N; Freshney, R I

    1997-01-01

    Alkaline phosphatase, a marker of differentiation in the human alveolar adenocarcinoma cell line A549, is inducible by conditioned medium from lung fibroblasts and by cytokines including oncostatin M and interleukin 6, but only in the presence of a glucocorticoid, dexamethasone. Dexamethasone was shown to induce incorporation of [3H]glucosamine into three fractions of medium and cell trypsinate from subconfluent A549 cells, eluting from DEAE ion-exchange chromatography. The first peak did not correspond to any of the unlabelled glycosaminoglycans and was not characterized further. Induction was seen in two other peaks, corresponding to hyaluronic acid and heparan sulphate. Of these, heparan sulphate, eluting as one well-defined peak (referred to as HS1) and another of lower activity and less well defined (HS2), was selected as the most likely to interact with growth factors and cytokines and was isolated from the eluate, concentrated and desalted, and used in alkaline phosphatase induction experiments in place of dexamethasone. HS1 isolated from the medium (HS1m) of subconfluent A549 cells was shown to replace dexamethasone in induction experiments with fibroblast-conditioned medium, oncostatin M and interleukin 6. HS1 from the cell trypsinate and HS2 from the medium and trypsinate were inactive. As the activity of HS1m could be abolished by heparinase and heparitinase but not by chondroitinase ABC, it was concluded that HS1m was a fraction of heparan sulphate involved in the regulation of paracrine growth factor activity in lung fibroblast-conditioned medium, and in the regulation of other growth factors with potential roles in the paracrine control of cell differentiation.

  3. Endotoxin potency in the A549 lung epithelial cell bioassay and the limulus amebocyte lysate assay.

    PubMed

    Hansen, L A; Poulsen, O M; Würtz, H

    1999-06-24

    The purpose of this study was to elucidate to what extent the potency of endotoxins measured by the limulus amebocyte lysate (LAL) assay is reflected in the potency in an in vitro assay based on release of interleukin-8 (IL-8) from a lung epithelial cell line, A549. Lipopolysaccharides (LPS) from Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella enteritidis and detoxified LPS from E. coli were applied in serial dilutions in the LAL assay and in the A549 bioassay. Also 19 organic dust samples from waste recycling plants were tested. The A549 cells were incubated for 24 h with LPS or dust, and the IL-8 secretion was determined by ELISA. The method for evaluation of the LAL assay showed linearity for the four endotoxins. Using the slope as a measure of the potency factor (PF), LPS from E. coli and S. enteritidis was about four times more potent than that for LPS from K. pneumoniae and P. aeruginosa. In the A549 bioassay each of the different types of endotoxin had characteristic and very different dose-response curves. The potency of the LPS, in the A549 bioassay, ranked as follows K. pneumoniae > P. aeruginosa > E. coli > or = S. enteritidis. The content of endotoxin in the dust samples did not correlate with their potency in the A549 bioassay. The present study indicates a poor correlation between the potency of endotoxin in the LAL assay compared with the A549 bioassay. The lack of correlation when organic dust samples are tested may reflect the fact that these samples contain biological active compounds, which are non-reactive in the LAL-assay but stimulate IL-8 secretion from epithelial cells.

  4. The influence of incubation time on adenovirus quantitation in A549 cells by most probable number.

    PubMed

    Cashdollar, Jennifer L; Huff, Emma; Ryu, Hodon; Grimm, Ann C

    2016-11-01

    Cell culture based assays used to detect waterborne viruses typically call for incubating the sample for at least two weeks in order to ensure that all the culturable virus present is detected. Historically, this estimate was based, at least in part, on the length of time used for detecting poliovirus. In this study, we have examined A549 cells infected with human adenovirus type 2, and have found that a three week incubation of virus infected cells results in a higher number of detected viruses by quantal assay than what is seen after two weeks of incubation, with an average 955% increase in Most Probable Number (MPN) from 2 weeks to 3 weeks. This increase suggests that the extended incubation time is essential for accurately estimating viral titer, particularly for slow-growing viruses, UV treated samples, or samples with low titers of virus. In addition, we found that for some UV-treated samples, there was no detectable MPN at 2 weeks, but after 3 weeks, MPN values were obtained. For UV-treated samples, the average increase in MPN from 2 weeks to 3 weeks was 1401%, while untreated samples averaged a change in MPN of 674%, leading us to believe that the UV-damaged viral DNA may be able to be repaired such that viral replication then occurs. Published by Elsevier B.V.

  5. Biosynthesis of gold nanoparticles and related cytotoxicity evaluation using A549 cells.

    PubMed

    Sathishkumar, M; Pavagadhi, S; Mahadevan, A; Balasubramanian, R

    2015-04-01

    Biosynthesis of gold nanoparticles (AuNPs) has become an attractive area of research as it is environmentally benign. The toxicity of AuNPs synthesized by chemical routes has been widely studied. However, little is known about the toxicity associated with the biological synthesis of AuNPs. The present study was carried out to synthesize AuNPs using star anise (Illicium verum; a commercially available spice in abundance)and evaluate its toxicity using human epithelial lung cells (A549) in comparison with AuNPs synthesized by the traditional chemical methods (using sodium citrate and sodium borohydride). Apart from cell viability, markers of oxidative stress (reduced glutathione) and cell death (caspases) were also evaluated to understand the mechanisms of toxicity. Cell viability was observed to be 65.7 percent and 72.3 percent in cells exposed to chemically synthesized AuNPs at the highest dose (200nM) as compared to 80.2 percent for biologically synthesized AuNPs. Protective coating/capping of AuNPs by various polyphenolic compounds present in star anise extract appears to be a major contributor to lower toxicity observed in biologically synthesized AuNPs. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Proteomic Analysis of Cellular Response Induced by Multi-Walled Carbon Nanotubes Exposure in A549 Cells

    PubMed Central

    Zhang, Xing; Jia, Zhenyu; Gao, Xiangjing; Jiang, Ying; Yan, Chunlan; Duerksen-Hughes, Penelope J.; Chen, Fanqing Frank; Li, Hongjuan; Zhu, Xinqiang; Yang, Jun

    2014-01-01

    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). PMID:24454774

  7. Therapeutic Targeting of CC Ligand 21 or CC Chemokine Receptor 7 Abrogates Pulmonary Fibrosis Induced by the Adoptive Transfer of Human Pulmonary Fibroblasts to Immunodeficient Mice

    PubMed Central

    Pierce, Elizabeth M.; Carpenter, Kristin; Jakubzick, Claudia; Kunkel, Steven L.; Flaherty, Kevin R.; Martinez, Fernando J.; Hogaboam, Cory M.

    2007-01-01

    Idiopathic interstitial pneumonias (IIPs) are a collection of pulmonary fibrotic diseases of unknown etiopathogenesis. CC chemokine receptor 7 (CCR7) is expressed in IIP biopsies and primary fibroblast lines, but its role in pulmonary fibrosis was not previously examined. To study the in vivo role of CCR7 in a novel model of pulmonary fibrosis, 1.0 × 106 primary fibroblasts grown from idiopathic pulmonary fibrosis/usual interstitial pneumonia, nonspecific interstitial pneumonia, or histologically normal biopsies were injected intravenously into C.B-17 severe combined immunodeficiency (SCID)/beige (bg) mice. At days 35 and 63 after idiopathic pulmonary fibrosis/usual interstitial pneumonia fibroblast injection, patchy interstitial fibrosis and increased hydroxyproline were present in the lungs of immunodeficient mice. Adoptively transferred nonspecific interstitial pneumonia fibroblasts caused a more diffuse interstitial fibrosis and increased hydroxyproline levels at both times, but injected normal human fibroblasts did not induce interstitial remodeling changes in C.B-17SCID/bg mice. Systemic therapeutic immunoneutralization of either human CCR7 or CC ligand 21, its ligand, significantly attenuated the pulmonary fibrosis in groups of C.B-17SCID/bg mice that received either type of IIP fibroblasts. Thus, the present study demonstrates that pulmonary fibrosis is initiated by the intravenous introduction of primary human fibroblast lines into immunodeficient mice, and this fibrotic response is dependent on the interaction between CC ligand 21 and CCR7. PMID:17392156

  8. The induction of nitric oxide-mediated relaxation of human isolated pulmonary arteries by PACAP

    PubMed Central

    Cardell, Lars Olaf; Hjert, Ola; Uddman, Rolf

    1997-01-01

    The effects of pituitary adenylate cyclase-activating peptide (PACAP) and vasoactive intestinal peptide (VIP) were analysed in human isolated circular segments of pulmonary arteries. Guinea-pig pulmonary arteries were used for comparison. The responses obtained were analysed in relation to the vascular endothelium and the nitric oxide (NO) synthase inhibitor NG-monomethyl L-arginine (L-NMMA).PACAP and VIP induced concentration-dependent relaxations of precontracted pulmonary arteries. The maximal dilator response (Imax,%) and the potency (pEC50 value) were the same for both peptides, and there were no differences in the effects obtained on human and guinea-pig segments. PACAP and VIP were both more potent that acetylcholine (ACh).Removal of the vascular endothelium abolished the PACAP induced dilator response in pulmonary arteries from both species. The VIP induced dilatation was unaffected, whereas the response to ACh was abolished. L-NMMA given before PACAP inhibited the dilatation. Furthermore, L-NMMA also reversed the dilatation already induced by PACAP and excess concentrations of L-arginine restored the dilator response of the L-NMMA treated arteries.PACAP is a potent dilator of human pulmonary arteries. Although the dilator effect seems to be similar in amplitude to the one induced by VIP, the present results suggest differences in the underlying mechanisms of action (endothelium-dependency) between the two peptides. PMID:9134222

  9. Glucocorticoid regulation of human pulmonary surfactant protein-B (SP-B) mRNA stability is independent of activated glucocorticoid receptor

    PubMed Central

    Tillis, Ceá C.; Huang, Helen W.; Bi, Weizhen; Pan, Su; Bruce, Shirley R.

    2011-01-01

    Adequate expression of surfactant protein-B (SP-B) is critical in the function of pulmonary surfactant to reduce alveolar surface tension. Expression of SP-B mRNA is restricted to specific lung-airway epithelial cells, and human SP-B mRNA stability is increased in the presence of the synthetic glucocorticoid dexamethasone (DEX). Although the mechanism of SP-B mRNA stabilization by DEX is unknown, studies suggest involvement of the glucocorticoid receptor (GR). We developed a dual-cistronic plasmid-based expression assay in which steady-state levels of SP-B mRNA, determined by Northern analysis, reproducibly reflect changes in SP-B mRNA stability. Using this assay, we found that steady-state levels of SP-B mRNA increased greater than twofold in transfected human-airway epithelial cells (A549) incubated with DEX (10−7 M). DEX-mediated changes in SP-B mRNA levels required the presence of the SP-B mRNA 3′-untranslated region but did not require ongoing protein synthesis. The effect of DEX on SP-B mRNA levels was dose dependent, with maximal effect at 10−7 M. DEX increased levels of SP-B mRNA in cells lacking GR, and the presence of the GR antagonist RU486 did not interfere with the effect of DEX. Surprisingly, other steroid hormones (progesterone, estradiol, and vitamin D; 10−7 M) significantly increased SP-B mRNA levels, suggesting a common pathway of steroid hormone action on SP-B mRNA stability. These results indicate that the effect of DEX to increase SP-B mRNA stability is independent of activated GR and suggests that the mechanism is mediated by posttranscriptional or nongenomic effects of glucocorticoids. PMID:21398497

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

    PubMed

    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. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

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

    PubMed

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

    2014-03-01

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

  12. Mitomycin-Induced Pulmonary Veno-Occlusive Disease: Evidence From Human Disease and Animal Models.

    PubMed

    Perros, Frédéric; Günther, Sven; Ranchoux, Benoit; Godinas, Laurent; Antigny, Fabrice; Chaumais, Marie-Camille; Dorfmüller, Peter; Hautefort, Aurélie; Raymond, Nicolas; Savale, Laurent; Jaïs, Xavier; Girerd, Barbara; Cottin, Vincent; Sitbon, Olivier; Simonneau, Gerald; Humbert, Marc; Montani, David

    2015-09-01

    Pulmonary veno-occlusive disease (PVOD) is an uncommon form of pulmonary hypertension characterized by the obstruction of small pulmonary veins and a dismal prognosis. PVOD may be sporadic or heritable because of biallelic mutations of the EIF2AK4 gene coding for GCN2. Isolated case reports suggest that chemotherapy may be a risk factor for PVOD. We reported on the clinical, functional, and hemodynamic characteristics and outcomes of 7 cases of PVOD induced by mitomycin-C (MMC) therapy from the French Pulmonary Hypertension Registry. All patients displayed squamous anal cancer and were treated with MMC alone or MMC plus 5-fluoruracil. The estimated annual incidence of PVOD in the French population that have anal cancer is 3.9 of 1000 patients, which is much higher than the incidence of PVOD in the general population (0.5/million per year). In rats, intraperitoneal administration of MMC induced PVOD, as demonstrated by pulmonary hypertension at right-heart catheterization at days 21 to 35 and major remodeling of small pulmonary veins associated with foci of intense microvascular endothelial-cell proliferation of the capillary bed. In rats, MMC administration was associated with dose-dependent depletion of pulmonary GCN2 content and decreased smad1/5/8 signaling. Amifostine prevented the development of MMC-induced PVOD in rats. MMC therapy is a potent inducer of PVOD in humans and rats. Amifostine prevents MMC-induced PVOD in rats and should be tested as a preventive therapy for MMC-induced PVOD in humans. MMC-induced PVOD in rats represents a unique model to test novel therapies in this devastating orphan disease. © 2015 American Heart Association, Inc.

  13. Activities of ten essential oils towards Propionibacterium acnes and PC-3, A-549 and MCF-7 cancer cells.

    PubMed

    Zu, Yuangang; Yu, Huimin; Liang, Lu; Fu, Yujie; Efferth, Thomas; Liu, Xia; Wu, Nan

    2010-04-30

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

  14. APE1 modulates cellular responses to organophosphate pesticide-induced oxidative damage in non-small cell lung carcinoma A549 cells.

    PubMed

    Thakur, Shweta; Dhiman, Monisha; Mantha, Anil K

    2018-04-01

    Monocrotophos (MCP) and chlorpyrifos (CP) are widely used organophosphate pesticides (OPPs), speculated to be linked with human pathologies including cancer. Owing to the fact that lung cells are most vulnerable to the environmental toxins, the development and progression of lung cancer can be caused by the exposure of OPPs. The present study investigates the oxidative DNA damage response evoked by MCP and CP in human non-small cell lung carcinoma A549 cells. A549 cells were exposed to MCP and CP; cytotoxicity and reactive oxygen species (ROS) generation were measured to select the non-toxic dose. In order to establish whether MCP and CP can initiate the DNA repair and cell survival signalling pathways in A549 cells, qRT-PCR and Western blotting techniques were used to investigate the mRNA and protein expression levels of DNA base excision repair (BER)-pathway enzymes and transcription factors (TFs) involved in cell survival mechanisms. A significant increase in cell viability and ROS generation was observed when exposed to low and moderate doses of MCP and CP at different time points (24, 48 and 72 h) studied. A549 cells displayed a dose-dependent accumulation of apurinic/apyrimidinic (AP) sites after 24 h exposure to MCP advocating for the activation of AP endonuclease-mediated DNA BER-pathway. Cellular responses to MCP- and CP-induced oxidative stress resulted in an imbalance in the mRNA and protein expression of BER-pathway enzymes, viz. PARP1, OGG1, APE1, XRCC1, DNA pol β and DNA ligase III α at different time points. The treatment of OPPs resulted in the upregulation of TFs, viz. Nrf2, c-jun, phospho-c-jun and inducible nitric oxide synthase. Immunofluorescent confocal imaging of A549 cells indicated that MCP and CP induces the translocation of APE1 within the cytoplasm at an early 6 h time point, whereas it promotes nuclear localization after 24 h of treatment, which suggests that APE1 subcellular distribution is dynamically regulated in response to

  15. PULMONARY SYSTEM LIMITATIONS TO ENDURANCE EXERCISE PERFORMANCE IN HUMANS

    PubMed Central

    Amann, Markus

    2014-01-01

    Accumulating evidence over the past 25 years depicts the healthy pulmonary system as a limiting factor of whole body endurance exercise performance. This brief overview emphasizes three respiratory system-related mechanisms which impair O2 transport to the locomotor musculature [arterial O2 content (CaO2) × leg blood flow (QL)], i.e. the key determinant of an individual’s aerobic capacity and ability to resist fatigue. First, the respiratory system often fails to prevent arterial desaturation substantially below resting values and thus compromises CaO2. Especially susceptible to this threat to convective O2 transport are well-trained endurance athletes characterized by high metabolic and ventilatory demands and, likely due to anatomical and morphologic gender differences, active females. Second, fatiguing respiratory muscle work (Wresp) associated with strenuous exercise elicits sympathetically-mediated vasoconstriction in limb-muscle vasculature which compromises QL. This impact on limb O2 transport is independent of fitness level and affects all individuals, however, only during sustained, high-intensity endurance exercise performed above ~85% VO2max. And third, excessive fluctuations in intrathoracic pressures accompanying Wresp can limit cardiac output and therefore QL. Exposure to altitude exacerbates the respiratory system limitations observed at sea level and further reduces CaO2 and substantially increases exercise-induced Wresp. Taken together, the intact pulmonary system of healthy endurance athletes impairs locomotor muscle O2 transport during strenuous exercise by failing to ensure optimal arterial oxygenation and compromising QL. This respiratory system-related impact exacerbates the exercise-induced development of fatigue and compromises endurance performance. PMID:22125308

  16. Pulmonary system limitations to endurance exercise performance in humans.

    PubMed

    Amann, Markus

    2012-03-01

    Accumulating evidence over the past 25 years depicts the healthy pulmonary system as a limiting factor of whole-body endurance exercise performance. This brief overview emphasizes three respiratory system-related mechanisms which impair O(2) transport to the locomotor musculature [arterial O(2) content (C(aO(2))) × leg blood flow (Q(L))], i.e. the key determinant of an individual's aerobic capacity and ability to resist fatigue. First, the respiratory system often fails to prevent arterial desaturation substantially below resting values and thus compromises C(aO(2)). Especially susceptible to this threat to convective O(2) transport are well-trained endurance athletes characterized by high metabolic and ventilatory demands and, probably due to anatomical and morphological gender differences, active women. Second, fatiguing respiratory muscle work (W(resp)) associated with strenuous exercise elicits sympathetically mediated vasoconstriction in limb-muscle vasculature, which compromises Q(L). This impact on limb O(2) transport is independent of fitness level and affects all individuals, but only during sustained, high-intensity endurance exercise performed above ∼85% maximal oxygen uptake. Third, excessive fluctuations in intrathoracic pressures accompanying W(resp) can limit cardiac output and therefore Q(L). Exposure to altitude exacerbates the respiratory system limitations observed at sea level, further reducing C(aO(2)) and substantially increasing exercise-induced W(resp). Taken together, the intact pulmonary system of healthy endurance athletes impairs locomotor muscle O(2) transport during strenuous exercise by failing to ensure optimal arterial oxygenation and compromising Q(L). This respiratory system-related impact exacerbates the exercise-induced development of fatigue and compromises endurance performance.

  17. Characterization of indoor dust from Brazil and evaluation of the cytotoxicity in A549 lung cells.

    PubMed

    Deschamps, E; Weidler, P G; Friedrich, F; Weiss, C; Diabaté, S

    2014-04-01

    Over the past decade, ambient air particulate matter (PM) has been clearly associated with adverse health effects. In Brazil, small and poor communities are exposed to indoor dust derived from both natural sources, identified as blowing soil dust, and anthropogenic particles from mining activities. This study investigates the physicochemical and mineralogical composition of indoor PM10 dust samples collected in Minas Gerais, Brazil, and evaluates its cytotoxicity and inflammatory potential. The mean PM10 mass concentration was 206 μg/m(3). The high dust concentration in the interior of the residences is strongly related to blowing soil dust. The chemical and mineralogical compositions were determined by ICP-OES and XRD, and the most prominent minerals were clays, Fe-oxide, quartz, feldspars, Al(hydr)oxides, zeolites, and anatase, containing the transition metals Fe, Cr, V, Ni, Cu, Zn, Ti, and Mn as well as the metalloid As. The indoor dust samples presented a low water solubility of about 6 %. In vitro experiments were carried out with human lung alveolar carcinoma cells (A549) to study the toxicological effects. The influence of the PM10 dust samples on cell viability, intracellular formation of reactive oxygen species (ROS), and release of the pro-inflammatory cytokine IL-8 was analysed. The indoor dust showed little effects on alamarBlue reduction indicating unaltered mitochondrial activity. However, significant cell membrane damage, ROS production, and IL-8 release were detected in dependence of dose and time. This study will support the implementation of mitigation actions in the investigated area in Brazil.

  18. Suitable parameter choice on quantitative morphology of A549 cell in epithelial–mesenchymal transition

    PubMed Central

    Ren, Zhou-Xin; Yu, Hai-Bin; Li, Jian-Sheng; Shen, Jun-Ling; Du, Wen-Sen

    2015-01-01

    Evaluation of morphological changes in cells is an integral part of study on epithelial to mesenchymal transition (EMT), however, only a few papers reported the changes in quantitative parameters and no article compared different parameters for demanding better parameters. In the study, the purpose was to investigate suitable parameters for quantitative evaluation of EMT morphological changes. A549 human lung adenocarcinoma cell line was selected for the study. Some cells were stimulated by transforming growth factor-β1 (TGF-β1) for EMT, and other cells were as control without TGF-β1 stimulation. Subsequently, cells were placed in phase contrast microscope and three arbitrary fields were captured and saved with a personal computer. Using the tools of Photoshop software, some cells in an image were selected, segmented out and exchanged into unique hue, and other part in the image was shifted into another unique hue. The cells were calculated with 29 morphological parameters by Image Pro Plus software. A parameter between cells with or without TGF-β1 stimulation was compared statistically and nine parameters were significantly different between them. Receiver operating characteristic curve (ROC curve) of a parameter was described with SPSS software and F-test was used to compare two areas under the curves (AUCs) in Excel. Among them, roundness and radius ratio were the most AUCs and were significant higher than the other parameters. The results provided a new method with quantitative assessment of cell morphology during EMT, and found out two parameters, roundness and radius ratio, as suitable for quantification. PMID:26182364

  19. Human fetal pulmonary artery velocimetry: repeatability and normal values with emphasis on middle and distal pulmonary vessels.

    PubMed

    Laudy, J A; de Ridder, M A; Wladimiroff, J W

    2000-06-01

    To establish the nature and gestational age dependency of flow velocity waveforms from fetal middle and distal arterial pulmonary branches in the second half of normal pregnancy and to determine repeatability and inter-relationship of flow velocity waveform recordings from proximal, middle and distal arterial pulmonary branches. Cross-sectional study. A total of 111 singleton normal pregnancies between 20 and 40 weeks of gestation were studied using a color-coded Doppler ultrasound system. Pulmonary waveforms were obtained at the level of the fetal cardiac four-chamber view. Repeatability was tested from two recordings at 15 min time-intervals in 25 separate normal pregnancies. Acceptable repeatability of flow velocity waveforms from fetal arterial pulmonary branches was established with coefficients of variation below 15%. The nature of middle arterial pulmonary flow velocity waveforms was similar to that of proximal waveforms and showed a gestational age-related change for diastolic velocity parameters, peak systolic/peak diastolic ratio and pulsatility index. The distal arterial pulmonary branch displayed a monophasic forward flow velocity profile throughout the cardiac cycle. All velocity parameters of the distal branch remained unchanged with advancing gestation, with the exception of the pulsatility index. Significant inter-pulmonary changes were found for all pulmonary arterial waveform parameters. Alteration in pulmonary vascular resistance may play a role in gestational age-related changes, whereas changes in vessel branching/diameter and in the distance between the heart and more distal arterial pulmonary vessels may cause inter-pulmonary differences.

  20. Nano zerovalent iron particles induce pulmonary and cardiovascular toxicity in an in vitro human co-culture model.

    PubMed

    Sun, Zhelin; Yang, Lingyan; Chen, Ku-Fan; Chen, Guan-Wen; Peng, Yen-Ping; Chen, Jen-Kun; Suo, Guangli; Yu, Jiantao; Wang, Wen-Cheng; Lin, Chia-Hua

    2016-09-01

    Despite promising environmental applications for nano zerovalent iron (nZVI), concerns remain about the potential accumulation and toxic effects of nZVI particles. Here, we use an alveolar-capillary co-culture model to investigate a possible link between low-level epithelial exposure to nZVI and pulmonary and cardiovascular toxicity. While nZVI was unable to pass through the epithelial barrier into the endothelium, nZVI exposure did cause oxidative and inflammatory responses in both epithelial and endothelial cells. Therefore, toxic effects induced by nZVI are not restricted to epithelial cells but can be transferred into the endothelium. Communication between A549 and EA.hy926 cells is responsible for amplification of nZVI-induced toxic responses. Decreases in transepithelial electrical resistance and zonula occludens proteins after epithelial exposure to nZVI impaired epithelial barrier integrity. Increases in oxidized α1-antitrypsin and oxidized low-density lipoprotein in the co-culture model suggest that nZVI exposure increases the risk of chronic obstructive pulmonary disease and atherosclerosis. Therefore, inhalation of nZVI has the potential to induce cardiovascular disease through oxidative and inflammatory mediators produced from the damaged lung epithelium in chronic lung diseases.

  1. Cold stress increases reactive oxygen species formation via TRPA1 activation in A549 cells.

    PubMed

    Sun, Wenwu; Wang, Zhonghua; Cao, Jianping; Cui, Haiyang; Ma, Zhuang

    2016-03-01

    Reactive oxygen species (ROS) are responsible for lung damage during inhalation of cold air. However, the mechanism of the ROS production induced by cold stress in the lung is still unclear. In this work, we measured the changes of ROS and the cytosolic Ca(2+) concentration ([Ca(2+)]c) in A549 cell. We observed that cold stress (from 20 to 5 °C) exposure of A549 cell resulted in an increase of ROS and [Ca(2+)]c, which was completely attenuated by removing Ca(2+) from medium. Further experiments showed that cold-sensing transient receptor potential subfamily member 1 (TRPA1) agonist (allyl isothiocyanate, AITC) increased the production of ROS and the level of [Ca(2+)]c in A549 cell. Moreover, HC-030031, a TRPA1 selective antagonist, significantly inhibited the enhanced ROS and [Ca(2+)]c induced by AITC or cold stimulation, respectively. Taken together, these data demonstrated that TRPA1 activation played an important role in the enhanced production of ROS induced by cold stress in A549 cell.

  2. [Pulmonary pathology in fatal human influenza A (H1N1) infection].

    PubMed

    Duan, Xue-jing; Li, Yong; Gong, En-cong; Wang, Jue; Lü, Fu-dong; Zhang, He-qiu; Sun, Lin; Yue, Zhu-jun; Song, Chen-chao; Zhang, Shi-Jie; Li, Ning; Dai, Jie

    2011-12-01

    To study the pulmonary pathology in patients died of fatal human influenza A(H1N1) infection. Eight cases of fatal human influenza A (H1N1) infection, including 2 autopsy cases and 6 paramortem needle puncture biopsies, were enrolled into the study. Histologic examination, immunohistochemitry, flow cytometry and Western blotting were carried out. The major pathologic changes included necrotizing bronchiolitis with surrounding inflammation, diffuse alveolar damage and pulmonary hemorrhage. Influenza viral antigen expression was detected in the lung tissue by Western blotting. Immunohistochemical study demonstrated the presence of nuclear protein and hemagglutinin virus antigens in parts of trachea, bronchial epithelium and glands, alveolar epithelium, macrophages and endothelium. Flow cytometry showed that the apoptotic rate of type II pneumocytes (32.15%, 78.15%) was significantly higher than that of the controls (1.93%, 3.77%). Necrotizing bronchiolitis, diffuse alveolar damage and pulmonary hemorrhage followed by pulmonary fibrosis in late stage are the major pathologic changes in fatal human influenza A (H1N1) infection.

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

    PubMed

    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.

  4. Inhibition of acrolein-stimulated MUC5AC expression by Platycodon grandiflorum root-derived saponin in A549 cells.

    PubMed

    Choi, Jae Ho; Hwang, Yong Pil; Han, Eun Hee; Kim, Hyung Gyun; Park, Bong Hwan; Lee, Hyun Sun; Park, Byung Keun; Lee, Young Chun; Chung, Young Chul; Jeong, Hye Gwang

    2011-09-01

    Mucin overproduction is a hallmark of chronic airway diseases such as chronic obstructive pulmonary disease. In this study, we investigated the inhibition of acrolein-induced expression of mucin 5, subtypes A and C (MUC5AC) by Changkil saponin (CKS) in A549 cells. Acrolein, a known toxin in tobacco smoke and an endogenous mediator of oxidative stress, increases the expression of airway MUC5AC, a major component of airway mucus. CKS, a Platycodon grandiflorum root-derived saponin, inhibited acrolein-induced MUC5AC expression and activity, through the suppression of NF-κB activation. CKS also repressed acrolein-induced phosphorylation of ERK1/2, JNK1/2, and p38MAPK, which are upstream signaling molecules that control MUC5AC expression. In addition, the MAPK inhibitors PD98059 (ERK1/2), SP600125 (JNK1/2), and SB203580 (p38 MAPK), and a PKC delta inhibitor (rottlerin; PKCδ) inhibited acrolein-induced MUC5AC expression and activity. CKS repressed acrolein-induced phosphorylation of PKCδ. Moreover, a reactive oxygen species (ROS) inhibitor, N-acetylcysteine, inhibited acrolein-induced MUC5AC expression and activity through the suppression of PKCδ and MAPK activation, and CKS repressed acrolein-induced ROS production. These results suggest that CKS suppresses acrolein-induced MUC5AC expression by inhibiting the activation of NF-κB via ROS-PKCδ-MAPK signaling. Copyright © 2011 Elsevier Ltd. All rights reserved.

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

    PubMed

    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. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. In vitro inflammatory effects of polyhexamethylene biguanide through NF-κB activation in A549 cells.

    PubMed

    Kim, Ha Ryong; Shin, Da Young; Chung, Kyu Hyuck

    2017-02-01

    Polyhexamethylene biguanide (PHMB) is a member of the polymeric guanidine family, which is used as a biocide and preservative in industrial, medicinal, and consumer products. Some studies reported that polyhexamethylene guanidine phosphate, which is also a member of the guanidine family, induced severe inflammation and fibrosis in the lungs. However, limited studies have evaluated the pulmonary toxicity of PHMB associated with inflammatory responses. The aim of this study was to elucidate the inflammatory responses and its mechanisms induced by PHMB in lung cells. A549 cells exposed to PHMB showed decreased viability, reactive oxygen species (ROS) generation, inflammatory cytokine secretion, and nuclear factor kappa B (NF-κB) activation. The cells showed dose-dependent cytotoxicity and slight generation of ROS. PHMB triggered inflammatory cytokine secretion and NF-κB activation by modulating the degradation of IκB-α and the accumulation of nuclear p65. TNF-α plays important roles in IL-8 expression as well as NF-κB activation. Moreover, IL-8 production induced by PHMB was completely suppressed by a NF-κB inhibitor, but not by a ROS scavenger. In conclusion, we suggest that PHMB induces the inflammatory responses via the NF-κB signaling pathway. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Rapid induction and persistence of paracrine-induced cellular antiviral states arrest viral infection spread in A549 cells

    PubMed Central

    Voigt, Emily A; Swick, Adam; Yin, John

    2016-01-01

    The virus/host interaction is a complex interplay between pro- and anti-viral factors that ultimately determines the spread or halt of virus infections in tissues. This interplay develops over multiple rounds of infection. The purpose of this study was to determine how cellular-level processes combine to impact the spatial spread of infection. We measured the kinetics of virus replication (VSV), antiviral paracrine signal upregulation and secretion, spatial spread of virus and paracrine antiviral signaling, and inhibition of virus production in antiviral-exposed A549 human lung epithelial cells. We found that initially infected cells released antiviral signals 4-to-7 hours following production of virus. However, the subsequent rapid dissemination of signal and fast induction of a robust and persistent antiviral state ultimately led to a suppression of infection spread. This work shows how cellular responses to infection and activation of antiviral responses can integrate to ultimately control infection spread across host cell populations. PMID:27254596

  8. Melatonin suppresses acrolein-induced IL-8 production in human pulmonary fibroblasts.

    PubMed

    Kim, Gun-Dong; Lee, Seung Eun; Kim, Tae-Ho; Jin, Young-Ho; Park, Yong Seek; Park, Cheung-Seog

    2012-04-01

    Cigarette smoke (CS) causes harmful alterations in the lungs and airway structures and functions that characterize chronic obstructive pulmonary disease (COPD). In addition to COPD, active cigarette smoking causes other respiratory diseases and diminishes health status. Furthermore, recent studies show that, α, β-unsaturated aldehyde acrolein in CS induces the production of interleukin (IL)-8, which is known to be related to bronchitis, rhinitis, pulmonary fibrosis, and asthma. In addition, lung and pulmonary fibroblasts secrete IL-8, which has a chemotactic effect on leukocytes, and which in turn, play a critical role in lung inflammation. On the other hand, melatonin regulates circadian rhythm homeostasis in humans and has many other effects, which include antioxidant and anti-inflammatory effects, as demonstrated by the reduced expressions of iNOS, IL-1β, and IL-6 and increased glutathione (GSH) and superoxide dismutase activities. In this study, we investigated whether melatonin suppresses acrolein-induced IL-8 secretion in human pulmonary fibroblasts (HPFs). It was found that acrolein-induced IL-8 production was accompanied by increased levels of phosphorylation of Akt and extracellular signal-regulated kinases (ERK1/2) in HPFs, and that melatonin suppressed IL-8 production in HPFs. These results suggest that melatonin suppresses acrolein-induced IL-8 production via ERK1/2 and phosphatidylinositol 3-kinase (PI3K)/Akt signal inhibition in HPFs. © 2011 John Wiley & Sons A/S.

  9. Determinants of ventilation and pulmonary artery pressure during early acclimatization to hypoxia in humans.

    PubMed

    Fatemian, Marzieh; Herigstad, Mari; Croft, Quentin P P; Formenti, Federico; Cardenas, Rosa; Wheeler, Carly; Smith, Thomas G; Friedmannova, Maria; Dorrington, Keith L; Robbins, Peter A

    2016-03-01

    Pulmonary ventilation and pulmonary arterial pressure both rise progressively during the first few hours of human acclimatization to hypoxia. These responses are highly variable between individuals, but the origin of this variability is unknown. Here, we sought to determine whether the variabilities between different measures of response to sustained hypoxia were related, which would suggest a common source of variability. Eighty volunteers individually underwent an 8-h isocapnic exposure to hypoxia (end-tidal P(O2)=55 Torr) in a purpose-built chamber. Measurements of ventilation and pulmonary artery systolic pressure (PASP) assessed by Doppler echocardiography were made during the exposure. Before and after the exposure, measurements were made of the ventilatory sensitivities to acute isocapnic hypoxia (G(pO2)) and hyperoxic hypercapnia, the latter divided into peripheral (G(pCO2)) and central (G(cCO2)) components. Substantial acclimatization was observed in both ventilation and PASP, the latter being 40% greater in women than men. No correlation was found between the magnitudes of pulmonary ventilatory and pulmonary vascular responses. For G(pO2), G(pCO2) and G(cC O2), but not the sensitivity of PASP to acute hypoxia, the magnitude of the increase during acclimatization was proportional to the pre-acclimatization value. Additionally, the change in G(pO2) during acclimatization to hypoxia correlated well with most other measures of ventilatory acclimatization. Of the initial measurements prior to sustained hypoxia, only G(pCO2) predicted the subsequent rise in ventilation and change in G(pO2) during acclimatization. We conclude that the magnitudes of the ventilatory and pulmonary vascular responses to sustained hypoxia are predominantly determined by different factors and that the initial G(pCO2) is a modest predictor of ventilatory acclimatization. © 2015 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological

  10. Side population cells separated from A549 lung cancer cell line possess cancer stem cell-like properties and inhibition of autophagy potentiates the cytotoxic effect of cisplatin.

    PubMed

    Yang, Yang; Fan, Yuxia; Qi, Yu; Liu, Donglei; Wu, Kai; Wen, Fengbiao; Zhao, Song

    2015-08-01

    Recent studies have suggested that cancer stem cells (CSCs) may be responsible for tumorigenesis and contribute to resistance to chemotherapy. Side population (SP) cells are thought to be enriched for CSCs in most types of human tumors. Therefore, the aim of the present study was to sort SP cells using an A549 lung cancer cell line, identify the cancer stem cell-like properties of SP and determine the role of autophagy in the survival of SP cells of lung cancer. SP cells were isolated by fluorescence-activated cell sorter (FACS) from A549 lung cancer cells, and the CSC-like properties were verified through confocal fluorescence imaging, sphere formation assays, cell proliferation and colony formation assay, gene expression in vitro and tumor formation in vivo. The role of autophagy in the survival of SP cells was assessed by western blotting and flow cytometric analysis. A549 lung cancer cells contained 1.10% SP cells. SP cells showed higher abilities of sphere and colony formation, cell proliferation and self-renewal. Moreover, compared to non-SP, SP cells demonstrated a higher mRNA expression of stem cell markers (MDR1, ABCG2 and OCT-4). The clone formation efficiency of SP cells was significantly higher than that non-SP cells under the same conditions. Expression of autophagosomes in SP cells was markedly lower than that in non-SP cells. However, the level of autophagy in SP cells was found to be markedly increased in the presence of cisplatin. In addition, inhibition of autophagy enhanced the effects of apoptosis induced by cisplatin. SP cells from the A549 lung cancer cell line possessed the properties of CSCs and were used to investigate the further characteristics of lung CSCs. SP cells were more resistant to chemotherapy and inhibition of autophagy enhanced the effects of apoptosis induced by the chemotherapeutic agent, cisplatin. These results may provide insight into novel therapeutic targets.

  11. Theophylline prevents NAD{sup +} depletion via PARP-1 inhibition in human pulmonary epithelial cells

    SciTech Connect

    Moonen, Harald J.J.; Geraets, Liesbeth; Vaarhorst, Anika

    2005-12-30

    Oxidative DNA damage, as occurs during exacerbations in chronic obstructive pulmonary disease (COPD), highly activates the nuclear enzyme poly(ADP-ribose)polymerase-1 (PARP-1). This can lead to cellular depletion of its substrate NAD{sup +}, resulting in an energy crisis and ultimately in cell death. Inhibition of PARP-1 results in preservation of the intracellular NAD{sup +} pool, and of NAD{sup +}-dependent cellular processes. In this study, PARP-1 activation by hydrogen peroxide decreased intracellular NAD{sup +} levels in human pulmonary epithelial cells, which was found to be prevented in a dose-dependent manner by theophylline, a widely used compound in the treatment of COPD. This enzymemore » inhibition by theophylline was confirmed in an ELISA using purified human PARP-1 and was found to be competitive by nature. These findings provide new mechanistic insights into the therapeutic effect of theophylline in oxidative stress-induced lung pathologies.« less

  12. Pulmonary hypertension

    MedlinePlus

    Pulmonary arterial hypertension; Sporadic primary pulmonary hypertension; Familial primary pulmonary hypertension; Idiopathic pulmonary arterial hypertension; Primary pulmonary hypertension; PPH; Secondary pulmonary ...

  13. The Pseudomonas aeruginosa Exopolysaccharide Psl Facilitates Surface Adherence and NF-κB Activation in A549 Cells

    PubMed Central

    Byrd, Matthew S.; Pang, Bing; Mishra, Meenu; Swords, W. Edward; Wozniak, Daniel J.

    2010-01-01

    In order for the opportunistic Gram-negative pathogen Pseudomonas aeruginosa to cause an airway infection, the pathogen interacts with epithelial cells and the overlying mucous layer. We examined the contribution of the biofilm polysaccharide Psl to epithelial cell adherence and the impact of Psl on proinflammatory signaling by flagellin. Psl has been implicated in the initial attachment of P. aeruginosa to biotic and abiotic surfaces, but its direct role in pathogenesis has not been evaluated (L. Ma, K. D. Jackson, R. M. Landry, M. R. Parsek, and D. J. Wozniak, J. Bacteriol. 188:8213–8221, 2006). Using an NF-κB luciferase reporter system in the human epithelial cell line A549, we show that both Psl and flagellin are necessary for full activation of NF-κB and production of the interleukin 8 (IL-8) chemokine. We demonstrate that Psl does not directly stimulate NF-κB activity, but indirectly as a result of increasing contact between bacterial cells and epithelial cells, it facilitates flagellin-mediated proinflammatory signaling. We confirm differential adherence of Psl and/or flagellin mutants by scanning electron microscopy and identify Psl-dependent membrane structures that may participate in adherence. Although we hypothesized that Psl would protect P. aeruginosa from recognition by the epithelial cell line A549, we instead observed a positive role for Psl in flagellin-mediated NF-κB activation, likely as a result of increasing contact between bacterial cells and epithelial cells. PMID:20802825

  14. Cytotoxicity of carbon nanotube variants: a comparative in vitro exposure study with A549 epithelial and J774 macrophage cells.

    PubMed

    Kumarathasan, Prem; Breznan, Dalibor; Das, Dharani; Salam, Mohamed A; Siddiqui, Yunus; MacKinnon-Roy, Christine; Guan, Jingwen; de Silva, Nimal; Simard, Benoit; Vincent, Renaud

    2015-03-01

    While production of engineered carbon nanotubes (CNTs) has escalated in recent years, knowledge of risk associated with exposure to these materials remains unclear. We report on the cytotoxicity of four CNT variants in human lung epithelial cells (A549) and murine macrophages (J774). Morphology, metal content, aggregation/agglomeration state, pore volume, surface area and modifications were determined for the pristine and oxidized single-walled (SW) and multi-walled (MW) CNTs. Cytotoxicity was evaluated by cellular ATP content, BrdU incorporation, lactate dehydrogenase (LDH) release, and CellTiter-Blue (CTB) reduction assays. All CNTs were more cytotoxic than respirable TiO2 and SiO2 reference particles. Oxidation of CNTs removed most metallic impurities but introduced surface polar functionalities. Although slopes of fold changes for cytotoxicity endpoints were steeper with J774 compared to A549 cells, CNT cytotoxicity ranking in both cell types was assay-dependent. Based on CTB reduction and BrdU incorporation, the cytotoxicity of the polar oxidized CNTs was higher compared to the pristine CNTs. In contrast, pristine CNTs were more cytotoxic than oxidized CNTs when assessed for cellular ATP and LDH. Correlation analyses between CNTs' physico-chemical properties and average relative potency revealed the impact of metal content and surface area on the potency values estimated using ATP and LDH assays, while surface polarity affected the potency values estimated from CTB and BrdU assays. We show that in order to reliably estimate the risk posed by these materials, in vitro toxicity assessment of CNTs should be conducted with well characterized materials, in multiple cellular models using several cytotoxicity assays that report on distinct cellular processes.

  15. Human mesenchymal stem cells attenuate pulmonary hypertension induced by prenatal lipopolysaccharide treatment in rats.

    PubMed

    Chou, Hsiu-Chu; Lin, Willie; Chen, Chung-Ming

    2016-10-01

    Intra-amniotic injection of lipopolysaccharide (LPS) induces pulmonary hypertension in newborn rats. This study was designed to test whether human mesenchymal stem cells (MSCs) reduce pulmonary hypertension and alleviate cardiac hypertrophy in prenatal LPS-treated rats. Pregnant Sprague-Dawley rats were injected intraperitoneally with LPS (0.5 mg/kg per day) or untreated on gestational days 20 and 21. Human MSCs (3×10(5) cells and 1×10(6) cells) in 0.03 mL of normal saline (NS) were transplanted intratracheally on postnatal day 5. Four study groups were considered: normal, LPS+NS, LPS+MSCs (3×10(5) cells), and LPS+MSCs (1×10(6) cells). On postnatal day 14, lung and heart tissues were collected for measuring the arterial medial wall thickness (MWT) and β-myosin heavy chain (β-MHC) level as markers of pulmonary hypertension and cardiac hypertrophy, respectively. The LPS+NS group exhibited a significantly higher right ventricle (RV)/[left ventricle (LV)+ interventricular septum (IVS)] thickness ratio and MWT, a greater cardiomyocyte width, a greater number of cardiomyocyte nuclei per squared millimeter, and higher β-MHC expression than those observed in the normal group. Human MSC transplantation (3×10(5) cells and 1×10(6) cells) in LPS-treated rats reduced MWT and the RV/(LV+IVS) thickness ratio to normal levels. This improvement in right ventricular hypertrophy was accompanied by a decrease in toll-like receptor 4 (TLR4), nuclear factor-κB, and tumor necrosis factor-α expression in the heart. Intratracheal human MSCs transplantation can attenuate pulmonary hypertension and right ventricular hypertrophy in prenatal LPS-treated rats; this attenuation may be associated with suppression of TLR4 expression via paracrine pathways. © 2016 John Wiley & Sons Australia, Ltd.

  16. Radiology of pulmonary turberculosis and human immunodeficiency virus infection in Gulu, Uganda.

    PubMed

    Awil, P O; Bowlin, S J; Daniel, T M

    1997-03-01

    Pulmonary tuberculosis is a major complication of human immunodeficiency virus (HIV) infection. The radiographic manifestations of pulmonary tuberculosis in HIV-infected patients are not typical of those seen in immunologically normal individuals. We sought to determine whether these manifestations provide clues to the pathogenesis of tuberculosis in HIV-infected persons. The radiographic manifestations of pulmonary tuberculosis were reviewed and classified in 82 HIV-positive and 53 HIV-negative tuberculous patients in Gulu, Uganda. Pulmonary presentations of tuberculosis were more acute in HIV-positive patients, and often included hilar or mediastinal adenopathy and pleural effusions, findings typical of primary tuberculosis in immunologically normal individuals. Many patients also had chronic forms of tuberculosis, either alone or in combination with acute disease. The findings of this study support the hypothesis that reactivation of latent infections and progression of pre-existent chronic disease produce a substantial portion of the tuberculosis burden of HIV-positive persons in Uganda. Tuberculosis control efforts should extend beyond efforts at decreasing transmission of new infections.

  17. Doppler echo evaluation of pulmonary venous-left atrial pressure gradients: human and numerical model studies

    NASA Technical Reports Server (NTRS)

    Firstenberg, M. S.; Greenberg, N. L.; Smedira, N. G.; Prior, D. L.; Scalia, G. M.; Thomas, J. D.; Garcia, M. J.

    2000-01-01

    The simplified Bernoulli equation relates fluid convective energy derived from flow velocities to a pressure gradient and is commonly used in clinical echocardiography to determine pressure differences across stenotic orifices. Its application to pulmonary venous flow has not been described in humans. Twelve patients undergoing cardiac surgery had simultaneous high-fidelity pulmonary venous and left atrial pressure measurements and pulmonary venous pulsed Doppler echocardiography performed. Convective gradients for the systolic (S), diastolic (D), and atrial reversal (AR) phases of pulmonary venous flow were determined using the simplified Bernoulli equation and correlated with measured actual pressure differences. A linear relationship was observed between the convective (y) and actual (x) pressure differences for the S (y = 0.23x + 0.0074, r = 0.82) and D (y = 0.22x + 0.092, r = 0.81) waves, but not for the AR wave (y = 0. 030x + 0.13, r = 0.10). Numerical modeling resulted in similar slopes for the S (y = 0.200x - 0.127, r = 0.97), D (y = 0.247x - 0. 354, r = 0.99), and AR (y = 0.087x - 0.083, r = 0.96) waves. Consistent with numerical modeling, the convective term strongly correlates with but significantly underestimates actual gradient because of large inertial forces.

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

    NASA Astrophysics Data System (ADS)

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

    2016-12-01

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

  19. Selective killing effect of oxytetracycline, propafenone and metamizole on A549 or Hela cells

    PubMed Central

    Feng, Guihua

    2013-01-01

    Objective To determine the selective killing effect of oxytetracycline, propafenone and metamizole on A549 or Hela cells. Methods Proliferation assay, lactate dehydrogenase (LDH) assay, apoptosis detecting, flow cytometry and western blot were performed. Results It was found that treatment with propafenone at the concentration of 0.014 g/L or higher for 48 h could induce apoptosis in Hela cells greatly, while it was not observed in oxytetracycline and metamizole at the concentration of 0.20 g/L for 48 h. Oxytetracycline, propafenone and metamizole all displayed evident inhibitory effects on the proliferation of A549 cells. The results of LDH assay demonstrated that the drugs at the test range of concentration did not cause necrosis in the cells. Propafenone could elevate the protein level of P53 effectively (P<0.01). Conclusions Oxytetracycline, propafenone and metamizol (dipyrone) all displayed evident inhibitory effects on the proliferation of A549 cells. Propafenone also displayed evident inhibitory effects on the proliferation of Hela cells. PMID:24385693

  20. A549 and MRC-5 cell aggregation in a microfluidic Lab-on-a-chip system.

    PubMed

    Zuchowska, A; Jastrzebska, E; Zukowski, K; Chudy, M; Dybko, A; Brzozka, Z

    2017-03-01

    In this paper, we present a culture of A549 and MRC-5 spheroids in a microfluidic system. The aim of our work was to develop a good lung cancer model for the evaluation of drug cytotoxicity. Our research was focused on determining the progress of cell aggregation depending on such factors as the depth of culture microwells in the microdevices, a different flow rate of the introduced cell suspensions, and the addition of collagen to cell suspensions. We showed that these factors had a significant influence on spheroid formation. It was found that both MRC-5 and A549 cells exhibited higher aggregation in 500  μ m microwells. We also noticed that collagen needs to be added to A549 cells to form the spheroids. Optimizing the mentioned parameters allowed us to form 3D lung tissue models in the microfluidic system during the 10-day culture. This study indicates how important an appropriate selection of the specified parameters is (e.g., geometry of the microwells in the microsystem) to obtain the spheroids characterized by high viability in the microfluidic system.

  1. Protocatechuic aldehyde ameliorates experimental pulmonary fibrosis by modulating HMGB1/RAGE pathway

    SciTech Connect

    Zhang, Liang, E-mail: countryspring@sina.com; Ji, Yunxia, E-mail: 413499057@qq.com; Kang, Zechun, E-mail: davidjiangwl@163.com

    2015-02-15

    An abnormal high mobility group box 1 (HMGB1) activation and a decrease in receptor for advanced glycation end-product (RAGE) play a key role in the pathogenesis of pulmonary fibrosis. Protocatechuic aldehyde (PA) is a naturally occurring compound, which is extracted from the degradation of phenolic acids. However, whether PA has anti-fibrotic functions is unknown. In this study, the effects of PA on the transforming growth factor-β1 (TGF-β1)-mediated epithelial–mesenchymal transition (EMT) in A549 cells, on the apoptosis of human type I alveolar epithelial cells (AT I), on the proliferation of human lung fibroblasts (HLF-1) in vitro, and on bleomycin (BLM)-induced pulmonarymore » fibrosis in vivo were investigated. PA treatment resulted in a reduction of EMT in A549 cells with a decrease in vimentin and HMGB, an increase of E-cadherin and RAGE, a reduction of HLF-1 proliferation with a decrease of fibroblast growth factor 2 (FGF-2) and platelet-derived growth factor (PDGF). Apoptosis of AT I was attenuated with an increase of RAGE. PA ameliorated BLM-induced pulmonary fibrosis in rats with a reduction of histopathological scores and collagen deposition, and a lower FGF-2, PDGF, α-smooth muscle actin (α-SMA) and HMGB1 expression, whereas higher RAGE was found in BLM-instilled lungs. Through the decrease of HGMB1 and the regulation of RAGE, PA reversed the EMT, inhibited HLF-1 proliferation as well as reduced apoptosis in AT I, and prevented pulmonary fibrosis in vivo. Collectively, our results demonstrate that PA prevents experimental pulmonary fibrosis by modulating HMGB1/RAGE pathway. - Highlights: • PA prevents EMT, reduces the apoptosis of AT1 in vitro. • PA decreases proliferation of HLF-1, reduces PDGF and FGF expression in vitro. • PA prevents experimental pulmonary fibrosis by modulating the HMGB1/RAGE pathway.« less

  2. Id proteins are critical downstream effectors of BMP signaling in human pulmonary arterial smooth muscle cells.

    PubMed

    Yang, Jun; Li, Xiaohui; Li, Ying; Southwood, Mark; Ye, Lingying; Long, Lu; Al-Lamki, Rafia S; Morrell, Nicholas W

    2013-08-15

    Bone morphogenetic protein type II receptor (BMPR-II) mutations are responsible for over 70% of cases of heritable pulmonary arterial hypertension (PAH). Loss of BMP signaling promotes pulmonary vascular remodeling via modulation of pulmonary artery smooth muscle cell (PASMC) proliferation. Id proteins (Id1-4) are major downstream transcriptional targets of BMP signaling. However, the impact of BMPR-II mutation on the expression of the range of Id proteins and the contribution of individual Id proteins to abnormal PASMC function remain unclear. Human PASMCs were used to determine the expression of Id proteins (Id1-4) by real-time PCR and immunoblotting. The BMP responses in control cells were compared with PASMCs harboring BMPR-II mutations and cells in which BMPR-II was knocked down by siRNA transfection. Id3 expression in pulmonary vessels was also investigated in BMPR-II mutant mice and in patients with heritable PAH. BMP4 and BMP6, but not BMP9, induced mRNA expression of Id1, Id2, and Id3. The BMP-stimulated induction of Id1 and Id3 was markedly reduced in BMPR-II mutant PASMCs and in control PASMCs following siRNA silencing of BMPR-II. Pulmonary arteries in BMPR-II mutant mice and patients with heritable PAH demonstrated reduced levels of Id3 compared with control subjects. Lentiviral overexpression of Id3 reduced cell cycle progression and inhibited proliferation of PASMCs. Lipopolysaccharide further reduced Id3 expression in mutant PASMCs. In conclusion, Id proteins, and particularly Id1 and Id3, are critical downstream effectors of BMP signaling in PASMCs. Loss of BMPR-II function reduces the induction of Id genes in PASMCs, Id1, and Id3 regulate the proliferation of PASMCs via cell cycle inhibition, an effect that may be exacerbated by inflammatory stimuli.

  3. [Expression of MADD in lung adenocarcinoma tissues and its effects on proliferation and apoptosis of lung adenocarcinoma A549 cells].

    PubMed

    Wei, Yu-Ping; Wu, Jin-Xiang; Guo, Yuan-Fang; Sun, Gao-Ying; Zhang, Qiang; Bi, Wen-Xiang; Dong, Liang

    2012-04-01

    To investigate the expression of MAPK-activating death domain protein (MADD) in lung adenocarcinoma tissues and its effects on proliferation and apoptosis of lung adenocarcinoma A549 cells. Immunohistochemistry was used to detect the expression of MADD in lung normal and tumor tissues. The expression of IG20 gene in A549 cells was measured by reverse transcription polymerase chain reaction. A549 cells were transfected with pEYFP-MADD plasmids carrying MADD gene or pNL-SIN-GFP-MID lentiviral vectors used for RNA interference. MADD expression and cell proliferation and apoptosis were determined by Western blot, MTT assay, and flow cytometry. The expression levels of MADD were higher in lung adenocarcinoma and squamous cell carcinoma tissues than that in lung normal tissues, and lung adenocarcinoma tissues expressed more MADD than lung squamous cell carcinoma tissues. The transcript encoding MADD was expressed in A549 cells. The transfection of pEYFP-MADD plasmids could increase MADD expression and cell proliferation of A549 cells, while the A549 cells transfected with pNL-SIN-GFP-MID lentiviral vectors showed significantly decreases in the MADD level and proliferation. It is shown that MADD overexpression could inhibit A549 cell apoptosis, and knock down of MADD could promote apoptosis of them. The expression of MADD increases obviously in lung adenocarcinoma, and MADD can promote survival of lung adenocarcinoma cells by inhibiting apoptosis.

  4. Decellularization of human and porcine lung tissues for pulmonary tissue engineering.

    PubMed

    O'Neill, John D; Anfang, Rachel; Anandappa, Annabelle; Costa, Joseph; Javidfar, Jeffrey; Wobma, Holly M; Singh, Gopal; Freytes, Donald O; Bacchetta, Matthew D; Sonett, Joshua R; Vunjak-Novakovic, Gordana

    2013-09-01

    The only definitive treatment for end-stage organ failure is orthotopic transplantation. Lung extracellular matrix (LECM) holds great potential as a scaffold for lung tissue engineering because it retains the complex architecture, biomechanics, and topologic specificity of the lung. Decellularization of human lungs rejected from transplantation could provide "ideal" biologic scaffolds for lung tissue engineering, but the availability of such lungs remains limited. The present study was designed to determine whether porcine lung could serve as a suitable substitute for human lung to study tissue engineering therapies. Human and porcine lungs were procured, sliced into sheets, and decellularized by three different methods. Compositional, ultrastructural, and biomechanical changes to the LECM were characterized. The suitability of LECM for cellular repopulation was evaluated by assessing the viability, growth, and metabolic activity of human lung fibroblasts, human small airway epithelial cells, and human adipose-derived mesenchymal stem cells over a period of 7 days. Decellularization with 3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) showed the best maintenance of both human and porcine LECM, with similar retention of LECM proteins except for elastin. Human and porcine LECM supported the cultivation of pulmonary cells in a similar way, except that the human LECM was stiffer and resulted in higher metabolic activity of the cells than porcine LECM. Porcine lungs can be decellularized with CHAPS to produce LECM scaffolds with properties resembling those of human lungs, for pulmonary tissue engineering. We propose that porcine LECM can be an excellent screening platform for the envisioned human tissue engineering applications of decellularized lungs. Copyright © 2013 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  5. Fused pulmonary lobes is a rat model of human Fraser syndrome

    SciTech Connect

    Kiyozumi, Daiji; Nakano, Itsuko; Takahashi, Ken L.

    2011-07-29

    Highlights: {yields} Fused pulmonary lobes (fpl) mutant rats exhibit similar phenotypes to Fraser syndrome. {yields} The fpl gene harbors a nonsense mutation in Fraser syndrome-associated gene Frem2. {yields} Fpl mutant is defined as a first model of human Fraser syndrome in rats. -- Abstract: Fused pulmonary lobes (fpl) is a mutant gene that is inherited in an autosomal recessive manner and causes various developmental defects, including fusion of pulmonary lobes, and eyelid and digit anomalies in rats. Since these developmental defects closely resemble those observed in patients with Fraser syndrome, a recessive multiorgan disorder, and its model animals, we investigatedmore » whether the abnormal phenotypes observed in fpl/fpl mutant rats are attributable to a genetic disorder similar to Fraser syndrome. At the epidermal basement membrane in fpl/fpl mutant neonates, the expression of QBRICK, a basement membrane protein whose expression is attenuated in Fraser syndrome model mice, was greatly diminished compared with control littermates. Quantitative RT-PCR analyses of Fraser syndrome-related genes revealed that Frem2 transcripts were markedly diminished in QBRICK-negative embryos. Genomic DNA sequencing of the fpl/fpl mutant identified a nonsense mutation that introduced a stop codon at serine 2005 in Frem2. These findings indicate that the fpl mutant is a rat model of human Fraser syndrome.« less

  6. Dielectric barrier discharge plasma in Ar/O2 promoting apoptosis behavior in A549 cancer cells

    NASA Astrophysics Data System (ADS)

    Huang, Jun; Li, Hui; Chen, Wei; Lv, Guo-Hua; Wang, Xing-Quan; Zhang, Guo-Ping; Ostrikov, Kostya; Wang, Peng-Ye; Yang, Si-Ze

    2011-12-01

    The Ar/O2 plasma needle in the induction of A549 cancer cells apoptosis process is studied by means of real-time observation. The entire process of programmed cell death is observed. The typical morphological changes of A549 apoptosis are detected by 4', 6-diamidino-2-phenylindole staining, for example, chromatin condensation and nuclear fragmentation. Cell viability is determined and quantified by neutral red uptake assay, and the survival rate of A549 from Ar/O2 plasmas is presented. Further spectral analysis indicates the reactive species, including O and OH play crucial roles in the cell inactivation.

  7. Determinants of ventilation and pulmonary artery pressure during early acclimatization to hypoxia in humans

    PubMed Central

    Fatemian, Marzieh; Herigstad, Mari; Croft, Quentin P. P.; Formenti, Federico; Cardenas, Rosa; Wheeler, Carly; Smith, Thomas G.; Friedmannova, Maria; Dorrington, Keith L.

    2015-01-01

    Key points Lung ventilation and pulmonary artery pressure rise progressively in response to 8 h of hypoxia, changes described as ‘acclimatization to hypoxia’. Acclimatization responses differ markedly between humans for unknown reasons.We explored whether the magnitudes of the ventilatory and vascular responses were related, and whether the degree of acclimatization could be predicted by acute measurements of ventilatory and vascular sensitivities.In 80 healthy human volunteers measurements of acclimatization were made before, during, and after a sustained exposure to 8 h of isocapnic hypoxia.No correlation was found between measures of ventilatory and pulmonary vascular acclimatization.The ventilatory chemoreflex sensitivities to acute hypoxia and hypercapnia all increased in proportion to their pre‐acclimatization values following 8 h of hypoxia. The peripheral (rapid) chemoreflex sensitivity to CO2, measured before sustained hypoxia against a background of hyperoxia, was a modest predictor of ventilatory acclimatization to hypoxia. This finding has relevance to predicting human acclimatization to the hypoxia of altitude. Abstract Pulmonary ventilation and pulmonary arterial pressure both rise progressively during the first few hours of human acclimatization to hypoxia. These responses are highly variable between individuals, but the origin of this variability is unknown. Here, we sought to determine whether the variabilities between different measures of response to sustained hypoxia were related, which would suggest a common source of variability. Eighty volunteers individually underwent an 8‐h isocapnic exposure to hypoxia (end‐tidal P O2=55 Torr) in a purpose‐built chamber. Measurements of ventilation and pulmonary artery systolic pressure (PASP) assessed by Doppler echocardiography were made during the exposure. Before and after the exposure, measurements were made of the ventilatory sensitivities to acute isocapnic hypoxia (GpO2) and

  8. Low level CO2 effects on pulmonary function in humans

    NASA Technical Reports Server (NTRS)

    Sexton, J.; Mueller, K.; Elliott, A.; Gerzer, D.; Strohl, K. P.; West, J. B. (Principal Investigator)

    1998-01-01

    The purpose of the study was to determine whether chamber exposure to low levels of CO2 results in functional alterations in gas mixing and closing volume in humans. Four healthy volunteer subjects were exposed to 0.7% CO2 and to 1.2% CO2. Spirometry, lung volumes, single breath nitrogen washout, diffusing capacity for carbon monoxide (DLCO) by two methods, and cardiac output were measured in triplicate. Values were obtained over two non-consecutive days during the training period (control) and on days 2 or 3, 4, 6, 10, 13, and 23 of exposure to each CO2 level. Measurements were made during the same time of day. There was one day of testing after exposure, while still in the chamber but off carbon dioxide. The order of testing, up until measurements of DLCO and cardiac output, were randomized to avoid presentation effects. The consistent findings were a reduction in diffusing capacity for carbon monoxide and a fall in cardiac output, occurring to a similar degree with both exposures. For the group as a whole, there was no indication of major effects on spirometry, lung volumes, gas mixing or dead space. We conclude that small changes may occur in the function of distal gas exchanging units; however, these effects were not associated with any adverse health effects. The likelihood of pathophysiologic changes in lung function or structure with 0.7 or 1.2% CO2 exposure for this period of time, is therefore, low.

  9. Air Pollution by Hydrothermal Volcanism and Human Pulmonary Function.

    PubMed

    Linhares, Diana; Ventura Garcia, Patrícia; Viveiros, Fátima; Ferreira, Teresa; dos Santos Rodrigues, Armindo

    2015-01-01

    The aim of this study was to assess whether chronic exposure to volcanogenic air pollution by hydrothermal soil diffuse degassing is associated with respiratory defects in humans. This study was carried in the archipelago of the Azores, an area with active volcanism located in the Atlantic Ocean where Eurasian, African, and American lithospheric plates meet. A cross-sectional study was performed on a study group of 146 individuals inhabiting an area where volcanic activity is marked by active fumarolic fields and soil degassing (hydrothermal area) and a reference group of 359 individuals inhabiting an area without these secondary manifestations of volcanism (nonhydrothermal area). Odds ratio (OR) and 95% confidence intervals (CIs) were adjusted for age, gender, fatigue, asthma, and smoking. The OR for restrictive defects and for exacerbation of obstructive defects (COPD) in the hydrothermal area was 4.4 (95% CI 1.78-10.69) and 3.2 (95% CI 1.82-5.58), respectively. Increased prevalence of restrictions and all COPD severity ranks (mild, moderate, and severe) was observed in the population from the hydrothermal area. These findings may assist health officials in advising and keeping up with these populations to prevent and minimize the risk of respiratory diseases.

  10. Water-pipe smoke condensate increases the internalization of Mycobacterium Bovis of type II alveolar epithelial cells (A549).

    PubMed

    Mortaz, Esmaeil; Alipoor, Shamila D; Movassaghi, Masoud; Varahram, Mohammad; Ghorbani, Jahangir; Folkerts, Gert; Garssen, Johan; Adcock, Ian M

    2017-04-21

    Tuberculosis (TB) is a major global health problem, and there is an association between tobacco smoke and TB. Water pipe smoking has become an increasing problem not only in Middle Eastern countries but also globally because users consider it as safer than cigarettes. The presence of high levels of toxic substances in water-pipe smoke may be a predisposing factor that enhances the incidence of pulmonary disorders. For example, uncontrolled macropinocytosis in alveolar epithelial cells following exposure to water-pipe smoke may predispose subjects to pulmonary infection. Here, we studied the effects of water-pipe condense (WPC) on the internalization of Mycobacterium Bovis BCG by macropinocytosis in the alveolar epithelial cell line A549. A549 cells were exposed to WPC (4 mg/ml) for 24, 48, 72 and 96 h. Cell viability was studied using the methyl thiazolyldipenyl-tetrazolium bromide (MTT) reduction assay and proliferation by bromodeoxyUridine (BrdU) incorporation. Cells were exposed to FITC-Dextran (1 mg/ml) (as a control) and FITC-BCG (MOI = 10) for 20 min at 37 °C before cells were collected and the uptake of BCG-FITC determined by flow cytometry. Similar experiments were performed at 4 °C as a control. The Rho-associated protein kinase (ROCK) inhibitor Y-27632 (1 μM) was used to assess the mechanism by which WPC enhanced BCG uptake. WPC (4 mg/ml) increased the uptake of BCG-FITC after 72 (1.3 ± 0.1 fold, p < 0.05) and 96 (1.4 ± 0.05 fold, p < 0.05) hours. No effect on BCG-FITC uptake was observed at 24 or 48 h. WPC also significantly increased the uptake of FITC-Dextran (2.9 ± 0.3 fold, p < 0.05) after 24 h. WPC significantly decreased cell viability after 24 (84 ± 2%, p < 0.05), 48 (78±, 3%, p < 0.05), 72 (64 ± 2%, p < 0.05) and 96 h (45 ± 2%, p < 0.05). Y-27632 completely attenuated the increased uptake of BCG by WPC. Cell proliferation showed a decreasing trend in a time

  11. Dectin-1 Is Expressed in Human Lung and Mediates the Proinflammatory Immune Response to Nontypeable Haemophilus influenzae

    PubMed Central

    Heyl, Kerstin A.; Klassert, Tilman E.; Heinrich, Annina; Müller, Mario M.; Klaile, Esther; Dienemann, Hendrik; Grünewald, Christiane; Bals, Robert; Singer, Bernhard B.

    2014-01-01

    ABSTRACT The C-type lectin receptor Dectin-1 is expressed mainly on myeloid cells mediating the immune response targeting respiratory pathogens such as Aspergillus fumigatus and Mycobacterium tuberculosis. The pulmonary epithelium serves as an important interface for interactions between these pathogens and the respiratory tract. Therefore, we analyzed the expression pattern of Dectin-1 in the human lung. Immunohistochemically stained human lung sections from 17 out of 19 individuals were positive for Dectin-1, which was expressed mainly apically on bronchial and alveolar epithelium. Our results showed no correlation with chronic obstructive pulmonary disease (COPD) or the smoking habits of the patients. Nontypeable Haemophilus influenzae (NTHI), an important bacterial pathogen of the respiratory tract with significant importance in COPD, has also been proposed to be recognized by Dectin-1, suggesting a possible impact on the NTHI-dependent immune response in human airways. Therefore, the involvement of Dectin-1 in NTHI-triggered cytokine responses was investigated in primary normal human bronchial epithelial (NHBE) cells and in the A549 cell line stably transfected with Dectin-1. The presence of Dectin-1 significantly increased cytokine release in response to NTHI in NHBE and A549 cells. In addition, phosphorylation of the Dectin-1 hem-immunoreceptor tyrosine-based activation motif (hemITAM) was essential for the Dectin-1-triggered response to NTHI in A549 cells. In conclusion, in human airways, epithelium-expressed Dectin-1 may play a significant role in generating an NTHI-mediated, proinflammatory immune response. PMID:25161190

  12. Ozone exposure and pulmonary effects in panel and human clinical studies: Considerations for design and interpretation.

    PubMed

    Rohr, Annette C

    2018-04-01

    A wealth of literature exists regarding the pulmonary effects of ozone, a photochemical pollutant produced by the reaction of nitrogen oxide and volatile organic precursors in the presence of sunlight. This paper focuses on epidemiological panel studies and human clinical studies of ozone exposure, and discusses issues specific to this pollutant that may influence study design and interpretation as well as other, broader considerations relevant to ozone-health research. The issues are discussed using examples drawn from the wider literature. The recent panel and clinical literature is also reviewed. Health outcomes considered include lung function, symptoms, and pulmonary inflammation. Issues discussed include adversity, reversibility, adaptation, variability in ozone exposure metric used and health outcomes evaluated, co-pollutants in panel studies, influence of temperature in panel studies, and multiple comparisons. Improvements in and standardization of panel study approaches are recommended to facilitate comparisons between studies as well as meta-analyses. Additional clinical studies at or near the current National Ambient Air Quality Standard (NAAQS) of 70 ppb are recommended, as are clinical studies in sensitive subpopulations such as asthmatics. The pulmonary health impacts of ozone exposure have been well documented using both epidemiological and chamber study designs. However, there are a number of specific methodological and related issues that should be considered when interpreting the results of these studies and planning additional research, including the standardization of exposure and health metrics to facilitate comparisons among studies.

  13. Signalling pathways involved in the contractile response to 5-HT in the human pulmonary artery.

    PubMed

    Rodat-Despoix, L; Aires, V; Ducret, T; Marthan, R; Savineau, J-P; Rousseau, E; Guibert, C

    2009-12-01

    Serotonin (5-hydroxytryptamine; 5-HT) is a potent pulmonary vasoconstrictor and mitogenic agent whose plasma level is increased in pulmonary hypertensive patients. Thus, we explored the signalling pathways involved in the contractile response to 5-HT in human pulmonary arteries (HPAs). Intact and beta-escin permeabilised rings from HPAs mounted in an organ bath system were used to assess both tension and myofilament Ca(2+)-sensitisation. Microspectrofluorimetry was used for intracellular Ca(2+) recordings in cultured HPA smooth muscle cells. Voltage-operated Ca(2+) channel blockers (nitrendipine and nifedipine) partially reduced the contraction to 5-HT. Thapsigargin or cyclopiazonic acid (CPA), known to deplete sarcoplasmic reticulum Ca(2+) stores, also partially inhibited the contraction, whereas removal of extracellular Ca(2+) under these conditions further inhibited the contraction. Changing from Ca(2+)-free to Ca(2+) containing solution, in the presence of nitrendipine and CPA, a protocol known to stimulate store-operated Ca(2+) channels, induced HPA contractions that were blocked by nickel. Nickel or gadolinium also reduced the contraction to 5-HT. Finally, 5-HT increased intracellular Ca(2+) responses in cultured HPA smooth muscle cells and myofilament Ca(2+)-sensitisation in HPA rings. Collectively, these results indicate that voltage-operated and voltage-independent Ca(2+) channels, as well as Ca(2+) release and myofilament Ca(2+)-sensitisation, participate in 5-HT-induced contraction in HPAs.

  14. Asthma causes inflammation of human pulmonary arteries and decreases vasodilatation induced by prostaglandin I2analogs.

    PubMed

    Foudi, Nabil; Badi, Aouatef; Amrane, Mounira; Hodroj, Wassim

    2017-12-01

    Asthma is a chronic inflammatory disease associated with increased cardiovascular events. This study assesses the presence of inflammation and the vascular reactivity of pulmonary arteries in patients with acute asthma. Rings of human pulmonary arteries obtained from non-asthmatic and asthmatic patients were set up in organ bath for vascular tone monitoring. Reactivity was induced by vasoconstrictor and vasodilator agents. Protein expression of inflammatory markers was detected by western blot. Prostanoid releases and cyclic adenosine monophosphate (cAMP) levels were quantified using specific enzymatic kits. Protein expression of cluster of differentiation 68, intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and cyclooxygenase-2 was significantly increased in arteries obtained from asthmatic patients. These effects were accompanied by an alteration of vasodilatation induced by iloprost and treprostinil, a decrease in cAMP levels and an increase in prostaglandin (PG) E 2 and PGI 2 synthesis. The use of forskolin (50 µmol/L) has restored the vasodilatation and cAMP release. No difference was observed between the two groups in reactivity induced by norepinephrine, angiotensin II, PGE 2 , KCl, sodium nitroprusside, and acetylcholine. Acute asthma causes inflammation of pulmonary arteries and decreases vasodilation induced by PGI 2 analogs through the impairment of cAMP pathway.

  15. Doppler velocimetry in branch pulmonary arteries of normal human fetuses during the second half of gestation.

    PubMed

    Laudy, J A; de Ridder, M A; Wladimiroff, J W

    1997-06-01

    The objective of the present study was to determine the characteristics of Doppler flow velocity wave forms in branch pulmonary arteries in relation to gestational age. A total of 111 singleton normal pregnancies were studied during the second half of pregnancy using a combined color-coded Doppler and two-dimensional real-time ultrasound system. Pulsed Doppler measurements of the most proximal branch of the right or left pulmonary artery were attempted during fetal apnea from a transverse cross-section of the fetal chest at the level of the cardiac four-chamber view after visualization with color Doppler. The success rate in obtaining the pulmonary arterial wave form was 85%. The wave form displayed a rapid systolic velocity acceleration, followed by an initially rapid but then more gradual velocity deceleration which was interrupted in most cases by a short reversed flow interval at the beginning of the diastolic phase of the cardiac cycle. The diastolic phase was characterized by forward flow. Peak systolic, end-diastolic and time-averaged velocity, pulsatility index, and systolic integral remained constant during gestation. Changes in vessel diameter or compliance may play a role in this. A gestational age-dependent rise was established for peak diastolic velocity, diastolic integral, and early peak diastolic reverse flow, whereas a gestational age-determined decline was found for the peak systolic/peak diastolic ratio. Fetal heart rate demonstrated a statistically significant increase relative to gestational age. However, the observed relation between the flow velocity wave form parameters, pulsatility index calculations, and gestational age was independent of fetal heart rate. It is speculated that peak diastolic velocity, diastolic integral, and peak systolic/peak diastolic ratio rather than the pulsatility index are useful in detecting gestational age-related changes in human fetal pulmonary vascular resistance.

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

    PubMed

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

    2016-06-01

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

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

    NASA Astrophysics Data System (ADS)

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

    2012-03-01

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

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

    PubMed

    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. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. [Pulmonary arterial hypertension associated with human immunodeficiency virus: thirty years after its description].

    PubMed

    Sandoval Gutiérrez, José Luis

    2018-04-11

    The development of pulmonary arterial hypertension associated with human immunodeficiency virus reduces the probability of survival in the patient affected compared to those without cardiopulmonary disease. The pathophysiology is uncertain. There are several lines of research to associate the different proteins of the virus in the endothelial lesion. From a therapeutic point of view there are treatment modalities that allow an acceptable life expectancy. Copyright © 2018 Instituto Nacional de Cardiología Ignacio Chávez. Publicado por Masson Doyma México S.A. All rights reserved.

  20. [Pulmonary cystic disease may be a rare complication to recurrent respiratory human papilloma virus infection].

    PubMed

    Laurberg, Peter Thaysen; Weinreich, Ulla M Øller

    2014-12-08

    A 19-year-old woman with a history of juvenile laryngeal papillomatosis (JLP), treated since childhood with multiple resections, was admitted with symptoms of pneumonia. A chest X-ray and CAT-scan revealed multiple lung cysts and a bronchoalveolar lavage detected human papilloma virus 11. The patient responded well to antibiotics. A body plethysmography showed small lung volumes and low diffusion capacity for carbon monoxide, but normal volume diffusion capacity divided by alveolar volume. Pulmonary cystic disease should be considered when patients with JLP have symptoms of pneumonia.

  1. SIRT1 is required for mitochondrial biogenesis reprogramming in hypoxic human pulmonary arteriolar smooth muscle cells

    PubMed Central

    Li, Pengyun; Liu, Yan; Burns, Nana; Zhao, Ke-Seng; Song, Rui

    2017-01-01

    Although recent studies have reported that mitochondria are putative oxygen sensors underlying hypoxic pulmonary vasoconstriction, little is known concerning the sirtuin 1 (SIRT1)-mediated mitochondrial biogenesis regulatory program in pulmonary arteriolar smooth muscle cells (PASMCs) during hypoxia/reoxygenation (H/R). We investigated the epigenetic regulatory mechanism of mitochondrial biogenesis and function in human PASMCs during H/R. Human PASMCs were exposed to hypoxia of 24–48 h and reoxygenation of 24–48 h. The expression of SIRT1 was reduced in a time-dependent manner. Mitochondrial transcription factor A (TFAM) expression was increased during hypoxia and decreased during reoxygenation, while the release of TFAM was increased in a time-dependent manner. Lentiviral overexpression of SIRT1 preserved SIRT3 deacetylase activity in human PASMCs exposed to H/R. Knockdown of PGC-1α suppressed the effect of SIRT1 on SIRT3 activity. Knockdown of SIRT3 abrogated SIRT1-mediated deacetylation of cyclophilin D (CyPD). Notably, knockdown of SIRT3 or PGC-1α suppressed the incremental effect of SIRT1 on mitochondrial TFAM, mitochondrial DNA (mtDNA) content and cellular ATP levels. Importantly, polydatin restored SIRT1 levels in human PASMCs exposed to H/R. Knockdown of SIRT1 suppressed the effect of polydatin on mitochondrial TFAM, mtDNA content and cellular ATP levels. In conclusion, SIRT1 expression is decreased in human PASMCs during H/R. TFAM expression in mitochondria is reduced and the release of TFAM is increased by H/R. PGC-1α/SIRT3/CyPD mediates the protective effect of SIRT1 on expression and release of TFAM and mitochondrial biogenesis and function. Polydatin improves mitochondrial biogenesis and function by enhancing SIRT1 expression in hypoxic human PASMCs. PMID:28339017

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

    NASA Astrophysics Data System (ADS)

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

    2015-10-01

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

  3. [Effect of Inhibiting NGAL Gene Expression on A549 Lung Cancer Cell Migration and Invasion].

    PubMed

    Tang, Jian; Li, Jie; Li, Shaojun; Li, Jingbo; Yu, Changhai; Wei, Chengze

    2015-04-01

    To detect the expression of neutrophil gelatinase-assoeiated lipocalin (NGAL) in the different differentiations of lung cancer tissues and to study the mechanism of invasion of A549 cells affected by NGAL. The expression of NGAL was detected by immunochemistry in lung cancer tissue and the tissue around edge of the cancer. The effect of NGAL expression on A549 cells was observed by using qRT-PCR and Western blot. The abilities of invasion and metastasis were evaluated by transwell invasion and migration assay, and cell scratch assay in vitro. The protein expression of E-cadherin, Vimentin was measured by immunofluoresence and Western blot. The positive expression rate of NGAL was 76.32% (58/76) in the lung cancer, 13.3% (4/30) in adjacent tissue by immunochemistry. NGAL expression levels in the lung cancer tissues were significantly higher than that in adjacent tissues. The rate of migration and invasion in NGAL-siRNA group was 60.4% ± 6.4% compared to 50.5% ± 4.4% in the control group, there was a significant difference (P < 0.05). Vimentin was suppressed, and E-cadherin was upregulated when NGAL was inhibited. MMP-2 and MMP-9 decreased when NGAL was knocked down. The expression level of NGAL is highly expressed in lung cancer. NGAL may be one of important indicators involved in lung cancer infiltrated and transferred. NGAL might be one of potential targets for lung cancer treatment.

  4. Role of Smac in apoptosis of lung cancer cells A549 induced by Taxol.

    PubMed

    Zhang, Ying; Hao, Yingtao; Sun, Qifeng; Peng, Chuanliang

    2015-01-01

    A series of structurally unique second mitochondria-derived activators of caspase (Smac) that act as antagonists of inhibitor of apoptosis proteins (IAPs) directly have been discovered and have been shown to promote chemotherapy-induced apoptosis. In this study, we investigate the role of Smac in Taxol-induced apoptosis of lung cancer cell in vitro. PcDNA3.1/Smac recombinants were transfected into the non-small cell lung cancer cell line A549. Smac expression was detected by RT-PCR and Western blot. The invasive ability of cells was examined. Flow cytometry was used to analyze apoptosis of cells induced by Taxol with Annexin V/PI double staining technique. Smac expression was significantly higher in the PcDNA3.1/Smac recombinant group than in the untransfected group at mRNA and protein level (p < 0.05) and lower invasion through a basal membrane was apparent after transfection (p < 0.05). A small number of cells were promoted to apoptosis in the PcDNA3.1/Smac group. There were significant differences compared to the empty vector group and control group. The apoptosis rate was significantly higher in PcDNA3.1/Smac + Taxol group than in other groups (p < 0.05). Transfected Smac can enhance the chemosensitivity of the non-small cell lung cancer cell line A549 to Taxol.

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

    PubMed

    Ghosh, Somnath; Narang, Himanshi; Sarma, Asitikantha; Krishna, Malini

    2011-11-01

    Carbon beams (5.16MeV/u, LET=290keV/μm) 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 γ-rays and carbon ion-irradiation. A549 cells were irradiated with 1Gy carbon or γ-rays. Carbon beam was found to be three times more cytotoxic than γ-rays despite the fact that the numbers of γ-H2AX foci were same. Percentage of cells showing ATM/ATR foci were more with γ-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 γ-rays irradiated cells and prosurvival ERK pathway was activated after γ-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. Copyright © 2011 Elsevier B.V. All rights reserved.

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

    PubMed

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

    2017-09-01

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

  7. Krüppel-like factor 5 contributes to pulmonary artery smooth muscle proliferation and resistance to apoptosis in human pulmonary arterial hypertension.

    PubMed

    Courboulin, Audrey; Tremblay, Véronique L; Barrier, Marjorie; Meloche, Jolyane; Jacob, Maria Helena; Chapolard, Mathilde; Bisserier, Malik; Paulin, Roxane; Lambert, Caroline; Provencher, Steeve; Bonnet, Sébastien

    2011-09-27

    Pulmonary arterial hypertension (PAH) is a vascular remodeling disease characterized by enhanced proliferation of pulmonary artery smooth muscle cell (PASMC) and suppressed apoptosis. This phenotype has been associated with the upregulation of the oncoprotein survivin promoting mitochondrial membrane potential hyperpolarization (decreasing apoptosis) and the upregulation of growth factor and cytokines like PDGF, IL-6 and vasoactive agent like endothelin-1 (ET-1) promoting PASMC proliferation. Krüppel-like factor 5 (KLF5), is a zinc-finger-type transcription factor implicated in the regulation of cell differentiation, proliferation, migration and apoptosis. Recent studies have demonstrated the implication of KLF5 in tissue remodeling in cardiovascular diseases, such as atherosclerosis, restenosis, and cardiac hypertrophy. Nonetheless, the implication of KLF5 in pulmonary arterial hypertension (PAH) remains unknown. We hypothesized that KLF5 up-regulation in PAH triggers PASMC proliferation and resistance to apoptosis. We showed that KFL5 is upregulated in both human lung biopsies and cultured human PASMC isolated from distal pulmonary arteries from PAH patients compared to controls. Using stimulation experiments, we demonstrated that PDGF, ET-1 and IL-6 trigger KLF-5 activation in control PASMC to a level similar to the one seen in PAH-PASMC. Inhibition of the STAT3 pathway abrogates KLF5 activation in PAH-PASMC. Once activated, KLF5 promotes cyclin B1 upregulation and promotes PASMC proliferation and triggers survivin expression hyperpolarizing mitochondria membrane potential decreasing PASMC ability to undergo apoptosis. We demonstrated for the first time that KLF5 is activated in human PAH and implicated in the pro-proliferative and anti-apoptotic phenotype that characterize PAH-PASMC. We believe that our findings will open new avenues of investigation on the role of KLF5 in PAH and might lead to the identification of new therapeutic targets.

  8. Krüppel-like Factor 5 contributes to pulmonary artery smooth muscle proliferation and resistance to apoptosis in human pulmonary arterial hypertension

    PubMed Central

    2011-01-01

    Background Pulmonary arterial hypertension (PAH) is a vascular remodeling disease characterized by enhanced proliferation of pulmonary artery smooth muscle cell (PASMC) and suppressed apoptosis. This phenotype has been associated with the upregulation of the oncoprotein survivin promoting mitochondrial membrane potential hyperpolarization (decreasing apoptosis) and the upregulation of growth factor and cytokines like PDGF, IL-6 and vasoactive agent like endothelin-1 (ET-1) promoting PASMC proliferation. Krüppel-like factor 5 (KLF5), is a zinc-finger-type transcription factor implicated in the regulation of cell differentiation, proliferation, migration and apoptosis. Recent studies have demonstrated the implication of KLF5 in tissue remodeling in cardiovascular diseases, such as atherosclerosis, restenosis, and cardiac hypertrophy. Nonetheless, the implication of KLF5 in pulmonary arterial hypertension (PAH) remains unknown. We hypothesized that KLF5 up-regulation in PAH triggers PASMC proliferation and resistance to apoptosis. Methods and results We showed that KFL5 is upregulated in both human lung biopsies and cultured human PASMC isolated from distal pulmonary arteries from PAH patients compared to controls. Using stimulation experiments, we demonstrated that PDGF, ET-1 and IL-6 trigger KLF-5 activation in control PASMC to a level similar to the one seen in PAH-PASMC. Inhibition of the STAT3 pathway abrogates KLF5 activation in PAH-PASMC. Once activated, KLF5 promotes cyclin B1 upregulation and promotes PASMC proliferation and triggers survivin expression hyperpolarizing mitochondria membrane potential decreasing PASMC ability to undergo apoptosis. Conclusion We demonstrated for the first time that KLF5 is activated in human PAH and implicated in the pro-proliferative and anti-apoptotic phenotype that characterize PAH-PASMC. We believe that our findings will open new avenues of investigation on the role of KLF5 in PAH and might lead to the identification of

  9. Study of Bioreductive Anticancer Agent RH-1-Induced Signals Leading the Wild-Type p53-Bearing Lung Cancer A549 Cells to Apoptosis.

    PubMed

    Stulpinas, Aurimas; Imbrasaitė, Aušra; Krestnikova, Natalija; Šarlauskas, Jonas; Čėnas, Narimantas; Kalvelytė, Audronė Valerija

    2016-01-19

    Aziridinylquinone RH-1 (2,5-diaziridinyl-3-hydroxymethyl-6-methyl-cyclohexa-2,5-diene-1,4-dione) is a potential anticancer agent. RH-1 action is associated with quinone oxidoreductase (NQO1) which reduces this diaziridinylbenzoquinone into DNA-alkylating hydroquinone and is overexpressed in many tumors. Another suggested mechanism of RH-1 toxicity is the formation of reactive oxygen species (ROS) arising from its redox cycling. In order to improve anticancer action of this and similar antitumor quinones, we investigated the involvement of different signaling molecules in cytotoxicity induced by RH-1 by using wild-type tumor suppressor p53 bearing nonsmall cell lung carcinoma A549 cells as a model. Gradual and prolonged increase of mitogen-activated protein kinases (MAPK) ERK, P38, and JNK phosphorylation was observed during 24-h RH-1 treatment. In parallel, activation of DNA damage-sensing ATM kinase, upregulation, and phosphorylation of TP53 (human p53) took place. Inhibition studies revealed that RH-1-induced A549 apoptosis involved the NQO1-ATM-p53 signaling pathway and ROS generation. TP53 participated in ROS- and DNA damage-induced cell death differently. Moreover, MAP kinase JNK was another TP53 activator and death inducer in A549 cells. At the same time, rapid and prolonged activation of AKT kinase during RH-1 treatment was found, and it proved to be antiapoptotic kinase in our model system. Therefore, we identified that different and opposite cell death regulating signaling pathways, which may counteract one another, are induced in cancer cells during chemotherapeutic RH-1 treatment.

  10. Identification of curcumin-inhibited extracellular matrix receptors in non-small cell lung cancer A549 cells by RNA sequencing.

    PubMed

    Li, Huiping; Wu, Hongjin; Zhang, Hongfang; Li, Ying; Li, Shuang; Hou, Qiang; Wu, Shixiu; Yang, Shuan-Ying

    2017-06-01

    Curcumin is a potent anti-cancer drug in several types of human cancers. Despite of several preclinical and clinical studies of curcumin, the precise mechanism of curcumin in cancer prevention has remained unclear. In our study, we for the first time investigated whole transcriptome alteration in A549 non-small cell lung cancer (NSCLC) cell lines after treatment with curcumin using RNA sequencing. We found that lots of genes and signaling pathways were significantly altered after curcumin treatment in A549 cells. With bioinformatics approaches (gene ontology, Kyoto Encyclopedia of Genes and Genomes, and STRING), we found that those curcumin altered genes were not only the genes that induce cell death but also those extracellular matrix receptors and mitogen-activated protein kinase signaling pathway genes which regulate cell migration and proliferation. Among those significantly altered genes, eight genes ( COL1A1, COL4A1, COL5A1, LAMA5, ITGA3, ITGA2B, DDIT3, and DUSP1) were further examined by quantitative reverse transcription polymerase chain reaction and western blot analysis in four non-small cell lung cancer cell lines. Both in cell lines and in mouse model, the extracellular matrix receptors including the integrin ( ITGA3 and ITGA2B), collagen ( COL5A1), and laminin ( LAMA5) were significantly inhibited by curcumin at messenger RNA and protein levels. Functional studies confirmed that curcumin not only induced A549 cell death but also repressed cell proliferation and migration by regulating extracellular matrix receptors. Collectively, our study suggests that curcumin may be used as a promising drug candidate for intervening lung cancer in future studies.

  11. α9- and α7-containing receptors mediate the pro-proliferative effects of nicotine in the A549 adenocarcinoma cell line.

    PubMed

    Mucchietto, Vanessa; Fasoli, Francesca; Pucci, Susanna; Moretti, Milena; Benfante, Roberta; Maroli, Annalisa; Di Lascio, Simona; Bolchi, Cristiano; Pallavicini, Marco; Dowell, Cheryl; McIntosh, Michael; Clementi, Francesco; Gotti, Cecilia

    2017-07-20

    Tobacco smoke contains many classes of carcinogens and although nicotine is unable to initiate tumourigenesis in humans and rodents, it promotes tumour growth and metastasis in lung tumours by acting on neuronal nicotinic ACh receptors (nAChRs). The aim of this study was to identify molecularly, biochemically and pharmacologically which nAChR subtypes are expressed and functionally activated by nicotine in lung cancer cell lines. We used A549 and H1975 adenocarcinoma cell lines derived from lung tumours to test the in vitro effects of nicotine, and nAChR subtype-specific peptides and compounds. The two adenocarcinoma cell lines express distinctive nAChR subtypes, and this affects their nicotine-induced proliferation. In A549 cells, nAChRs containing the α7 or α9 subunits not only regulate nicotine-induced cell proliferation but also the activation of the Akt and ERK pathways. Blocking these nAChRs by means of subtype-specific peptides, or silencing their expression by means of subunit-specific siRNAs, abolishes nicotine-induced proliferation and signalling. Moreover, we found that the α7 antagonist MG624 also acts on α9-α10 nAChRs, blocks the effects of nicotine on A549 cells and has dose-dependent cytotoxic activity. These results highlight the pathophysiological role of α7- and α9-containing receptors in promoting non-small cell lung carcinoma cell growth and intracellular signalling and provide a framework for the development of new drugs that specifically target the receptors expressed in lung tumours. © 2017 The British Pharmacological Society.

  12. [The stimulation of human pulmonary artery endothelial cells by cigarette smoke extract contributed to cell senescence and induced human pulmonary artery smooth cell migration].

    PubMed

    Cai, L; Zhu, P C; Wang, Y E; Gao, Y T; Ao, Q L

    2017-06-12

    Objective: To observe the senescent effect of human pulmonary arterial endothelial cells (HPAEC) stimulated by cigarette smoke extract (CSE) and the effect of secretion of senescent cells on human pulmonary arterial smooth muscles cell (HPASMC) proliferation and migration. Methods: HPAEC was treated with different concentrations of CSE in vitro and cell proliferation was determined by CCK8, senescence cells analyzed by detecting the β-gal activity, and the senescent proteins of cells measured by Western blot. The concentration of senescence-associated secretory phenotype (SASP) was detected by ELISA and the expression of MCP-1 and TGF-β1 was measured by Real-time PCR. The number of the proliferated cells was measured by Transwell assay and immunoflurescence. Results: The HPAEC was aging with the stimulation concentration of CSE increasing and the stimulation time prolonging ( P <0.05). Western blot indicated that the senescent associated protein p53 or p21 increased markedly after 48 h and 72 h CSE-exposure ( n =3, P <0.05). The SA-β-Gal staining showed that the number of senescent cells increased as the exposure time prolonged. Compared with the control group, cell viability of 48 h group(1.8±0.1) and 72 h group (1.8±0.1) decreased significantly. The flow cytometry showed a significant difference between the CSE group(14.1±1.2) and the control group(28.5±1.8) in S phase( P <0.01), indicating cell cycle arrest. The SASP was increasing as the CSE-exposure prolonged. Compared with the control group(177±39), the 48 h group(460±43) and the 72 h group(609±64) showed a marked increase in MCP-1( P <0.05). For TGF-β1, it had a same tendency and a significant difference between the control group(121±18) and the 48 h group(413±32) or 72 h group(606±67, both P <0.05). In the meantime, the bFGF increased after 48 h stimulation(291±13, P <0.05). Besides MCP-1, TGF-β1 showed a significant difference between the control group and the 72 h CSE-exposure group ( P <0

  13. Neutrophil-Derived MMP-8 Drives AMPK-Dependent Matrix Destruction in Human Pulmonary Tuberculosis.

    PubMed

    Ong, Catherine W M; Elkington, Paul T; Brilha, Sara; Ugarte-Gil, Cesar; Tome-Esteban, Maite T; Tezera, Liku B; Pabisiak, Przemyslaw J; Moores, Rachel C; Sathyamoorthy, Tarangini; Patel, Vimal; Gilman, Robert H; Porter, Joanna C; Friedland, Jon S

    2015-05-01

    Pulmonary cavities, the hallmark of tuberculosis (TB), are characterized by high mycobacterial load and perpetuate the spread of M. tuberculosis. The mechanism of matrix destruction resulting in cavitation is not well defined. Neutrophils are emerging as key mediators of TB immunopathology and their influx are associated with poor outcomes. We investigated neutrophil-dependent mechanisms involved in TB-associated matrix destruction using a cellular model, a cohort of 108 patients, and in separate patient lung biopsies. Neutrophil-derived NF-kB-dependent matrix metalloproteinase-8 (MMP-8) secretion was up-regulated in TB and caused matrix destruction both in vitro and in respiratory samples of TB patients. Collagen destruction induced by TB infection was abolished by doxycycline, a licensed MMP inhibitor. Neutrophil extracellular traps (NETs) contain MMP-8 and are increased in samples from TB patients. Neutrophils lined the circumference of human pulmonary TB cavities and sputum MMP-8 concentrations reflected TB radiological and clinical disease severity. AMPK, a central regulator of catabolism, drove neutrophil MMP-8 secretion and neutrophils from AMPK-deficient patients secrete lower MMP-8 concentrations. AMPK-expressing neutrophils are present in human TB lung biopsies with phospho-AMPK detected in nuclei. These data demonstrate that neutrophil-derived MMP-8 has a key role in the immunopathology of TB and is a potential target for host-directed therapy in this infectious disease.

  14. Fenugreek extract diosgenin and pure diosgenin inhibit the hTERT gene expression in A549 lung cancer cell line.

    PubMed

    Rahmati-Yamchi, Mohammad; Ghareghomi, Somayyeh; Haddadchi, Gholamreza; Milani, Morteza; Aghazadeh, Mohammad; Daroushnejad, Hasan

    2014-09-01

    Trigonella foenum-graecum generally known as fenugreek, has been normally cultivated in Asia and Africa for the edible and medicinal values of its seeds. Fenugreek leaves and seeds have been used widely for therapeutic purposes. Fenugreek seed is recognized to show anti-diabetic and anti-nociceptive properties and other things such as hypocholesterolaemic, and anti-cancer. Diosgenin is a steroidal saponin from therapeutic herbs, fenugreek (T. foenum-graceum L.), has been well-known to have anticancer properties. Telomerase activity is not identified in usual healthy cells, while in carcinogenic cell telomerase expression is reactivated. Therefore telomerase illustrates a promising cancer therapeutic target. We deliberate the inhibitory effect of pure diosgenin and fenugreek extract diosgenin on human telomerase reverse transcriptase gene (hTERT) expression which is critical for telomerase activity. MTT-assay and qRT-PCR analysis were achieved to discover cytotoxicity effects and hTERT gene expression inhibition properties, separately. MTT results exhibited that IC50 for pure diosgenin were 47, 44 and 43 µM and for fenugreek extract diosgenin were 49, 48 and 47 µM for 24, 48 and 72 h after treatment. Culturing cells with pure diosgenin and fenugreek extract diosgenin treatment caused in down regulation of hTERT expression. These results indication that pure and impure diosgenin prevents telomerase activity by down regulation of the hTERT gene expression in A549 lung cancer cell line, with the difference that pure compound is more effective than another.

  15. Deletion of Adipose Triglyceride Lipase Links Triacylglycerol Accumulation to a More-Aggressive Phenotype in A549 Lung Carcinoma Cells.

    PubMed

    Tomin, Tamara; Fritz, Katarina; Gindlhuber, Juergen; Waldherr, Linda; Pucher, Bettina; Thallinger, Gerhard G; Nomura, Daniel K; Schittmayer, Matthias; Birner-Gruenberger, Ruth

    2018-04-06

    Adipose triglyceride lipase (ATGL) catalyzes the rate limiting step in triacylglycerol breakdown in adipocytes but is expressed in most tissues. The enzyme was shown to be lost in many human tumors, and its loss may play a role in early stages of cancer development. Here, we report that loss of ATGL supports a more-aggressive cancer phenotype in a model system in which ATGL was deleted in A549 lung cancer cells by CRISPR/Cas9. We observed that loss of ATGL led to triacylglycerol accumulation in lipid droplets and higher levels of cellular phospholipid and bioactive lipid species (lyso- and ether-phospholipids). Label-free quantitative proteomics revealed elevated expression of the pro-oncogene SRC kinase in ATGL depleted cells, which was also found on mRNA level and confirmed on protein level by Western blot. Consistently, higher expression of phosphorylated (active) SRC (Y416 phospho-SRC) was observed in ATGL-KO cells. Cells depleted of ATGL migrated faster, which was dependent on SRC kinase activity. We propose that loss of ATGL may thus increase cancer aggressiveness by activation of pro-oncogenic signaling via SRC kinase and increased levels of bioactive lipids.

  16. Rapid induction and persistence of paracrine-induced cellular antiviral states arrest viral infection spread in A549 cells.

    PubMed

    Voigt, Emily A; Swick, Adam; Yin, John

    2016-09-01

    The virus/host interaction is a complex interplay between pro- and anti-viral factors that ultimately determines the spread or halt of virus infections in tissues. This interplay develops over multiple rounds of infection. The purpose of this study was to determine how cellular-level processes combine to impact the spatial spread of infection. We measured the kinetics of virus replication (VSV), antiviral paracrine signal upregulation and secretion, spatial spread of virus and paracrine antiviral signaling, and inhibition of virus production in antiviral-exposed A549 human lung epithelial cells. We found that initially infected cells released antiviral signals 4-to-7h following production of virus. However, the subsequent rapid dissemination of signal and fast induction of a robust and persistent antiviral state ultimately led to a suppression of infection spread. This work shows how cellular responses to infection and activation of antiviral responses can integrate to ultimately control infection spread across host cell populations. Copyright © 2016. Published by Elsevier Inc.

  17. Delphinidin inhibits angiogenesis through the suppression of HIF-1α and VEGF expression in A549 lung cancer cells.

    PubMed

    Kim, Mun-Hyeon; Jeong, Yun-Jeong; Cho, Hyun-Ji; Hoe, Hyang-Sook; Park, Kwan-Kyu; Park, Yoon-Yub; Choi, Yung Hyun; Kim, Cheorl-Ho; Chang, Hyeun-Wook; Park, Young-Ja; Chung, Il-Kyung; Chang, Young-Chae

    2017-02-01

    Delphinidin, a polyphenol that belongs to the group of anthocyanidins and is abundant in many pigmented fruits and vegetables, possesses important antioxidant, anti‑inflammatory, anti-mutagenic and anticancer properties. In the present study, we investigated the inhibitory effects of delphinidin on vascular endothelial growth factor (VEGF) expression, an important factor involved in angiogenesis and tumor progression, in A549 human lung cancer cells. Delphinidin inhibited CoCl2- and epidermal growth factor (EGF)-induced VEGF mRNA expression and VEGF protein production. Delphinidin also decreased CoCl2- and EGF-stimulated expression of hypoxia‑inducible factor (HIF)‑1α, which is a transcription factor of VEGF. Delphinidin suppressed CoCl2- and EGF-induced hypoxia‑response element (HRE) promoter activity, suggesting that the inhibitory effects of delphinidin on VEGF expression are caused by the suppression of the binding of HIF-1 to the HRE promoter. We also found that delphinidin specifically decreased the CoCl2- and EGF-induced HIF-1α protein expression by blocking the ERK and PI3K/Akt/mTOR/p70S6K signaling pathways, whereas the p38-mediated pathways were not involved. In animal models, EGF-induced new blood vessel formation was significantly inhibited by delphinidin. Therefore, our results indicate that delphinidin has a potentially new role in anti‑angiogenic action by inhibiting HIF-1α and VEGF expression.

  18. Bio-fabrication of catalytic platinum nanoparticles and their in vitro efficacy against lungs cancer cells line (A549).

    PubMed

    Ullah, Sadeeq; Ahmad, Aftab; Wang, Aoke; Raza, Muslim; Jan, Amin Ullah; Tahir, Kamran; Rahman, Aziz Ur; Qipeng, Yuan

    2017-08-01

    Platinum based drugs are considered as effective agents against various types of carcinoma; however, the severe toxicity associated with the chemically prepared platinum complexes limit their practical applications. Similarly, water pollution caused by various organic moieties is another serious health problem worldwide. Hence, an intense need exists to develop new, effective and biocompatible materials with catalytic and biomedical applications. In the present contribution, we prepared platinum nanoparticles (PtNPs) by a green route using phytochemicals as a source of reducing and stabilizing agents. Well dispersed and crystalline PtNPs of spherical shapes were prepared and characterized. The bio-fabricated PtNPs were used as catalyst and anticancer agents. Catalytic performance of the PtNPs showed that 84% of the methylene blue can be reduced in 32min under visible light irradiation (K=0.078min -1 ). Similarly the catalytic conversion of 4-nitrophenol to 4-aminophenol was achieved in <20min (K=0.124min -1 ). The in vitro anticancer study revealed that biogenic PtNPs are the efficient nano-agents possessing strong anticancer activity against the lungs cancer cells line (A549). Interestingly, the as prepared PtNPs were well tolerated by normal human cells, and therefore, could be effective and biocompatible agents in the treatment of different cancer cells. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    NASA Astrophysics Data System (ADS)

    Di Bucchianico, Sebastiano; Migliore, Lucia; Marsili, Paolo; Vergari, Chiara; Giammanco, Francesco; Giorgetti, Emilia

    2015-05-01

    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.

  20. Activity of interferon alpha, interleukin 6 and insulin in the regulation of differentiation in A549 alveolar carcinoma cells.

    PubMed

    McCormick, C; Freshney, R I; Speirs, V

    1995-02-01

    The differentiation of A549, a human tumour cell line from type II pneumocytes, can be induced by a crude fibroblast-derived factor (FDF) isolated from the conditioned medium of glucocorticoid-treated lung fibroblasts. In the present report, we have used alkaline phosphatase as a differentiation marker to investigate the activity of a number of growth factors as potential candidates for this paracrine activity. This showed that insulin, interleukin 6 (IL-6), and interferon alpha (IFN-alpha) could simulate the activity of conditioned medium. Their effects were dexamethasone (DX) dependent, additive and reversible with a half-life of 1 week. Transforming growth factor alpha and beta, IL-1 alpha and epidermal growth factor, were all inhibitory, and inhibition was opposed, partially or completely, by DX. The most potent inducer was IL-6, but as DX was shown to decrease the concentration of IL-6 in lung fibroblast-conditioned medium it seems an unlikely candidate for FDF. Unlike FDF, all of the positive-acting factors were shown to induce plasminogen activator. FDF has also been shown to be active in the absence of DX. This suggests that differentiation-inducing activity may be present in several paracrine factors, but that so far a candidate for FDF has not been identified.

  1. Activity of interferon alpha, interleukin 6 and insulin in the regulation of differentiation in A549 alveolar carcinoma cells.

    PubMed Central

    McCormick, C.; Freshney, R. I.; Speirs, V.

    1995-01-01

    The differentiation of A549, a human tumour cell line from type II pneumocytes, can be induced by a crude fibroblast-derived factor (FDF) isolated from the conditioned medium of glucocorticoid-treated lung fibroblasts. In the present report, we have used alkaline phosphatase as a differentiation marker to investigate the activity of a number of growth factors as potential candidates for this paracrine activity. This showed that insulin, interleukin 6 (IL-6), and interferon alpha (IFN-alpha) could simulate the activity of conditioned medium. Their effects were dexamethasone (DX) dependent, additive and reversible with a half-life of 1 week. Transforming growth factor alpha and beta, IL-1 alpha and epidermal growth factor, were all inhibitory, and inhibition was opposed, partially or completely, by DX. The most potent inducer was IL-6, but as DX was shown to decrease the concentration of IL-6 in lung fibroblast-conditioned medium it seems an unlikely candidate for FDF. Unlike FDF, all of the positive-acting factors were shown to induce plasminogen activator. FDF has also been shown to be active in the absence of DX. This suggests that differentiation-inducing activity may be present in several paracrine factors, but that so far a candidate for FDF has not been identified. PMID:7841035

  2. Deguelin inhibits the migration and invasion of lung cancer A549 and H460 cells via regulating actin cytoskeleton rearrangement.

    PubMed

    Zhao, Honggang; Jiao, Yan; Zhang, Zuncheng

    2015-01-01

    Deguelin, the main components from Mundulea sericea, was reported to suppress the growth of various cancer cells. However, the effect of Deguelin on tumor cell invasion and metastasis and its mechanism still unclear so far. In this study, we investigated the effects of Deguelin on the cell invasion in human lung cancer A549 and H460 cells. Our results demonstrate that Deguelin can significantly inhibited cell proliferation, cell migration and cell invasion. Moreover, Deguelin could also affected reorganization of the actin cytoskeleton and decreased filopodia and lamellipodia formation. Furthermore, deguelin-treated tumors showed decreased the tumor metastasis related genes such as CD44, MMP2 and MMP9 at protein and mRNA levels and the content of CEA, SCC, NSE, CYFAR21-1. In addition, Deguelin down-regulated protein expression of Rac1 and Rock1, which are impotent in actin cytoskeleton rearrangements and cell motility. Together, our results suggest that Deguelin inhibit tumor growth and metastasis of lung cancer cells and might be a candidate compound for curing lung cancer.

  3. [Effects of airborne fine particulate matter on human respiratory symptoms and pulmonary function].

    PubMed

    Gao, Zhi-Yi; Li, Peng-Kun; Zhao, Jin-Zhuo; Jiang, Rong-Fang; Yang, Bin-Jie; Zhang, Min-Hua; Song, Wei-Min

    2010-10-01

    to explore effects of airborne fine particulate matter exposure on human respiratory symptoms and pulmonary function. one hundred and seven field traffic policemen were recruited as airborne fine particulate matter high-exposure group and one hundred and one male residents as common exposure group. The individual sampler was used to measure fine particulate matter exposure levels of the two groups. To obtain personal information, especially respiratory symptoms such as cough, sputum, etc. a questionnaire survey was used. The pulmonary ventilation function was detected: forced expiratory vital capacity (FVC), the first 1 second forced expiratory volume (FEV1.0), FVC/FEV1.0% and peak flow values (PEF), and the difference of fine particulate matter exposure level and respiratory function of the two groups was compared. 24 h individual average fine particulate matter exposure concentration of traffic police and residents were respectively (115.4 ± 46.17) microg/m(3) and (74.94 ± 40.09) microg/m(3), the traffic police PM2.5 exposure levels were significantly higher than the residents. In the incidence of respiratory symptoms, compared with high-exposure group and common exposure group, coughing, expectoration, throat unwell, asthma, short of breath and nose discomfort, traffic police group was higher than residents group (P < 0.05). The abnormal rate of lung ventilation function indexes, such as FVC and FEV1.0 was 25.23% and 12.15% respectively in high-exposure group, 11.88% and 2.97% in common exposure group, there was no statistical difference between two groups. Besides, the abnormal rate of FVC and FEV1.0, showed rising trend in high-exposure group with seniority. long-term higher levels of airborne fine particulate matter exposure, may impact respiratory health and impair pulmonary function.

  4. Haemophilus influenzae genome evolution during persistence in the human airways in chronic obstructive pulmonary disease.

    PubMed

    Pettigrew, Melinda M; Ahearn, Christian P; Gent, Janneane F; Kong, Yong; Gallo, Mary C; Munro, James B; D'Mello, Adonis; Sethi, Sanjay; Tettelin, Hervé; Murphy, Timothy F

    2018-04-03

    Nontypeable Haemophilus influenzae (NTHi) exclusively colonize and infect humans and are critical to the pathogenesis of chronic obstructive pulmonary disease (COPD). In vitro and animal models do not accurately capture the complex environments encountered by NTHi during human infection. We conducted whole-genome sequencing of 269 longitudinally collected cleared and persistent NTHi from a 15-y prospective study of adults with COPD. Genome sequences were used to elucidate the phylogeny of NTHi isolates, identify genomic changes that occur with persistence in the human airways, and evaluate the effect of selective pressure on 12 candidate vaccine antigens. Strains persisted in individuals with COPD for as long as 1,422 d. Slipped-strand mispairing, mediated by changes in simple sequence repeats in multiple genes during persistence, regulates expression of critical virulence functions, including adherence, nutrient uptake, and modification of surface molecules, and is a major mechanism for survival in the hostile environment of the human airways. A subset of strains underwent a large 400-kb inversion during persistence. NTHi does not undergo significant gene gain or loss during persistence, in contrast to other persistent respiratory tract pathogens. Amino acid sequence changes occurred in 8 of 12 candidate vaccine antigens during persistence, an observation with important implications for vaccine development. These results indicate that NTHi alters its genome during persistence by regulation of critical virulence functions primarily by slipped-strand mispairing, advancing our understanding of how a bacterial pathogen that plays a critical role in COPD adapts to survival in the human respiratory tract.

  5. A human disease model of drug toxicity-induced pulmonary edema in a lung-on-a-chip microdevice.

    PubMed

    Huh, Dongeun; Leslie, Daniel C; Matthews, Benjamin D; Fraser, Jacob P; Jurek, Samuel; Hamilton, Geraldine A; Thorneloe, Kevin S; McAlexander, Michael Allen; Ingber, Donald E

    2012-11-07

    Preclinical drug development studies currently rely on costly and time-consuming animal testing because existing cell culture models fail to recapitulate complex, organ-level disease processes in humans. We provide the proof of principle for using a biomimetic microdevice that reconstitutes organ-level lung functions to create a human disease model-on-a-chip that mimics pulmonary edema. The microfluidic device, which reconstitutes the alveolar-capillary interface of the human lung, consists of channels lined by closely apposed layers of human pulmonary epithelial and endothelial cells that experience air and fluid flow, as well as cyclic mechanical strain to mimic normal breathing motions. This device was used to reproduce drug toxicity-induced pulmonary edema observed in human cancer patients treated with interleukin-2 (IL-2) at similar doses and over the same time frame. Studies using this on-chip disease model revealed that mechanical forces associated with physiological breathing motions play a crucial role in the development of increased vascular leakage that leads to pulmonary edema, and that circulating immune cells are not required for the development of this disease. These studies also led to identification of potential new therapeutics, including angiopoietin-1 (Ang-1) and a new transient receptor potential vanilloid 4 (TRPV4) ion channel inhibitor (GSK2193874), which might prevent this life-threatening toxicity of IL-2 in the future.

  6. Role for miR-204 in human pulmonary arterial hypertension

    PubMed Central

    Courboulin, Audrey; Paulin, Roxane; Giguère, Nellie J.; Saksouk, Nehmé; Perreault, Tanya; Meloche, Jolyane; Paquet, Eric R.; Biardel, Sabrina; Provencher, Steeve; Côté, Jacques; Simard, Martin J.

    2011-01-01

    Pulmonary arterial hypertension (PAH) is characterized by enhanced proliferation and reduced apoptosis of pulmonary artery smooth muscle cells (PASMCs). Because microRNAs have been recently implicated in the regulation of cell proliferation and apoptosis, we hypothesized that these regulatory molecules might be implicated in the etiology of PAH. In this study, we show that miR-204 expression in PASMCs is down-regulated in both human and rodent PAH. miR-204 down-regulation correlates with PAH severity and accounts for the proliferative and antiapoptotic phenotypes of PAH-PASMCs. STAT3 activation suppresses miR-204 expression, and miR-204 directly targets SHP2 expression, thereby SHP2 up-regulation, by miR-204 down-regulation, activates the Src kinase and nuclear factor of activated T cells (NFAT). STAT3 also directly induces NFATc2 expression. NFAT and SHP2 were needed to sustain PAH-PASMC proliferation and resistance to apoptosis. Finally, delivery of synthetic miR-204 to the lungs of animals with PAH significantly reduced disease severity. This study uncovers a new regulatory pathway involving miR-204 that is critical to the etiology of PAH and indicates that reestablishing miR-204 expression should be explored as a potential new therapy for this disease. PMID:21321078

  7. Dissociation between systemic and pulmonary anti‐inflammatory effects of dexamethasone in humans

    PubMed Central

    Bartko, Johann; Stiebellehner, Leopold; Derhaschnig, Ulla; Schoergenhofer, Christian; Schwameis, Michael; Prosch, Helmut

    2016-01-01

    Aims The local pulmonary inflammatory response has a different temporal and qualitative profile compared with the systemic inflammatory response. Although glucocorticoids substantially downregulate the systemic release of acute‐phase mediators, it is not clear whether they have comparable inhibitory effects in the human lung compartment. Therefore, we compared the anti‐inflammatory effects of a pure glucocorticoid agonist, dexamethasone, on bronchoalveolar lavage and blood cytokine concentrations in response to bronchially instilled endotoxin. Methods In this randomized, double‐blind and placebo‐controlled trial, 24 volunteers received dexamethasone or placebo and had endotoxin instilled into a lung segment and saline instilled into a contralateral segment, followed by bronchoalveolar lavage. Results Bronchially instilled endotoxin induced a local and systemic inflammatory response. Dexamethasone strongly blunted the systemic interleukin (IL) 6 and C‐reactive protein release. In sharp contrast, dexamethasone left the local release of acute‐phase mediators in the lungs virtually unchanged: bronchoalveolar lavage levels of IL‐6 were only 18% lower and levels of IL‐8 were even higher with dexamethasone compared with placebo, although the differences between treatments were not statistically significant (P = 0.07 and P = 0.08, respectively). However, dexamethasone had inhibitory effects on pulmonary protein extravasation and neutrophil migration. Conclusions The present study demonstrated a remarkable dissociation between the systemic anti‐inflammatory effects of glucocorticoids and its protective effects on capillary leak on the one hand and surprisingly low anti‐inflammatory effects in the lungs on the other. PMID:26647918

  8. Non-ionic surfactant vesicles in pulmonary glucocorticoid delivery: characterization and interaction with human lung fibroblasts.

    PubMed

    Marianecci, Carlotta; Paolino, Donatella; Celia, Christian; Fresta, Massimo; Carafa, Maria; Alhaique, Franco

    2010-10-01

    Non-ionic surfactant vesicles (NSVs) were proposed for the pulmonary delivery of glucocorticoids such as beclomethasone dipropionate (BDP) for the treatment of inflammatory lung diseases, e.g. asthma, chronic obstructive pulmonary disease and various type of pulmonary fibrosis. The thin layer evaporation method followed by sonication was used to prepare small non-ionic surfactant vesicles containing beclomethasone dipropionate. Light scattering experiments showed that beclomethasone dipropionate-loaded non-ionic surfactant vesicles were larger than unloaded ones and showed a significant (P<0.001) decrease of the zeta potential. The morphological analysis, by freeze-fracture transmission electron microscopy, showed the maintenance of a vesicular structure in the presence of the drug. The colloidal and storage stability were evaluated by Turbiscan Lab Expert, which evidenced the good stability of BDP-loaded non-ionic surfactant vesicles, thus showing no significant variations of mean size and no colloidal phase segregation. Primary human lung fibroblast (HLF) cells were used for in vitro investigation of vesicle tolerability, carrier-cell interaction, intracellular drug uptake and drug-loaded vesicle anti-inflammatory activity. The investigated NSVs did not show a significant cytotoxic activity at all incubation times for concentrations ranging from 0.01 to 1 μM. Confocal laser scanning microscopy showed vesicular carrier localization at the level of the cytoplasm compartment, where the glucocorticoid receptor (target site) is localized. BDP-loaded non-ionic surfactant vesicles elicited a significant improvement of the HLF intracellular uptake of the drug with respect to the free drug solution, drug/surfactant mixtures and empty vesicles used as references. The treatment of HLF cells with BDP-loaded non-ionic surfactant vesicles determined a noticeable increase of the drug anti-inflammatory activity by reducing the secretion of both constitutive and interleukin-1

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

    PubMed

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

    2015-06-01

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

  10. Aspirin reduces lipopolysaccharide-induced pulmonary inflammation in human models of ARDS.

    PubMed

    Hamid, U; Krasnodembskaya, A; Fitzgerald, M; Shyamsundar, M; Kissenpfennig, A; Scott, C; Lefrancais, E; Looney, M R; Verghis, R; Scott, J; Simpson, A J; McNamee, J; McAuley, D F; O'Kane, C M

    2017-11-01

    Platelets play an active role in the pathogenesis of acute respiratory distress syndrome (ARDS). Animal and observational studies have shown aspirin's antiplatelet and immunomodulatory effects may be beneficial in ARDS. To test the hypothesis that aspirin reduces inflammation in clinically relevant human models that recapitulate pathophysiological mechanisms implicated in the development of ARDS. Healthy volunteers were randomised to receive placebo or aspirin 75  or 1200 mg (1:1:1) for seven days prior to lipopolysaccharide (LPS) inhalation, in a double-blind, placebo-controlled, allocation-concealed study. Bronchoalveolar lavage (BAL) was performed 6 hours after inhaling 50 µg of LPS. The primary outcome measure was BAL IL-8. Secondary outcome measures included markers of alveolar inflammation (BAL neutrophils, cytokines, neutrophil proteases), alveolar epithelial cell injury, systemic inflammation (neutrophils and plasma C-reactive protein (CRP)) and platelet activation (thromboxane B2, TXB2). Human lungs, perfused and ventilated ex vivo (EVLP) were randomised to placebo or 24 mg aspirin and injured with LPS. BAL was carried out 4 hours later. Inflammation was assessed by BAL differential cell counts and histological changes. In the healthy volunteer (n=33) model, data for the aspirin groups were combined. Aspirin did not reduce BAL IL-8. However, aspirin reduced pulmonary neutrophilia and tissue damaging neutrophil proteases (Matrix Metalloproteinase (MMP)-8/-9), reduced BAL concentrations of tumour necrosis factor α and reduced systemic and pulmonary TXB2. There was no difference between high-dose and low-dose aspirin. In the EVLP model, aspirin reduced BAL neutrophilia and alveolar injury as measured by histological damage. These are the first prospective human data indicating that aspirin inhibits pulmonary neutrophilic inflammation, at both low and high doses. Further clinical studies are indicated to assess the role of aspirin in the

  11. Anticancer effects of novel thalidomide analogs in A549 cells through inhibition of vascular endothelial growth factor and matrix metalloproteinase-2.

    PubMed

    El-Aarag, Bishoy; Kasai, Tomonari; Masuda, Junko; Agwa, Hussein; Zahran, Magdy; Seno, Masaharu

    2017-01-01

    Lung cancer is one of the major causes of cancer-related mortality worldwide, and non-small-cell lung cancer is the most common form of lung cancer. Several studies had shown that thalidomide has potential for prevention and therapy of cancer. Therefore, the current study aimed to investigate the antitumor effects of two novel thalidomide analogs in human lung cancer A549 cells. The antiproliferative, antimigratory, and apoptotic effects in A549 cells induced by thalidomide analogs were examined. In addition, their effects on the expression of mRNAs encoding vascular endothelial growth factor165 (VEGF165) and matrix metalloproteinase-2 (MMP-2) were evaluated. Their influence on the tumor volume in nude mice was also determined. Results revealed that thalidomide analogs exhibited antiproliferative, antimigratory, and apoptotic activities with more pronounced effect than thalidomide drug. Furthermore, analogs 1 and 2 suppressed the expression levels of VEGF165 by 42% and 53.2% and those of MMP-2 by 45% and 52%, respectively. Thalidomide analogs 1 and 2 also reduced the tumor volume by 30.11% and 53.52%, respectively. Therefore, this study provides evidence that thalidomide analogs may serve as a new therapeutic option for treating lung cancer. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

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

    SciTech Connect

    Ohnuma, Tomokazu; Matsumoto, Takashi; Itoi, Ayano

    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 amore » 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.« less

  13. Revelation of Different Nanoparticle-Uptake Behavior in Two Standard Cell Lines NIH/3T3 and A549 by Flow Cytometry and Time-Lapse Imaging.

    PubMed

    Jochums, André; Friehs, Elsa; Sambale, Franziska; Lavrentieva, Antonina; Bahnemann, Detlef; Scheper, Thomas

    2017-07-19

    The uptake of nanomaterials into different cell types is a central pharmacological issue for the determination of nanotoxicity as well as for the development of drug delivery strategies. Most responses of the cells depend on their intracellular interactions with nanoparticles (NPs). Uptake behavior can be precisely investigated in vitro, with sensitive high throughput methods such as flow cytometry. In this study, we investigated two different standard cell lines, human lung carcinoma (A549) and mouse fibroblast (NIH/3T3) cells, regarding their uptake behavior of titanium dioxide NPs. Cells were incubated with different concentrations of TiO₂ NPs and samples were taken at certain time points to compare the uptake kinetics of both cell lines. Samples were analyzed with the help of flow cytometry by studying changes in the side and forward scattering signal. To additionally enable a detection via fluorescence, NPs were labeled with the fluorescent dye fluorescein isothiocyanate (FITC) and propidium iodide (PI). We found that NIH/3T3 cells take up the studied NPs more efficiently than A549 cells. These findings were supported by time-lapse microscopic imaging of the cells incubated with TiO₂ NPs. Our results confirm that the uptake behavior of individual cell types has to be considered before interpreting any results of nanomaterial studies.

  14. Revelation of Different Nanoparticle-Uptake Behavior in Two Standard Cell Lines NIH/3T3 and A549 by Flow Cytometry and Time-Lapse Imaging

    PubMed Central

    Jochums, André; Friehs, Elsa; Sambale, Franziska; Lavrentieva, Antonina; Bahnemann, Detlef; Scheper, Thomas

    2017-01-01

    The uptake of nanomaterials into different cell types is a central pharmacological issue for the determination of nanotoxicity as well as for the development of drug delivery strategies. Most responses of the cells depend on their intracellular interactions with nanoparticles (NPs). Uptake behavior can be precisely investigated in vitro, with sensitive high throughput methods such as flow cytometry. In this study, we investigated two different standard cell lines, human lung carcinoma (A549) and mouse fibroblast (NIH/3T3) cells, regarding their uptake behavior of titanium dioxide NPs. Cells were incubated with different concentrations of TiO2 NPs and samples were taken at certain time points to compare the uptake kinetics of both cell lines. Samples were analyzed with the help of flow cytometry by studying changes in the side and forward scattering signal. To additionally enable a detection via fluorescence, NPs were labeled with the fluorescent dye fluorescein isothiocyanate (FITC) and propidium iodide (PI). We found that NIH/3T3 cells take up the studied NPs more efficiently than A549 cells. These findings were supported by time-lapse microscopic imaging of the cells incubated with TiO2 NPs. Our results confirm that the uptake behavior of individual cell types has to be considered before interpreting any results of nanomaterial studies. PMID:29051447

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

    PubMed

    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. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. An imaging flow cytometry method to assess ricin trafficking in A549 human lung epithelial cells.

    PubMed

    Jenner, Dominic; Chong, Damien; Walker, Nicola; Green, A Christopher

    2018-02-01

    The endocytosis and trafficking of ricin in mammalian cells is an important area of research for those producing ricin anti-toxins and other ricin therapeutics. Ricin trafficking is usually observed by fluorescence microscopy techniques. This gives good resolution and leads to a detailed understanding of the internal movement of ricin within cells. However, microscopy techniques are often hampered by complex analysis and quantification techniques, and the inability to look at ricin trafficking in large populations of cells. In these studies we have directly labelled ricin and assessed if its trafficking can be observed using Imaging Flow Cytometry (IFC) both to the cytoplasmic region of cells and specifically to the Golgi apparatus. Using IDEAS® data analysis software the specific fluorescence location of the ricin within the cells was analysed. Then, using cytoplasmic masking techniques to quantify the number of cells with endocytosed cytoplasmic ricin or cells with Golgi-associated ricin, kinetic endocytosis curves were generated. Here we present, to the authors' knowledge, the first example of using imaging flow cytometry for evaluating the subcellular transport of protein cargo, using the trafficking of ricin toxin in lung cells as a model. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

  17. Hypoxia induces arginase II expression and increases viable human pulmonary artery smooth muscle cell numbers via AMPKα1 signaling

    PubMed Central

    Xue, Jianjing; Nelin, Leif D.

    2017-01-01

    Pulmonary artery smooth muscle cell (PASMC) proliferation is one of the hallmark features of hypoxia-induced pulmonary hypertension. With only supportive treatment options available for this life-threatening disease, treating and preventing the proliferation of PASMCs is a viable therapeutic option. A key promoter of hypoxia-induced increases in the number of viable human PASMCs is arginase II, with attenuation of viable cell numbers following pharmacologic inhibition or siRNA knockdown of the enzyme. Additionally, increased levels of arginase have been demonstrated in the pulmonary vasculature of patients with pulmonary hypertension. The signaling pathways responsible for the hypoxic induction of arginase II in PASMCs, however, remain unknown. Hypoxia is a recognized activator of AMPK, which is known to be expressed in human PASMCs (hPASMCs). Activation of AMPK by hypoxia has been shown to promote cell survival in PASMCs. In addition, pharmacologic agents targeting AMPK have been shown to attenuate chronic hypoxia-induced pulmonary hypertension in animal models. The present studies tested the hypothesis that hypoxia-induced arginase II expression in hPASMCs is mediated through AMPK signaling. We found that pharmacologic inhibitors of AMPK, as well as siRNA knockdown of AMPKα1, prevented hypoxia-induced arginase II. The hypoxia-induced increase in viable hPASMC numbers was also prevented following both pharmacologic inhibition and siRNA knockdown of AMPK. Furthermore, we demonstrate that overexpression of AMPK induced arginase II protein expression and viable cells numbers in hPASMCs. PMID:28213467

  18. In vitro effects of water-pipe smoke condensate on the endocytic activity of Type II alveolar epithelial cells (A549) with bacillus Calmette-Guérin.

    PubMed

    Adcock, Ian M; Mortaz, Esmaeil; Alipoor, Shamila D; Garssen, Johan; Akbar Velayati, Ali

    2016-12-01

    Tuberculosis (TB) is a major global health problem and poses immense threats to many populations. The association between tobacco smoke and TB has already been studied. Water-pipe smoking has become an increasing problem not only in Middle Eastern countries but also globally as it is considered by users as being safer than cigarettes. The presence of high levels of toxic substances in water-pipe smoke may be predisposing factors that enhance the incidence of pulmonary disorders in water-pipe smokers. For example, uncontrolled macropinocytosis occurs in alveolar epithelial cells following exposure to water-pipe smoke, which may predispose individuals to pulmonary infection. In this work, we studied the effects of water-pipe condense (WPC) on the internalization of Mycobacterium bovis (bacillus Calmette-Guérin [BCG]) by macropinocytosis in Type II alveolar epithelial cells (A549). A549 cells were treated by WPC (4mg/mL) for 24 h, 48 h, 72 h, and 96 h, respectively. The effect on cell proliferation was studied using a methylthiazolyldiphenyl-tetrazolium bromide (MTT) reduction assay. Cells were exposed to fluorescein isothiocyanate (FITC)-dextran (1mg/mL; control) and FITC-BCG (multiplicity of infection, 10) for 20min at 37°C before their collection and the uptake of BCG-FITC was determined by flow cytometry. Similar experiments were performed at 4°C as a control. WPC (4mg/mL) after 72h (1.4±0.2-fold, p<0.05) and 96h (1.6±0.2-fold, p<0.05) hours increased the uptake of BCG-FITC. No effect on BCG-FITC uptake was observed at 24h or 48h. WPC also significantly increased the uptake of FITC-dextran (2.9±0.3-fold, p<0.05) after 96h. WPC also significantly decreased cell proliferation after 24h (84±2%), 48h (78±3%), 72h (64±2%, p<0.05), and 96h (45±2%, p<0.05). WPC exposure increased epithelial cells' permeability and death and enhanced their capacity for macropinocytosis. Our in vitro data suggest possible harmful effects of WPC on the ability of lung epithelial

  19. RELM-β promotes human pulmonary artery smooth muscle cell proliferation via FAK-stimulated surviving

    SciTech Connect

    Lin, Chunlong, E-mail: lclmd@sina.com; Li, Xiaohui; Luo, Qiong

    2017-02-01

    Resistin-like molecule-β (RELM-β), focal adhesion kinase (FAK), and survivin may be involved in the proliferation of cultured human pulmonary artery smooth muscle cells (HPAMSCs), which is involved in pulmonary hypertension. HPAMSCs were treated with human recombinant RELM-β (rhRELM-β). siRNAs against FAK and survivin were transfected into cultured HPASMCs. Expression of FAK and survivin were examined by RT-PCR and western blot. Immunofluorescence was used to localize FAK. Flow cytometry was used to examine cell cycle distribution and cell death. Compared to the control group, all rhRELM-β-treated groups demonstrated significant increases in the expression of FAK and survivin (P<0.05). rhRELM-β significantly increasedmore » the proportion of HPASMCs in the S phase and decreased the proportion in G0/G1. FAK siRNA down-regulated survivin expression while survivin siRNA did not affect FAK expression. FAK siRNA effectively inhibited FAK and survivin expression in RELM-β-treated HPASMCs and partially suppressed cell proliferation. RELM-β promoted HPASMC proliferation and upregulated FAK and survivin expression. In conclusion, results suggested that FAK is upstream of survivin in the signaling pathway mediating cell proliferation. FAK seems to be important in RELM-β-induced HPASMC proliferation, partially by upregulating survivin expression. - Highlights: • rhRELM-β increased the expression of FAK and survivin. • rhRELM-β increased the proportion of HPASMCs in the S phase. • FAK is upstream of survivin in the signaling pathway mediating cell proliferation. • FAK is important in RELM-β-induced HPASMC proliferation, partly via survivin.« less

  20. Absence of viscerosomatic inhibition with injections of lobeline designed to activate human pulmonary C fibres.

    PubMed

    Gandevia, S C; Butler, J E; Taylor, J L; Crawford, M R

    1998-08-15

    1. Activation of pulmonary C fibres (J receptors) in animals produces inhibition of spinal motoneurones. Intravenous bolus injections of lobeline are believed to activate pulmonary C fibres (J receptors) in human subjects and to produce characteristic sensations and cardiorespiratory responses. This study quantified the respiratory sensations evoked by such injections and then used a range of suprathreshold doses of lobeline and tested for the presence of reflex or descending inhibition of motoneuronal output. 2. Injections of lobeline produced dose-dependent sensations of respiratory discomfort referred to the throat and upper chest beginning within about 10 s and often associated with coughing. As the dose increased the latency for the sensations decreased while their duration and intensity increased. Reflex changes in blood pressure, heart rate and ventilation also occurred. 3. Injections of lobeline at doses sufficient to evoke respiratory discomfort lasting 25-32 s (37-73 microgram kg-1) increased the size of the H reflex in soleus with an onset latency of about 10 s and lasting about 20 s. 4. The size of EMG responses evoked in upper limb muscles by transcranial magnetic stimulation of the motor cortex increased shortly after injections and remained elevated for about 30-35 s. 5. Injections of lobeline during sustained voluntary contractions of the elbow flexors at submaximal or maximal levels did not impair the ability to produce force. 6. Walking was not disrupted by repeated suprathreshold doses of lobeline. 7. It is concluded that injections of lobeline sufficient to evoke cardiorespiratory reflexes and sensations of severe respiratory discomfort are not associated with functionally important inhibition of motor performance. The results cast doubt on the ability of the J reflex to limit exercise in humans.

  1. Absence of viscerosomatic inhibition with injections of lobeline designed to activate human pulmonary C fibres

    PubMed Central

    Gandevia, S C; Butler, J E; Taylor, J L; Crawford, M R

    1998-01-01

    Activation of pulmonary C fibres (J receptors) in animals produces inhibition of spinal motoneurones. Intravenous bolus injections of lobeline are believed to activate pulmonary C fibres (J receptors) in human subjects and to produce characteristic sensations and cardiorespiratory responses. This study quantified the respiratory sensations evoked by such injections and then used a range of suprathreshold doses of lobeline and tested for the presence of reflex or descending inhibition of motoneuronal output.Injections of lobeline produced dose-dependent sensations of respiratory discomfort referred to the throat and upper chest beginning within about 10 s and often associated with coughing. As the dose increased the latency for the sensations decreased while their duration and intensity increased. Reflex changes in blood pressure, heart rate and ventilation also occurred.Injections of lobeline at doses sufficient to evoke respiratory discomfort lasting 25-32 s (37-73 μg kg−1) increased the size of the H reflex in soleus with an onset latency of about 10 s and lasting about 20 s.The size of EMG responses evoked in upper limb muscles by transcranial magnetic stimulation of the motor cortex increased shortly after injections and remained elevated for about 30-35 s.Injections of lobeline during sustained voluntary contractions of the elbow flexors at submaximal or maximal levels did not impair the ability to produce force.Walking was not disrupted by repeated suprathreshold doses of lobeline.It is concluded that injections of lobeline sufficient to evoke cardiorespiratory reflexes and sensations of severe respiratory discomfort are not associated with functionally important inhibition of motor performance. The results cast doubt on the ability of the J reflex to limit exercise in humans. PMID:9679182

  2. Evaluation of six different airway devices regarding regurgitation and pulmonary aspiration during cardio-pulmonary resuscitation (CPR) - A human cadaver pilot study.

    PubMed

    Piegeler, Tobias; Roessler, Bernard; Goliasch, Georg; Fischer, Henrik; Schlaepfer, Martin; Lang, Susanna; Ruetzler, Kurt

    2016-05-01

    Chest compressions and ventilation are lifesaving tasks during cardio-pulmonary resuscitation (CPR). Besides oxygenation, endotracheal intubation (ETI) during CPR is performed to avoid aspiration of gastric contents. If intubation is difficult or impossible, supraglottic airway devices are utilized. We tested six different airway devices regarding their potential to protect against regurgitation and aspiration during CPR in a randomized experimental human cadaver study. Five-hundred ml of 0.01% methylene-blue-solution were instilled into the stomach of 30 adult human cadavers via an oro-gastric tube. The cadavers were then randomly assigned to one of six groups, resulting in 5 cadavers in each group. Airway management was performed with either bag-valve ventilation, Laryngeal Tube, EasyTube, Laryngeal Mask (Classic), I-Gel, or ETI. Thereafter 5min of CPR were performed according to the 2010 Guidelines of the European Resuscitation Council. Pulmonary aspiration was defined as the presence of methylene-blue-solution below the vocal cords or the ETI cuff as assessed by fiber-optic bronchoscopy. Thirty cadavers were included (14 females, 16 males). Aspiration was detected in three out of five cadavers receiving bag-valve ventilation and in two out of five intubated with LMA or I-Gel. In cadavers intubated with the LT, aspiration occurred in one out of five cases. No aspiration could be detected in cadavers intubated with ETI and EasyTube. This study provides experimental evidence that, during CPR, ETI offers superior protection against regurgitation and pulmonary aspiration of gastric contents than supraglottic airway devices or bag-valve ventilation. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  3. 9,10-Phenanthrenequinone promotes secretion of pulmonary aldo-keto reductases with surfactant.

    PubMed

    Matsunaga, Toshiyuki; Haga, Mariko; Watanabe, Gou; Shinoda, Yuhki; Endo, Satoshi; Kajiwara, Yu; Tanaka, Hiroyuki; Inagaki, Naoki; El-Kabbani, Ossama; Hara, Akira

    2012-02-01

    9,10-Phenanthrenequinone (9,10-PQ), a major quinone in diesel exhaust particles, induces apoptosis via the generation of reactive oxygen species (ROS) because of 9,10-PQ redox cycling. We have found that intratracheal infusion of 9,10-PQ facilitates the secretion of surfactant into rat alveolus. In the cultured rat lung, treatment with 9,10-PQ results in an increase in a lower-density surfactant by ROS generation through redox cycling of the quinone. The surfactant contains aldo-keto reductase (AKR) 1C15, which reduces 9,10-PQ and the enzyme level in the surfactant increases on treatment with 9,10-PQ suggesting an involvement of AKR1C15 in the redox cycling of the quinone. In six human cell types (A549, MKN45, Caco2, Hela, Molt4 and U937) only type II epithelial A549 cells secrete three human AKR1C subfamily members (AKR1C1, AKR1C2 and AKR1C3) with the surfactant into the medium; this secretion is highly increased by 9,10-PQ treatment. Using in vitro enzyme inhibition analysis, we have identified AKR1C3 as the most abundantly secreted AKR1C member. The AKR1C enzymes in the medium efficiently reduce 9,10-PQ and initiate its redox cycling accompanied by ROS production. The exposure of A549 cells to 9,10-PQ provokes viability loss, which is significantly protected by the addition of the AKR1C3 inhibitor and antioxidant enzyme and by the removal of the surfactants from the culture medium. Thus, the AKR1C enzymes secreted in pulmonary surfactants probably participate in the toxic mechanism triggered by 9,10-PQ.

  4. Silencing the Snail-dependent RNA splice regulator ESRP1 drives malignant transformation of human pulmonary epithelial cells.

    PubMed

    Walser, Tonya C; Jing, Zhe; Tran, Linh M; Lin, Ying Q; Yakobian, Natalie; Wang, Gerald; Krysan, Kostyantyn; Zhu, Li X; Sharma, Sherven; Lee, Mi-Heon; Belperio, John A; Ooi, Aik T; Gomperts, Brigitte N; Shay, Jerry W; Larsen, Jill E; Minna, John D; Hong, Long-Sheng; Fishbein, Michael C; Dubinett, Steven M

    2018-02-05

    Epithelial-to-mesenchymal transition (EMT) is organized in cancer cells by a set of key transcription factors, but the significance of this process is still debated including in non-small cell lung cancer (NSCLC). Here we report increased expression of the EMT-inducing transcription factor Snail in premalignant pulmonary lesions, relative to histologically normal pulmonary epithelium. In immortalized human pulmonary epithelial cells and isogenic derivatives, we documented Snail-dependent anchorage-independent growth in vitro and primary tumor growth and metastatic behavior in vivo. Snail-mediated transformation relied upon silencing of the tumor suppressive RNA splicing regulatory protein ESRP1. In clinical specimens of NSCLC, ESRP1 loss was documented in Snail-expressing premalignant pulmonary lesions. Mechanistic investigations showed that Snail drives malignant progression in an ALDH+CD44+CD24- pulmonary stem cell subset in which ESRP1 and stemness-repressing microRNAs are inhibited. Collectively, our results show how ESRP1 loss is a critical event in lung carcinogenesis, and they identify new candidate directions for targeted therapy of NSCLC. Copyright ©2018, American Association for Cancer Research.

  5. Normal development of the pulmonary veins in human embryos and formulation of a morphogenetic concept for sinus venosus defects.

    PubMed

    Blom, N A; Gittenberger-de Groot, A C; Jongeneel, T H; DeRuiter, M C; Poelmann, R E; Ottenkamp, J

    2001-02-01

    A sinus venosus defect is a form of interatrial communication associated with abnormal drainage of the right pulmonary veins. Its morphogenesis still remains unclear. We therefore studied the normal development of pulmonary veins in human embryos in relation to the sinus venosus and the dorsal mesocardium using graphic reconstructions and HNK-1 immunohistochemistry. Twenty embryos, ranging from 4 to 7 weeks' gestation, were examined. At 4 weeks, the orifice of the nonlumenized common pulmonary vein is visible as an endothelial invagination within the sinus venosus segment. Development of the muscular septum primum and the ventral proliferation of extracardiac mesenchyme from the dorsal mesocardium positions the common pulmonary vein (CPV) eventually into the left atrium. The right wall of the CPV contributes to the posterior part of the atrial septum and is continuous with the dorsal sinuatrial fold (the future left venous valve). With the use of HNK-1 antigen expression as a marker for sinus venosus myocardium, this common wall between the right-sided sinus venosus and the CPV is demonstrated, and at 7 weeks the proximal part of the right upper pulmonary vein also becomes part of this common wall. This study demonstrates that the CPV develops within the sinus venosus segment and that later a common myocardial wall is present between the sinus venosus in the right atrium and the CPV. A deficiency of this wall explains the development of sinus venosus defects.

  6. Differential activation of inflammatory pathways in A549 type II pneumocytes by Streptococcus pneumoniae strains with different adherence properties

    PubMed Central

    Robson, Rachel L; Reed, Natalie A; Horvat, Rebecca T

    2006-01-01

    Background Adherence of Streptococcus pneumoniae bacteria to lung cells is a first step in the progression from asymptomatic carriage to pneumonia. Adherence abilities vary widely among S. pneumoniae patient isolates. In this study, the binding properties of S. pneumoniae isolates and the effects of binding on activation of the Nuclear Factor-Kappa-B (NFκB) pathway and cytokine secretion by type II pneumocytes were measured. Methods Mechanisms of high- and low-binding S. pneumoniae adherence to A549 cells were investigated by blocking putative receptors on bacteria and host cells with antibody and by eluting choline-binding proteins off of bacterial surfaces. NFκB activation was measured by western blot and immunocytochemistry and cytokine secretion was detected by a protein array. Results This study shows that S. pneumoniae isolates from pneumonia patients (n = 298) can vary by as much as 1000-fold in their ability to bind to human lung epithelial cells. This difference resulted in differential activation of the NFκB pathway. High-, but not low-binding S. pneumoniae used Choline-binding protein A (CbpA) to bind to complement component C3 on epithelial cell surfaces. Interleukin-8 (IL-8) was the only cytokine secreted by cells treated with either low- or high-binding S. pneumoniae. Conclusion These results indicate that S. pneumoniae clinical isolates are not homogeneous in their interaction with host epithelial cells. The differential activation of host cells by high- and low-binding S. pneumoniae strains could have implications for the treatment of pneumococcal pneumonia and for vaccine development. PMID:16606470

  7. Cytotoxic (A549) and antimicrobial effects of Methylobacterium sp. isolate (ERI-135) from Nilgiris forest soil, India.

    PubMed

    Balachandran, C; Duraipandiyan, V; Ignacimuthu, S

    2012-09-01

    To assess the antimicrobial and cytotoxic effects of Methylobacterium sp. isolated from soil sample of Doddabetta forest, Nilgiris, Western Ghats of Tamil Nadu. Isolation of Methylobacterium was performed from soils by serial dilution plate technique. The strain was grown in modified nutrient gulucose agar (MNGA) medium to study the morphology and biochemical characteristics. Methylobacterium sp. was screened for its antimicrobial activity against pathogenic bacteria and fungi. The strain was subjected to 16S rRNA analysis and was identified as Methylobacterium sp. The nucleotide sequence of the 16S rRNA gene of the isolate exhibited close similarity with other Methylobacterium sp. and has been submitted to Genbank. The antibacterial substances were extracted using chloroform and ethyl acetate from MNGA medium in which ERI-135 had grown for 5 d at 30 °C. Cytotoxic effect was also studied. GC-MS analysis was carried out. The antimicrobial activity was assessed using broth micro dilution technique. Ethyl acetate extract showed activity against bacteria such as Bacillus subtilis, Klebsiella pneumoniae (K. pneumoniae), Pseudomonas aeruginosa, Salmonella typhimurium, Shigella flexneri, Enterobacter aerogenes, Staphylococcus aureu and Staphylococcus epidermidis (S. epidermidis) and fungi such as, Candida albicans and Trichophyton rubrum. The lowest minimum inhibitory concentrations were: 250 µg/mL against S. epidermidis and 250µg/mL against K. pneumonia. The isolate had the ability to produce enzymes such as protease. The exyract showed cytotoxic effect in human adenocarcinoma cancer cell line (A549). GC-MS analysis showed the presence of isovaleric acid (3.64%), 2-Methylbutanoic acid (5.03%), isobutyramide (5.05%), N,N-oimethylformamide-di-t-butylacetal (9.79%), benzeneacetamide (15.56%), octyl butyl phthalate (3.59%) and diisooctyl phthalate (5.79) in the extract. Methylobacterium sp. (ERI-135) showed promising antibacterial and cytotoxic activity. This is the first

  8. Human Metapneumovirus Infection in Chronic Obstructive Pulmonary Disease: Impact of Glucocorticosteroids and Interferon.

    PubMed

    Kan-O, Keiko; Ramirez, Ruben; MacDonald, Martin I; Rolph, Michael; Rudd, Penny A; Spann, Kirsten M; Mahalingam, Suresh; Bardin, Philip G; Thomas, Belinda J

    2017-05-15

    Human metapneumovirus (hMPV) infection is implicated in exacerbations of asthma and chronic obstructive pulmonary disease (COPD). Research into the pathogenesis of infection is restricted to animal models, and information about hMPV replication and inflammatory and immune responses in human disease is limited. Human primary bronchial epithelial cells (PBECs) from healthy and asthmatic subjects and those with COPD were infected with hMPV, with or without glucocorticosteroid (GCS) exposure. Viral replication, inflammatory and immune responses, and apoptosis were analyzed. We also determined whether adjuvant interferon (IFN) can blunt hMPV infection in vitro and in a murine model. hMPV infected human PBECs and viral replication was enhanced in cells from patients with COPD. The virus induced gene expression of IFN-stimulated gene 56 (ISG56) and IFN-β, as well as IFN-γ-inducible protein 10 (IP-10) and regulated on activation, normal T cell expressed and secreted (RANTES), and more so in cells from patients with COPD. GCS exposure enhanced hMPV replication despite increased IFN expression. Augmented virus replication associated with GCS was mediated by reduced apoptosis via induction of antiapoptotic genes. Adjuvant IFN treatment suppressed hMPV replication in PBECs and reduced hMPV viral titers and inflammation in vivo. hMPV infects human PBECs, eliciting innate and inflammatory responses. Replication is enhanced by GCS and adjuvant IFN is an effective treatment, restricting virus replication and proinflammatory consequences of hMPV infections. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  9. Decellularization of human donor aortic and pulmonary valved conduits using low concentration sodium dodecyl sulfate.

    PubMed

    Vafaee, Tayyebeh; Thomas, Daniel; Desai, Amisha; Jennings, Louise M; Berry, Helen; Rooney, Paul; Kearney, John; Fisher, John; Ingham, Eileen

    2018-02-01

    The clinical use of decellularized cardiac valve allografts is increasing. Long-term data will be required to determine whether they outperform conventional cryopreserved allografts. Valves decellularized using different processes may show varied long-term outcomes. It is therefore important to understand the effects of specific decellularization technologies on the characteristics of donor heart valves. Human cryopreserved aortic and pulmonary valved conduits were decellularized using hypotonic buffer, 0.1% (w/v) sodium dodecyl sulfate and nuclease digestion. The decellularized tissues were compared to cellular cryopreserved valve tissues using histology, immunohistochemistry, quantitation of total deoxyribose nucleic acid, collagen and glycosaminoglycan content, in vitro cytotoxicity assays, uniaxial tensile testing and subcutaneous implantation in mice. The decellularized tissues showed no histological evidence of cells or cell remnants and >97% deoxyribose nucleic acid removal in all regions (arterial wall, muscle, leaflet and junction). The decellularized tissues retained collagen IV and von Willebrand factor staining with some loss of fibronectin, laminin and chondroitin sulfate staining. There was an absence of major histocompatibility complex Class I staining in decellularized pulmonary valve tissues, with only residual staining in isolated areas of decellularized aortic valve tissues. The collagen content of the tissues was not decreased following decellularization however the glycosaminoglycan content was reduced. Only moderate changes in the maximum load to failure of the tissues were recorded postdecellularization. The decellularized tissues were noncytotoxic in vitro, and were biocompatible in vivo in a mouse subcutaneous implant model. The decellularization process will now be translated into a good manufacturing practices-compatible process for donor cryopreserved valves with a view to future clinical use. Copyright © 2016 The Authors Tissue

  10. Diameters and cross-sectional areas of branches in the human pulmonary arterial tree.

    PubMed

    Horsfield, K; Woldenberg, M J

    1989-03-01

    Measurements were made of the diameters of the three branches meeting at each of 1,937 bifurcations in the pulmonary arterial tree, using resin casts from two fully inflated human lungs. Cross-sectional areas of the parent branch and of the daughter branches were calculated and plotted on a log-log plot, which showed that mean cross-sectional area increases in a constant proportion of 1.0879 at bifurcations. The mean value of the ratio of daughter branch diameters at bifurcations was 0.7849. The mean value of the exponent z in the equation flow = k (diameter(z)) was found to be 2.3 +/- 0.1, which is equal to the optimal value for minimizing power and metabolic costs for fully developed turbulent flow. Although Reynolds number may exceed 2,000 in the larger branches of the pulmonary artery, turbulent flow probably does not occur, and in the peripheral branches Reynolds number is always low, excluding turbulent flow in these branches. This finding seems to be incompatible with the observed value of z. A possible explanation may be that other factors may need to be taken into account when calculating the theoretical optimum value of z for minimum power dissipation, such as the relatively short branches and the disturbances of flow occurring at bifurcations. Alternatively, higher arterial diameters reduce acceleration of the blood during systole, reduce turbulent flow, and increase the reservoir function of the larger arteries. These higher diameters result in a lower value of z.

  11. Decellularization of human donor aortic and pulmonary valved conduits using low concentration sodium dodecyl sulfate

    PubMed Central

    Vafaee, Tayyebeh; Thomas, Daniel; Desai, Amisha; Jennings, Louise M.; Berry, Helen; Rooney, Paul; Kearney, John; Fisher, John

    2017-01-01

    Abstract The clinical use of decellularized cardiac valve allografts is increasing. Long‐term data will be required to determine whether they outperform conventional cryopreserved allografts. Valves decellularized using different processes may show varied long‐term outcomes. It is therefore important to understand the effects of specific decellularization technologies on the characteristics of donor heart valves. Human cryopreserved aortic and pulmonary valved conduits were decellularized using hypotonic buffer, 0.1% (w/v) sodium dodecyl sulfate and nuclease digestion. The decellularized tissues were compared to cellular cryopreserved valve tissues using histology, immunohistochemistry, quantitation of total deoxyribose nucleic acid, collagen and glycosaminoglycan content, in vitro cytotoxicity assays, uniaxial tensile testing and subcutaneous implantation in mice. The decellularized tissues showed no histological evidence of cells or cell remnants and >97% deoxyribose nucleic acid removal in all regions (arterial wall, muscle, leaflet and junction). The decellularized tissues retained collagen IV and von Willebrand factor staining with some loss of fibronectin, laminin and chondroitin sulfate staining. There was an absence of major histocompatibility complex Class I staining in decellularized pulmonary valve tissues, with only residual staining in isolated areas of decellularized aortic valve tissues. The collagen content of the tissues was not decreased following decellularization however the glycosaminoglycan content was reduced. Only moderate changes in the maximum load to failure of the tissues were recorded postdecellularization. The decellularized tissues were noncytotoxic in vitro, and were biocompatible in vivo in a mouse subcutaneous implant model. The decellularization process will now be translated into a good manufacturing practices‐compatible process for donor cryopreserved valves with a view to future clinical use. Copyright © 2016 The Authors

  12. Pulmonary and extrapulmonary complications of human rhinovirus infection in critically ill patients.

    PubMed

    To, Kelvin K W; Lau, Susanna K P; Chan, Kwok-Hei; Mok, Ka-Yi; Luk, Hayes K H; Yip, Cyril C Y; Ma, Yat-Kwan; Sinn, Lorraine H Y; Lam, Sonia H Y; Ngai, Chun-Wai; Hung, Ivan F N; Chan, Kwok-Hung; Yuen, Kwok-Yung

    2016-04-01

    Human rhinovirus (HRV) is frequently detected in patients with respiratory tract infection. However, the full clinical spectrum of HRV infection in critically ill patients is not well characterized. To evaluate the clinical and virological characteristics of critically ill patients with HRV infection. HRV-specific reverse transcription-polymerase chain reaction (RT-PCR) was performed on nasopharyngeal aspirate (NPA) specimens from 294 adult patients who required admission into the intensive care unit (ICU). Clinical characteristics were analyzed. HRV genotyping using the 5'UTR-VP4-VP2 region was performed. HRV was detected in NPA specimens of 22 patients (7.5%) by RT-PCR. Dyspnea was the most common presenting symptom (16/22; 72.7%), but seizure also occurred in 5 (22.7%) patients. Exacerbation of underlying disease occurred in 12 (54.5%) patients. Four (18.2%) patients died, and HRV was considered to play a role as the cause of death in 3 patients. Thirteen (59.1%) patients had pneumonia, and the most common radiological finding was consolidation (6/13; 46.2%). Streptococcus pneumoniae was the most common co-pathogen among patients with pneumonia. Among the 9 patients without pneumonia, 3 patients had exacerbation of underlying lung diseases, 3 patients had acute pulmonary edema, 2 patients with diabetes mellitus had acute complications from poor glycemic control, and 1 patient had status epilepticus. HRV-A was the most common species (64.3%), but there was no clear relationship between HRV species and clinical presentation. Both pulmonary and extrapulmonary complications of HRV were common in critically ill patients. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Enhanced production of nitric oxide in A549 cells through activation of TRPA1 ion channel by cold stress.

    PubMed

    Sun, Wenwu; Wang, Zhonghua; Cao, Jianping; Wang, Xu; Han, Yaling; Ma, Zhuang

    2014-08-31

    The respiratory epithelium is exposed to the external environment, and inhalation of cold air is common during the season of winter. In addition, the lung is a major source of nitric oxide (NO). However, the effect of cold stress on the production of NO is still unclear. In the present work, We measured the change of NO in single cell with DACF-DA and the change in cytosolic Ca(2+) concentration ([Ca(2+)]c) in A549 cell. We observed that cold stress (from 20 °C to 5 °C) induced an increase of NO in A549 cell, which was completely abolished by applying an extracellular Ca(2+) free medium. Further experiments showed that cold-sensing transient receptor potential subfamily member 1 (TRPA1) channel agonist (allyl isothiocyanate, AITC) increased the production of NO and the level of [Ca(2+)]c in A549 cell. Additionally, TRPA1 inhibitor, Ruthenium red (RR) and camphor, significantly blocked the enhanced production of NO and the rise of [Ca(2+)]c induced by AITC or cold stimulation, respectively. Taken together, these data indicated that cold-induced TRPA1 activation was responsible for the enhanced production of NO in A549 cell. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. SU-F-T-675: Down-Regulating the Expression of Cdc42 and Inhibition of Migration of A549 with Combined Treatment of Ionizing Radiation and Sevoflurane

    SciTech Connect

    Feng, Y; Feng, J; Huang, Z

    2016-06-15

    Purpose: Cdc42 is involved in cell transformation, proliferation, invasion and metastasis of human cancer cells. Cdc42 overexpression has been reported in several types of cancers. This study investigated the combined treatment effects of ionizing radiation and sevoflurane on down-regulating Cdc42 expression and suppressing migration of human adenocarcinoma cell line A549. Methods: Samples of A549 cells with Cdc42 overexpression were created and Cdc42 expression was determined by Western blotting. Increase of migration speed by Cdc42-HA overexpression was confirmed with an initial in-vitro scratch assay. The cells grown in culture media were separated into 2 groups of 6 samples: one for themore » control and the other was treated with 4% sevoflurane for 5hrs prior to a single-fraction radiation of 4Gy using a 6MV beam. Cell migration speeds of the 2 groups were measured with an initial in-vitro scratch assay. The scratch was created with a pipette tip immediately after treatment and images at 4 post-treatment time points (0h, 3h, 6h, 12h) were acquired. The distance between the two separated sides at 0h was used as reference and subsequent changes of the distance over time was defined as the cell migration speed. Image processing and measurement were performed with an in-house software. The experiment was repeated three times independently to evaluate the repeatability and reliability. Statistical analysis was performed with SPSS 19.0. Results: Western blotting showed the treatment down-regulated Cdc42 overexpression. Quantitative analysis and two-tailed t-test showed that cell migration speed of the treated group was higher than the control group at all time points after treatment (p < 0.02). Conclusion: Combined treatment of 6MV photon and sevoflurane can cause the effects of down-regulating Cdc42 overexpression and decrease of migration speed of A549 cells which provides potential of clinical benefit for the cancer therapy. More investigation is needed to

  15. Expression of Fibroblast Growth Factor 9 in Normal Human Lung and Idiopathic Pulmonary Fibrosis

    PubMed Central

    Coffey, Emily; Newman, Donna R.

    2013-01-01

    The fibroblast growth factor (FGF) family of signaling ligands contributes significantly to lung development and maintenance in the adult. FGF9 is involved in control of epithelial branching and mesenchymal proliferation and expansion in developing lungs. However, its activity and expression in the normal adult lung and by epithelial and interstitial cells in fibroproliferative diseases like idiopathic pulmonary fibrosis (IPF) are unknown. Tissue samples from normal organ donor human lungs and those of a cohort of patients with mild to severe IPF were sectioned and stained for the immunolocalization of FGF9. In normal lungs, FGF9 was confined to smooth muscle surrounding airways, alveolar ducts and sacs, and blood vessels. In addition to these same sites, lungs of IPF patients expressed FGF9 in a population of myofibroblasts within fibroblastic foci, hypertrophic and hyperplastic epithelium of airways and alveoli, and smooth muscle cells surrounding vessels embedded in thickened interstitium. The results demonstrate that FGF9 protein increased in regions of active cellular hyperplasia, metaplasia, and fibrotic expansion of IPF lungs, and in isolated human lung fibroblasts treated with TGF-β1 and/or overexpressing Wnt7B. The cellular distribution and established biologic activity of FGF9 make it a potentially strong candidate for contributing to the progression of IPF. PMID:23797050

  16. Variations in Alveolar Partial Pressure for Carbon Dioxide and Oxygen Have Additive Not Synergistic Acute Effects on Human Pulmonary Vasoconstriction

    PubMed Central

    Croft, Quentin P. P.; Formenti, Federico; Talbot, Nick P.; Lunn, Daniel; Robbins, Peter A.; Dorrington, Keith L.

    2013-01-01

    The human pulmonary vasculature constricts in response to hypercapnia and hypoxia, with important consequences for homeostasis and adaptation. One function of these responses is to direct blood flow away from poorly-ventilated regions of the lung. In humans it is not known whether the stimuli of hypercapnia and hypoxia constrict the pulmonary blood vessels independently of each other or whether they act synergistically, such that the combination of hypercapnia and hypoxia is more effective than the sum of the responses to each stimulus on its own. We independently controlled the alveolar partial pressures of carbon dioxide (Paco 2) and oxygen (Pao 2) to examine their possible interaction on human pulmonary vasoconstriction. Nine volunteers each experienced sixteen possible combinations of four levels of Paco 2 (+6, +1, −4 and −9 mmHg, relative to baseline) with four levels of Pao 2 (175, 100, 75 and 50 mmHg). During each of these sixteen protocols Doppler echocardiography was used to evaluate cardiac output and systolic tricuspid pressure gradient, an index of pulmonary vasoconstriction. The degree of constriction varied linearly with both Paco 2 and the calculated haemoglobin oxygen desaturation (1-So 2). Mixed effects modelling delivered coefficients defining the interdependence of cardiac output, systolic tricuspid pressure gradient, ventilation, Paco 2 and So 2. No interaction was observed in the effects on pulmonary vasoconstriction of carbon dioxide and oxygen (p>0.64). Direct effects of the alveolar gases on systolic tricuspid pressure gradient greatly exceeded indirect effects arising from concurrent changes in cardiac output. PMID:23935847

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

    PubMed

    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. Copyright © 2015. Published by Elsevier Ltd.

  18. Influenza virus infection induces translocation of apoptosis-inducing factor (AIF) in A549 cells: role of AIF in apoptosis and viral propagation.

    PubMed

    Qu, Xinyan; Ding, Xiaoran; Duan, Ming; Yang, Jing; Lin, Ruxian; Zhou, Zhe; Wang, Shengqi

    2017-03-01

    It is recognized that influenza virus induces caspase-dependent apoptosis by activating caspase-3. Apoptosis-inducing factor (AIF) is a caspase-independent cell death effector, and its mitochondrial-nuclear translocation plays an important role in apoptosis. It is demonstrated in this study how influenza virus infection can induce caspase-independent apoptosis in the human alveolar epithelial cell line A549. AIF is translocated from the mitochondria to the nucleus in a caspase-independent manner in response to infection with influenza virus. Knockdown of AIF expression by small interfering RNA (siRNA) led to a reduction in virus-infection-induced apoptosis and virus yield. These results indicate that AIF translocation has a role in influenza-virus-induced apoptosis.

  19. Assessing the survival of MRC5 and a549 cell lines upon exposure to pyruvic Acid, sodium citrate and sodium bicarbonate - biomed 2013.

    PubMed

    Farah, Ibrahim O; Lewis, Veshell L; Ayensu, Wellington K; Cameron, Joseph A

    2013-01-01

    Lung cancer is among the most prevalent and deadly cancers in United States. In general, cancer cells are known to exhibit higher rates of glycolysis in comparison to normal cells. In attempting to exploit this unique cancer-dependent ATP generation phenomenon, it was our hypothesis that upon exposure to organic inhibitors of glycolysis, cancer cells would not survive normally and that their growth and viability would be vastly decreased; essential glycolytic ATP production will be exhausted to the point of collapsing energy utilization. Furthermore, we hypothesize that no negative effect would be seen with exposures to organic inhibitors for normal lung cells. The human lung fibroblast MRC-5 and the human A549 alveolar epithelial cell lines were used as in vitro models of normal lung and lung cancers respectively. Using standard methods, both cell lines were maintained and exposed to pyruvic acid, sodium citrate and sodium bicarbonate reagents at concentration levels ranging from 31.3-2,000 µg/ml in 96 well plates in quadruplets and experiments repeated at least three times using MTT, and cell counting (T4 Cellometer) assays as well as phase-contrast photo-imaging for parallel morphological displays of any changes in the course of their vitality and metabolic activities. Our results indicate that exposure of both cell lines to these organics resulted in concentration dependent cell destruction/cell survival depending on the cell line exposed. Pyruvic acid, sodium citrate and sodium bicarbonate showed statistically significant (p<0.05) differential negative effects on the A549 cell line in comparison to its unexposed control as well as to their effects on the MRC-5 cell line, presenting a potential promise for their use as cancer biotherapeutics.

  20. Preliminary Proteomic Analysis of A549 Cells Infected with Avian Influenza Virus H7N9 and Influenza A Virus H1N1

    PubMed Central

    Ding, Xiaoman; Lu, Jiahai; Yu, Ruoxi; Wang, Xin; Wang, Ting; Dong, Fangyuan; Peng, Bo; Wu, Weihua; Liu, Hui; Geng, Yijie; Zhang, Renli; Ma, Hanwu; Cheng, Jinquan; Yu, Muhua; Fang, Shisong

    2016-01-01

    A newly emerged H7N9 influenza virus poses high risk to human beings. However, the pathogenic mechanism of the virus remains unclear. The temporal response of primary human alveolar adenocarcinoma epithelial cells (A549) infected with H7N9 influenza virus and H1N1 influenza A virus (H1N1, pdm09) were evaluated using the proteomics approaches (2D-DIGE combined with MALDI-TOF-MS/MS) at 24, 48 and 72 hours post of the infection (hpi). There were 11, 12 and 33 proteins with significant different expressions (P<0.05) at 24, 48 and 72hpi, especially F-actin-capping protein subunit alpha-1 (CAPZA1), Ornithine aminotransferase (OAT), Poly(rC)-binding protein 1 (PCBP1), Eukaryotic translation initiation factor 5A-1 (EIF5A) and Platelet-activating factor acetylhydrolaseⅠb subunit beta (PAFAH1B2) were validated by western-blot analysis. The functional analysis revealed that the differential proteins in A549 cells involved in regulating cytopathic effect. Among them, the down-regulation of CAPZA1, OAT, PCBP1, EIF5A are related to the death of cells infected by H7N9 influenza virus. This is the first time show that the down-regulation of PAFAH1B2 is related to the later clinical symptoms of patients infected by H7N9 influenza virus. These findings may improve our understanding of pathogenic mechanism of H7N9 influenza virus in proteomics. PMID:27223893

  1. Regional pulmonary blood flow measurement in humans with electron-beam computed tomography

    NASA Astrophysics Data System (ADS)

    Holt, William W.; Konhilas, John; Wolfkiel, Christopher J.

    1995-05-01

    Electron beam computed tomography (EBCT) is a potentially useful modality to quantitate regional pulmonary flow (RPF) with minimal invasiveness, in part because it has good spatial and temporal resolution. The present studies used a single compartment model of indicator transport and EBCT to measure regional tissue flow in the lungs of human subjects. The model postulates that flow is proportional to maximal enhancement and assumes complete tissue accumulation of indicator before significant indicator washout (WO). EBCT flow studies were retrospectively analyzed with respect to RPF in 10 adult patients who had undergone clinically indicated or research cardiovascular studies. Time density curves from the left atrial (LA) cavity and one-third segments of left (LL) and right (RL) lungs (A: anterior, M: middle, and P: posterior segments) were used to calculate RPF. Washout was determined as the percent of the LA curve at the time of peak parenchymal opacification using gamma curve fits to both tissue data and the LA curve data. Mean +/- standard deviation RPF in ml/min/ml was 0.8 +/- 0.4, 1.1 +/- 0.4, and 1.3 +/- 0.4 for A, M, and P respectively for one-third regions in the left lung. Similar results were found in the right lung. No difference in RPF was found when images were measured either by including the largest of visible parenchymal vessels or when such vessels were excluded. Flow in A of LL and RL was less than that in M or P. Average WO was about 10%, with a range of 0-41% of the LA curve area. There was no significant difference between one-third segment WO using pairwise comparison on the left and right sides when tested separately. RPF values were greater in the posterior vs anterior regions of these supine patients. In conclusion, EBCT can detect gravity related flow differences in the human lung. EBCT has potential for clinical assessment of absolute regional pulmonary flow determination in animals and man.

  2. Tumor necrosis factor-{alpha} induces MMP-9 expression via p42/p44 MAPK, JNK, and nuclear factor-{kappa}B in A549 cells

    SciTech Connect

    Lin, C.-C.; Tseng, Hsiao-Wei; Hsieh, Hsi-Lung

    2008-06-15

    Matrix metalloproteinases (MMPs), in particular MMP-9, have been shown to be induced by cytokines including tumor necrosis factor-{alpha} (TNF-{alpha}) and contributes to airway inflammation. However, the mechanisms underlying MMP-9 expression induced by TNF-{alpha} in human A549 cells remain unclear. Here, we showed that TNF-{alpha} induced production of MMP-9 protein and mRNA is determined by zymographic, Western blotting, RT-PCR and ELISA assay, which were attenuated by inhibitors of MEK1/2 (U0126), JNK (SP600125), and NF-{kappa}B (helenalin), and transfection with dominant negative mutants of ERK2 ({delta}ERK) and JNK ({delta}JNK), and siRNAs for MEK1, p42 and JNK2. TNF-{alpha}-stimulated phosphorylation of p42/p44 MAPK and JNKmore » were attenuated by pretreatment with the inhibitors U0126 and SP600125 or transfection with dominant negative mutants of {delta}ERK and {delta}JNK. Furthermore, the involvement of NF-{kappa}B in TNF-{alpha}-induced MMP-9 production was consistent with that TNF-{alpha}-stimulated degradation of I{kappa}B-{alpha} and translocation of NF-{kappa}B into the nucleus which were blocked by helenalin, but not by U0126 and SP600125, revealed by immunofluorescence staining. The regulation of MMP-9 gene transcription by MAPKs and NF-{kappa}B was further confirmed by gene luciferase activity assay. MMP-9 promoter activity was enhanced by TNF-{alpha} in A549 cells transfected with wild-type MMP-9-Luc, which was inhibited by helenalin, U0126, or SP600125. In contrast, TNF-{alpha}-stimulated MMP-9 luciferase activity was totally lost in cells transfected with mutant-NF-{kappa}B MMP-9-luc. Moreover, pretreatment with actinomycin D and cycloheximide attenuated TNF-{alpha}-induced MMP-9 expression. These results suggest that in A549 cells, phosphorylation of p42/p44 MAPK, JNK, and transactivation of NF-{kappa}B are essential for TNF-{alpha}-induced MMP-9 gene expression.« less

  3. Cinchonine induces apoptosis of HeLa and A549 cells through targeting TRAF6.

    PubMed

    Qi, Yonghao; Pradipta, Ambara R; Li, Miao; Zhao, Xuan; Lu, Lulu; Fu, Xuegang; Wei, Jing; Hsung, Richard P; Tanaka, Katsunori; Zhou, Lijun

    2017-02-23

    Cancer cells are known to over-express TRAF6 that is critical for both AKT and TAK1 activations. The Really Interesting New Gene (RING) domain of TRAF6 is believed to be responsible for the E3 ligase activity, ZINC fingers of TRAF6 provide critical support for the activity of the RING domain which is critical for both AKT and TAK1 activations. We employed computational docking program to identify small molecules that could effectively and competitively bind with the RING domain of TRAF6, which is believed to be responsible for its E3 ligase activity. MTT assay and flow cytometry were employed to analyze apoptosis of cancer cells. Signaling pathways were detected using immunoprecipitation and western blotting, and immunofluorescence was pursued to assess the nature of binding of cinchonine to TRAF6. We also performed animal experiments to test effect of cinchonine in vivo. Cinchonine, a naturally occurring Cinchona alkaloid identified from the docking study, could bind to TRAF6 in HeLa and A549 cells and induce apoptosis of these cancer cells. We found that AKT ubiquitination and phosphorylation as well as phosphorylation of TAK1 were decreased. These activities would lead to subsequent suppression anti-apoptotic protein Bcl-2, while elevating pro-apoptotic protein Bax. Immunofluorescence staining unambiguously demonstrated the binding of cinchonine specifically at the RING domain of TRAF6 in cells, thereby validating the computational modeling. Animal experiments showed that cinchonine could suppress tumor growth in mice without showing significant acute toxicity. These investigations suggest that through competitive binding with the RING domain of TRAF6, cinchonine could induce apoptosis via inhibiting AKT and TAK1 signaling pathways.

  4. [Pulmonary arterial hypertension associated with human immunodeficiency virus infection: study of 4 cases].

    PubMed

    Pousada, Guillermo; Baloira, Adolfo; Castro-Añón, Olalla; Valverde, Diana

    2016-04-15

    Pulmonary arterial hypertension (PAH) is a rare and progressive disease that can be inherited as autosomal dominant form. The BMPR2, ACVRL1 and ENG genes are main genes involved in the pathology. PAH associated to human immunodeficiency virus (HIV) is another rare disease with a low incidence, prevalence and survival. The main objective of this analysis was to study the clinical and molecular characteristics of PAH associated to HIV patients. We present 4 cases of HIV patients who developed PAH and have been treated with ambrisentan. Pathogenic mutations have been identify in analyzed genes in 3 of the four analyzed patients. In addition, these patients present other changes classified as benign after a thorough in silico analysis. We identified some changes in genetic modifiers that predispose to these patients to more severe phenotype. The clinical analysis can help to define monitoring for these patients and the administration of appropriate treatment. These patients also have shown several pathogenic mutations. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

  5. Atrial electrical and structural remodeling associated with longstanding pulmonary hypertension and right ventricular hypertrophy in humans.

    PubMed

    Medi, Caroline; Kalman, Jonathan M; Ling, Liang-Han; Teh, Andrew W; Lee, Geoffrey; Lee, Geraldine; Spence, Steven J; Kaye, David M; Kistler, Peter M

    2012-06-01

    Pulmonary hypertension (PH) is common to a range of cardiopulmonary conditions and is associated with atrial arrhythmias. However, little is known of the isolated atrial effects of PH and right atrial dilatation (RA) in humans. To avoid the confounding effects of PH-associated disease states, we performed detailed electrophysiological (EP) and electroanatomic (EA) mapping of the RA in patients with idiopathic PH. Eight PH patients (mean pulmonary arterial [PA] pressure 39.0 ± 15.8 mmHg) and 16 age-matched controls (mean PA pressure 11.5 ± 4.1 mmHg, P < 0.0001) were studied. Corrected sinus node recovery times (cSNRT), atrial effective refractory periods (ERPs), conduction delay at the crista terminalis (CT), and inducibility of atrial fibrillation (AF) were evaluated. EA mapping (pacing cycle length 600 and 300 milliseconds) was performed to determine RA global and regional voltage, conduction velocities, atrial activation times, fractionated electrograms and double potentials. Patients with PH demonstrated a prolongation in cSNRT without significant change in atrial ERP and an increase in AF inducibility. PH was associated with lower tissue voltage (1.8 ± 0.4 mV in PH vs 2.2 ± 0.4 mV in controls, P = 0.02), increased low voltage areas (13.7 ± 8.2% in PH vs 6.2 ± 3.7% in controls, P < 0.01) and the presence of electrically silent areas. Conduction velocities were slower (global 67.3 ± 5.6 cm/s vs 92.8 ± 4.0 cm/s, P < 0.001) and fractionated electrograms and double potentials were more prevalent (14.7 ± 4.4% vs 6.3 ± 4.1, P < 0.01) in PH compared with controls, respectively. Idiopathic PH is associated with RA remodeling characterized by: generalized conduction slowing with marked regional abnormalities; reduced tissue voltage; and regions of electrical silence. These changes provide important insights into the isolated effects of PH fundamental to a range of clinical conditions associated with AF. © 2012 Wiley Periodicals, Inc.

  6. Increase in cell motility by carbon ion irradiation via the Rho signaling pathway and its inhibition by the ROCK inhibitor Y-27632 in lung adenocarcinoma A549 cells.

    PubMed

    Murata, Kazutoshi; Noda, Shin-ei; Oike, Takahiro; Takahashi, Akihisa; Yoshida, Yukari; Suzuki, Yoshiyuki; Ohno, Tatsuya; Funayama, Tomoo; Kobayashi, Yasuhiko; Takahashi, Takeo; Nakano, Takashi

    2014-07-01

    This study aimed to investigate the effect of carbon ion (C-ion) irradiation on cell motility through the ras homolog gene family member (Rho) signaling pathway in the human lung adenocarcinoma cell line A549. Cell motility was assessed by a wound-healing assay, and the formation of cell protrusions was evaluated by F-actin staining. Cell viability was examined by the WST-1 assay. The expression of myosin light chain 2 (MLC2) and the phosphorylation of MLC2 at Ser19 (P-MLC2-S19) were analyzed by Western blot. At 48 h after irradiation, the wound-healing assay demonstrated that migration was significantly greater in cells irradiated with C-ion (2 or 8 Gy) than in unirradiated cells. Similarly, F-actin staining showed that the formation of protrusions was significantly increased in cells irradiated with C-ion (2 or 8 Gy) compared with unirradiated cells. The observed increase in cell motility due to C-ion irradiation was similar to that observed due to X-ray irradiation. Western-blot analysis showed that C-ion irradiation (8 Gy) increased P-MLC2-S19 expression compared with in unirradiated controls, while total MLC2 expression was unchanged. Exposure to a non-toxic concentration of Y-27632, a specific inhibitor of Rho-associated coiled-coil-forming protein kinase (ROCK), reduced the expression of P-MLC2-S19 after C-ion irradiation (8 Gy), resulting in a significant reduction in migration. These data suggest that C-ion irradiation increases cell motility in A549 cells via the Rho signaling pathway and that ROCK inhibition reduces that effect. © The Author 2014. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.

  7. Effect of three fatty acids from the leaf extract of Tiliacora triandra on P-glycoprotein function in multidrug-resistant A549RT-eto cell line

    PubMed Central

    Kaewpiboon, Chutima; Winayanuwattikun, Pakorn; Yongvanich, Tikamporn; Phuwapraisirisan, Preecha; Assavalapsakul, Wanchai

    2014-01-01

    Background: Cancer cells have the ability to develop resistance to chemotherapy drugs, which then leads to a reduced effectiveness and success of the treatment. Multidrug resistance (MDR) involves the resistance in the same cell/tissue to a diverse range of drugs of different structures. One of the characteristics of MDR is an overexpression of P-glycoprotein (P-gp), which causes the efflux of the accumulated drug out of the cell. The MDR human non-small cell lung carcinoma cell line with a high P-gp expression level (A549RT-eto) was used to investigate the bioactive compounds capable of reversing the etoposide resistance in this cell line. Materials and Methods: The leaves of Tiliacora triandra were sequentially extracted with hexane, dichloromethane, methanol and water. Only the hexane extract reduced the etoposide resistance of the A549RT-eto cell line, and was further fractionated by column chromatography using the TLC-pattern and the restoration of etoposide sensitivity as the selection criteria. Results: The obtained active fraction (F22) was found by nuclear magnetic resonance and gas chromatography-mass spectroscopy analyses to be comprised of a 49.5:19.6:30.9 (w/w/w) mixture of hexadecanoic: octadecanoic acid: (Z)-6-octadecenoic acids. This stoichiometric mixture was recreated using pure fatty acids (MSFA) and gave a similar sensitization to etoposide and enhanced the relative rate of rhodamine-123 accumulation to a similar extent as F22, supporting the action via reducing P-gp activity. In contrast, the fatty acids alone did not show this effect. Conclusion: This is the first report of the biological activity from the leaves of T. triandra as a potential source of a novel chemosensitizer. PMID:25298673

  8. Effect of three fatty acids from the leaf extract of Tiliacora triandra on P-glycoprotein function in multidrug-resistant A549RT-eto cell line.

    PubMed

    Kaewpiboon, Chutima; Winayanuwattikun, Pakorn; Yongvanich, Tikamporn; Phuwapraisirisan, Preecha; Assavalapsakul, Wanchai

    2014-08-01

    Cancer cells have the ability to develop resistance to chemotherapy drugs, which then leads to a reduced effectiveness and success of the treatment. Multidrug resistance (MDR) involves the resistance in the same cell/tissue to a diverse range of drugs of different structures. One of the characteristics of MDR is an overexpression of P-glycoprotein (P-gp), which causes the efflux of the accumulated drug out of the cell. The MDR human non-small cell lung carcinoma cell line with a high P-gp expression level (A549RT-eto) was used to investigate the bioactive compounds capable of reversing the etoposide resistance in this cell line. The leaves of Tiliacora triandra were sequentially extracted with hexane, dichloromethane, methanol and water. Only the hexane extract reduced the etoposide resistance of the A549RT-eto cell line, and was further fractionated by column chromatography using the TLC-pattern and the restoration of etoposide sensitivity as the selection criteria. The obtained active fraction (F22) was found by nuclear magnetic resonance and gas chromatography-mass spectroscopy analyses to be comprised of a 49.5:19.6:30.9 (w/w/w) mixture of hexadecanoic: octadecanoic acid: (Z)-6-octadecenoic acids. This stoichiometric mixture was recreated using pure fatty acids (MSFA) and gave a similar sensitization to etoposide and enhanced the relative rate of rhodamine-123 accumulation to a similar extent as F22, supporting the action via reducing P-gp activity. In contrast, the fatty acids alone did not show this effect. This is the first report of the biological activity from the leaves of T. triandra as a potential source of a novel chemosensitizer.

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

    PubMed

    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 < 0.01, sensitization enhancement ratio in CNE-1 increased 1.32-fold to 1.61-fold, p < 0.05). We explored the mechanisms for the radiosensitization efficiency and the difference between docetaxel and docetaxel-loaded poly(lactic-co-glycolic acid) (PLGA) 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. © The Author(s) 2015.

  10. MicroRNA-9 and Cell Proliferation in Lipopolysaccharide and Dexamethasone-Treated Naïve and Desialylated A549 Cells Grown in Cigarette Smoke Conditioned Medium.

    PubMed

    Holownia, A; Wielgat, P; Eljaszewicz, A

    2018-03-01

    In this study we assessed microRNA-9 (miR-9) levels (RT-PCR) and cell proliferation (flow cytometry) in naïve and desialylated human alveolar epithelial cells (A549 cells), grown for 24 h in cigarette smoke-conditioned medium. Cells were additionally treated with lipopolysaccharide (LPS) and/or dexamethasone. Proliferation positively correlated with miR-9 levels in both naïve and desialylated cells. Cigarette smoke decreased miR-9 levels in both cell types by about three-fold but there was no significant correlation between both parameters. Dexamethasone was without substantial effect on cigarette smoke-induced changes in proliferation of naïve cells, but some normalization was observed in desialylated cells. Dexamethasone increased miR-9 levels in both cell types grown in cigarette smoke-medium but the effect was stronger in desialylated cells. LPS increased cell proliferation and miR-9 by more than six-fold only in naïve cells, while correlation coefficient for both parameters in cigarette smoke-LPS group was 0.41. Herein we identify miR-9 as the cigarette smoke (decrease) and LPS-responsive but dexamethasone-unresponsive microRNA. It is possible that increased miR-9 levels in naïve A549 cells treated with LPS may be related to the activation of Toll-like receptor 4. Moreover, differences in cell response (both miR-9 and proliferation) to dexamethasone in naïve and desialylated cells may point to non-genomic dexamethasone effects.

  11. Properties and inflammatory effects of various size fractions of ambient particulate matter from Beijing on A549 and J774A.1 cells.

    PubMed

    Wang, Bin; Li, Kexin; Jin, Wenjie; Lu, Yan; Zhang, Yuzhong; Shen, Guofeng; Wang, Rong; Shen, Huizhong; Li, Wei; Huang, Ye; Zhang, Yanyan; Wang, Xilong; Li, Xiqing; Liu, Wenxin; Cao, Hongying; Tao, Shu

    2013-09-17

    Particulate matter (PM) is a major ambient air pollutant causing millions of premature deaths each year in China. The toxicity of PM is property and size dependent. In this study, ambient PM samples collected in Beijing were divided into five size fractions with nominal aerodynamic ranges of <0.40, 0.40-1.1, 1.1-3.3, 3.3-5.8, and 5.8-10 μm. Individual size fractions were characterized for a number of properties including particle size distribution, specific surface area, zeta potential, dithiothreitol (DTT)-based redox ability, and contents of water-soluble organic carbon (WSOC), polycyclic aromatic hydrocarbons (PAHs), selected metals, and endotoxin. Human adenocarcinomic alveolar epithelial cell line A549 and small mouse monocyte-macrophage cell line J774A.1 were tested for their relative viabilities and inflammatory effects (interleukine-8 for A549 and tumor necrosis factor-α for J774A.1) after exposure to PM of various sizes. It was found that PM specific area was positively correlated with WSOC, high molecular weight PAHs, DTT-based redox ability, negatively correlated with surface zeta potential and lithophile metals. Several trace metals from combustion sources were enriched in intermediate size fractions. For both endotoxin concentrations of the PM and PM induced inflammatory cytokine expressions by the two cell lines, there were general increasing trends as PM size increased with an exception of the finest fraction, which induced the highest inflammatory effects. It seems that the size dependence of cytokine expression was associated with a number of properties including endotoxin content, zeta potential, settling velocity, metal content, and DTT-based redox ability.

  12. A confocal microscopic study of solitary pulmonary neuroendocrine cells in human airway epithelium.

    PubMed

    Weichselbaum, Markus; Sparrow, Malcolm P; Hamilton, Elisha J; Thompson, Philip J; Knight, Darryl A

    2005-10-10

    Pulmonary neuroendocrine cells (PNEC) are specialized epithelial cells that are thought to play important roles in lung development and airway function. PNEC occur either singly or in clusters called neuroepithelial bodies. Our aim was to characterize the three dimensional morphology of PNEC, their distribution, and their relationship to the epithelial nerves in whole mounts of adult human bronchi using confocal microscopy. Bronchi were resected from non-diseased portions of a lobe of human lung obtained from 8 thoracotomy patients (Table 1) undergoing surgery for the removal of lung tumors. Whole mounts were stained with antibodies to reveal all nerves (PGP 9.5), sensory nerves (calcitonin gene related peptide, CGRP), and PNEC (PGP 9.5, CGRP and gastrin releasing peptide, GRP). The analysis and rendition of the resulting three-dimensional data sets, including side-projections, was performed using NIH-Image software. Images were colorized and super-imposed using Adobe Photoshop. PNEC were abundant but not homogenously distributed within the epithelium, with densities ranging from 65/mm2 to denser patches of 250/mm2, depending on the individual wholemount. Rotation of 3-D images revealed a complex morphology; flask-like with the cell body near the basement membrane and a thick stem extending to the lumen. Long processes issued laterally from its base, some lumenal and others with feet-like processes. Calcitonin gene-related peptide (CGRP) was present in about 20% of PNEC, mainly in the processes. CGRP-positive nerves were sparse, with some associated with the apical part of the PNEC. Our 3D-data demonstrates that PNEC are numerous and exhibit a heterogeneous peptide content suggesting an active and diverse PNEC population.

  13. Activation of endothelial and epithelial KCa2.3 calcium-activated potassium channels by NS309 relaxes human small pulmonary arteries and bronchioles

    PubMed Central

    Kroigaard, Christel; Dalsgaard, Thomas; Nielsen, Gorm; Laursen, Britt E; Pilegaard, Hans; Köhler, Ralf; Simonsen, Ulf

    2012-01-01

    BACKGROUND AND PURPOSE Small (KCa2) and intermediate (KCa3.1) conductance calcium-activated potassium channels (KCa) may contribute to both epithelium- and endothelium-dependent relaxations, but this has not been established in human pulmonary arteries and bronchioles. Therefore, we investigated the expression of KCa2.3 and KCa3.1 channels, and hypothesized that activation of these channels would produce relaxation of human bronchioles and pulmonary arteries. EXPERIMENTAL APPROACH Channel expression and functional studies were conducted in human isolated small pulmonary arteries and bronchioles. KCa2 and KCa3.1 currents were examined in human small airways epithelial (HSAEpi) cells by whole-cell patch clamp techniques. RESULTS While KCa2.3 expression was similar, KCa3.1 protein was more highly expressed in pulmonary arteries than bronchioles. Immunoreactive KCa2.3 and KCa3.1 proteins were found in both endothelium and epithelium. KCa currents were present in HSAEpi cells and sensitive to the KCa2.3 blocker UCL1684 and the KCa3.1 blocker TRAM-34. In pulmonary arteries contracted by U46619 and in bronchioles contracted by histamine, the KCa2.3/ KCa3.1 activator, NS309, induced concentration-dependent relaxations. NS309 was equally potent in relaxing pulmonary arteries, but less potent in bronchioles, than salbutamol. NS309 relaxations were blocked by the KCa2 channel blocker apamin, while the KCa3.1 channel blocker, charybdotoxin failed to reduce relaxation to NS309 (0.01–1 µM). CONCLUSIONS AND IMPLICATIONS KCa2.3 and KCa3.1 channels are expressed in the endothelium of human pulmonary arteries and epithelium of bronchioles. KCa2.3 channels contributed to endo- and epithelium-dependent relaxations suggesting that these channels are potential targets for treatment of pulmonary hypertension and chronic obstructive pulmonary disease. PMID:22506557

  14. Acute Meteorite Dust Exposure and Pulmonary Inflammation - Implications for Human Space Exploration

    NASA Technical Reports Server (NTRS)

    Harrington, A. D.; McCubbin, F. M.; Kaur, J.; Smirnov, A.; Galdanes, K.; Schoonen, M. A. A.; Chen, L. C.; Tsirka, S. E.; Gordon, T.

    2017-01-01

    The previous manned missions to the Moon represent milestones of human ingenuity, perseverance, and intellectual curiosity. However, one of the major ongoing concerns is the array of hazards associated with lunar surface dust. Not only did the dust cause mechanical and structural integrity issues with the suits, the dust 'storm' generated upon reentrance into the crew cabin caused "lunar hay fever" and "almost blindness [1-3]" (Figure 1). It was further reported that the allergic response to the dust worsened with each exposure [4]. The lack of gravity exacerbated the exposure, requiring the astronauts to wear their helmet within the module in order to avoid breathing the irritating particles [1]. Due to the prevalence of these high exposures, the Human Research Roadmap developed by NASA identifies the Risk of Adverse Health and Performance Effects of Celestial Dust Exposure as an area of concern [5]. Extended human exploration will further increase the probability of inadvertent and repeated exposures to celestial dusts. Going forward, hazard assessments of celestial dusts will be determined through sample return efforts prior to astronaut deployment. Studies on the lunar highland regolith indicate that the dust is not only respirable but also reactive [2, 6-9], and previous studies concluded that it is moderately toxic; generating a greater response than titanium oxide but a lower response than quartz [6]. The presence of reactive oxygen species (ROS) on the surface of the dust has been implicated. However, there is actually little data related to physicochemical characteristics of particulates and pulmonary toxicity, especially as it relates to celestial dust exposure. As a direct response to this deficit, the present study evaluates the role of a particulate's innate geochemical features (e.g., bulk chemistry, internal composition, morphology, size, and reactivity) in generating adverse toxicological responses in vitro and in vivo. This highly interdisciplinary

  15. Pulmonary Inflammatory Responses To Acute Meteorite Dust Exposures - Implications For Human Space Exploration

    NASA Technical Reports Server (NTRS)

    Harrington, A. D.; McCubbin, F. M.; Kaur, J.; Smirnov, A.; Galdanes, K.; Schoonen, M. A. A.; Chen, L. C.; Tsirka, S. E.; Gordon, T.

    2017-01-01

    The previous manned missions to the Moon represent milestones of human ingenuity, perseverance, and intellectual curiosity. However, one of the major ongoing concerns is the array of hazards associated with lunar surface dust. Not only did the dust cause mechanical and structural integrity issues with the suits, the dust 'storm' generated upon reentrance into the crew cabin caused "lunar hay fever" and "almost blindness" (Figure 1). It was further reported that the allergic response to the dust worsened with each exposure. The lack of gravity exacerbated the exposure, requiring the astronauts to wear their helmet within the module in order to avoid breathing the irritating particles. Due to the prevalence of these high exposures, the Human Research Roadmap developed by NASA identifies the Risk of Adverse Health and Performance Effects of Celestial Dust Exposure as an area of concern. Extended human exploration will further increase the probability of inadvertent and repeated exposures to celestial dusts. Going forward, hazard assessments of celestial dusts will be determined through sample return efforts prior to astronaut deployment. Studies on the lunar highland regolith indicate that the dust is not only respirable but also reactive, and previous studies concluded that it is moderately toxic; generating a greater response than titanium oxide but a lower response than quartz. The presence of reactive oxygen species (ROS) on the surface of the dust has been implicated. However, there is actually little data related to physicochemical characteristics of particulates and pulmonary toxicity, especially as it relates to celestial dust exposure. As a direct response to this deficit, the present study evaluates the role of a particulate's innate geochemical features (e.g., bulk chemistry, internal composition, morphology, size, and reactivity) in generating adverse toxicological responses in vitro and in vivo. This highly interdisciplinary study evaluates the relative

  16. Inhibition of growth and induction of apoptosis in A549 cells by compounds from oxheart cabbage extract.

    PubMed

    Wang, Nan; Wang, Wei; Liu, Caiqin; Jin, Jianchang; Shao, Bo; Shen, Lianqing

    2016-08-01

    Oxheart cabbage (Brassica oleracea var. capitata) is a member of the Brassica genus. Although some studies on the anticancer effects of extracts from oxheart cabbage have been reported, comprehensive information on the bioactive fractions and components from oxheart cabbage extracts is still lacking. The aim of this study was to isolate and identify the bioactive fractions and components from oxheart cabbage seeds using activity-guided isolation methods. The cytotoxicity and apoptotic effects of fraction II, fraction III, iberverin, sulforaphane and iberin from oxheart cabbage seed extract were investigated. The results showed that all five components had inhibitory effects on the in vitro growth of A549 cells which were dose-dependent. These compounds also changed the morphology of A549 cells, and their inhibitory activity on A549 cells was as follows: sulforaphane > iberin > iberverin > fraction III > fraction II. The IC50 values were 3.53 ± 0.63, 4.93 ± 1.02, 7.07 ± 0.51, 15.56 ± 0.24 and 27.32 ± 0.63 µg mL(-1) respectively. Fraction II, fraction III, iberverin, sulforaphane and iberin induced cell apoptosis by increasing early apoptosis and late apoptosis/necrosis, and activation of caspase-3, -8 and -9. These results indicated that the decrease in A549 cell viability by active compounds from oxheart cabbage seed extract was due to the induction of apoptosis. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

  17. Oligomeric proanthocyanidins protects A549 cells against H2O2-induced oxidative stress via the Nrf2-ARE pathway.

    PubMed

    Sun, Chao; Jin, Weiguo; Shi, Hongcan

    2017-06-01

    Oxidative signaling and oxidative stress contribute to aging, cancer and diseases resulting from lung fibrosis. In this study, we explored the anti-oxidative potential of oligomeric proanthocyanidins (OPCs), natural flavonoid compounds. We examined the protective effects of OPCs against hydrogen peroxide (H2O2)-induced oxidative stress in non-small cell lung cancer cells (A549). We demonstrated that OPC markedly attenuated H2O2-induced A549 cell viability, as shown by by 3-[4,5-dimethylthiazol-2-yl)]-2,5-diphenyl-tetrazolium bromide (MTT) assay. At the same time, OPC inhibited H2O2-induced oxidative stress by significantly increasing the activities of superoxide dismutase, catalase and glutathione, and reducing the levels of reactive oxygen species (ROS) and malondialdehyde (MDA). Treatment of the A549 cells with OPC significantly promoted the nuclear translocation of NF-E2-related factor 2 (Nrf2) and significantly enhanced the expression of its target genes [heme oxygenase-1 (HO-1), NAD(P)H quinone dehydrogenase 1 (NQO1) and thioredoxin reductase 1 (TXNRD1)] with different fold change values at both the mRNA and protein level. The knockout of Nrf2 using CRISPR/Cas9 technology attenuated OPC-mediated ARE gene transcription, and almost abolished the OPC-mediated protective effects against H2O2-induced oxidative stress. On the whole, our study suggests that OPC plays an important role in controlling the antioxidant response of A549 cells via the Nrf2-ARE pathway.

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

    SciTech Connect

    Liu, Juntao; Mao, Zhangfan; Huang, Jie

    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 treatmentsmore » 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

  19. Discovery of novel HSP90 inhibitors that induced apoptosis and impaired autophagic flux in A549 lung cancer cells.

    PubMed

    Wei, Qun; Ning, Jun-Ya; Dai, Xi; Gao, Yuan-Di; Su, Le; Zhao, Bao-Xiang; Miao, Jun-Ying

    2018-02-10

    Heat shock protein 90 (HSP90) inhibition has aroused increasing enthusiasm in antitumor strategies in recent years. According to our previous studies, we synthesized a series of coumarin pyrazoline compounds HCP1-HCP6 that might be HSP90 inhibitors. Interactions between HCP1-HCP6 and HSP90 were examined and antitumor activities of them were investigated in A549 lung cancer cells. Results showed that all the six derivatives could interact with HSP90, in which HCP1 exhibited the best binding ability and inhibited the activity of HSP90. Meanwhile, HCP1-HCP6 reduced the cell viability of A549 cells and HCP1 possessed the lowest IC 50 value. Above all HCP1 exerted better HSP90 inhibitory and anticancer effects than our initially identified HSP90 inhibitor DPB. As to the underlying mechanism, HCP1-HCP6 not only induced apoptosis as DPB but also blocked autophagic flux in A549 cells. Therefore, we discovered a novel HSP90 inhibitor HCP1 that had better biological activity and provided us a useful tool to explore the underlying mechanism of lung cancer therapy. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

  20. Effects of Green Tea Extract on Lung Cancer A549 Cells: Proteomic Identification of Proteins Associated with Cell Migration

    PubMed Central

    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

    2009-01-01

    Green tea polyphenols exhibit multiple anti-tumor 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 two-dimensional gel electrophoresis LC-tandem mass spectrometry of A549 cells before and after GTE exposure. We have identified 14 protein spots that changed in expression (≥2 fold) after GTE treatment. These proteins are involved in calcium-binding, cytoskeleton and motility, metabolism, detoxification or gene regulation. In particular we found up-regulation of several genes that modulate actin remodeling and cell migration, including lamin A/C. Our data indicated that GTE-induced lamin A/C up-regulation appears to be at the transcriptional level and the increased expression results in the decrease in the 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 anti-tumor activities of GTE. PMID:19137550

  1. [Effect of ginsenoside on the cellular proliferation, apoptosis and cell cycles in LC A549 and HUVEC 304 cell lines].

    PubMed

    Chen, Ming-wei; Ma, Ai-qun; Ni, Lei; Huang, Chen; Zhang, Dian-zeng; Niu, Xiao-ying

    2005-04-01

    To determine the effect of ginsenoside on the cellular proliferation, apoptosis and cell cycles in LC A549 and HUVEC 304 cell lines. A549 and HUVEC 304 cell lines were cultured with different concentrations of ginsenoside. Cellular proliferation was detected with MTT, apoptosis and cell cycles were checked with Flow Cytometer, and change of microstructure was observed by transmission electron microscope. The apoptosis rate was 29.8% in A549 cell lines after being interfered with ginsenoside at 3 x 10(-6) mol/L, significantly higher than that in the control group ( P < 0.05). No change was observed in the cell cycles after being interfered with ginsenoside. The inhibitive rate of ginsenoside was 12.53% for HUVEC 304 cell line at 1 x 10(-4) mol/L (P < 0.05 ). The cells induced by conditioned medium could be inhibited by ginsenoside, and apoptotic body could be found in cells induced by conditioned medium at 10(-6) mol/L. The proliferation of vascular endothelial cell could be inhibited by ginsenoside, and apoptosis could also be found in both tumor cells and cells induced by conditioned medium after being interfered with ginsenoside.

  2. Pulmonary angiography

    MedlinePlus

    The test is used to detect blood clots ( pulmonary embolism ) and other blockages in the blood flow in ... of pulmonary vessels Blood clot in the lungs (pulmonary embolism) Narrowed blood vessel Primary pulmonary hypertension Tumor in ...

  3. BZML, a novel colchicine binding site inhibitor, overcomes multidrug resistance in A549/Taxol cells by inhibiting P-gp function and inducing mitotic catastrophe.

    PubMed

    Bai, Zhaoshi; Gao, Meiqi; Zhang, Huijuan; Guan, Qi; Xu, Jingwen; Li, Yao; Qi, Huan; Li, Zhengqiang; Zuo, Daiying; Zhang, Weige; Wu, Yingliang

    2017-08-28

    Multidrug resistance (MDR) interferes with the efficiency of chemotherapy. Therefore, developing novel anti-cancer agents that can overcome MDR is necessary. Here, we screened a series of colchicine binding site inhibitors (CBSIs) and found that 5-(3, 4, 5-trimethoxybenzoyl)-4-methyl-2-(p-tolyl) imidazol (BZML) displayed potent cytotoxic activity against both A549 and A549/Taxol cells. We further explored the underlying mechanisms and found that BZML caused mitosis phase arrest by inhibiting tubulin polymerization in A549 and A549/Taxol cells. Importantly, BZML was a poor substrate for P-glycoprotein (P-gp) and inhibited P-gp function by decreasing P-gp expression at the protein and mRNA levels. Cell morphology changes and the expression of cycle- or apoptosis-related proteins indicated that BZML mainly drove A549/Taxol cells to die by mitotic catastrophe (MC), a p53-independent apoptotic-like cell death, whereas induced A549 cells to die by apoptosis. Taken together, our data suggest that BZML is a novel colchicine binding site inhibitor and overcomes MDR in A549/Taxol cells by inhibiting P-gp function and inducing MC. Our study also offers a new strategy to solve the problem of apoptosis-resistance. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Targeting Interleukin-13 with Tralokinumab Attenuates Lung Fibrosis and Epithelial Damage in a Humanized SCID Idiopathic Pulmonary Fibrosis Model

    PubMed Central

    Zhang, Huilan; Oak, Sameer R.; Coelho, Ana Lucia; Herath, Athula; Flaherty, Kevin R.; Lee, Joyce; Bell, Matt; Knight, Darryl A.; Martinez, Fernando J.; Sleeman, Matthew A.; Herzog, Erica L.; Hogaboam, Cory M.

    2014-01-01

    The aberrant fibrotic and repair responses in the lung are major hallmarks of idiopathic pulmonary fibrosis (IPF). Numerous antifibrotic strategies have been used in the clinic with limited success, raising the possibility that an effective therapeutic strategy in this disease must inhibit fibrosis and promote appropriate lung repair mechanisms. IL-13 represents an attractive target in IPF, but its disease association and mechanism of action remains unknown. In the present study, an overexpression of IL-13 and IL-13 pathway markers was associated with IPF, particularly a rapidly progressive form of this disease. Targeting IL-13 in a humanized experimental model of pulmonary fibrosis using tralokinumab (CAT354) was found to therapeutically block aberrant lung remodeling in this model. However, targeting IL-13 was also found to promote lung repair and to restore epithelial integrity. Thus, targeting IL-13 inhibits fibrotic processes and enhances repair processes in the lung. PMID:24325475

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

    PubMed

    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. © 2016 Wiley Periodicals, Inc.

  6. Pulmonary surfactant inhibits activation of human monocytes by recombinant interferon-gamma.

    PubMed Central

    Geertsma, M F; Teeuw, W L; Nibbering, P H; van Furth, R

    1994-01-01

    During an inflammatory reaction in the alveoli, the functional activities of monocytes, macrophages and granulocytes are regulated by a complex network of inflammatory mediators. The primary cytokine involved in activation of these phagocytes is interferon-gamma (IFN-gamma). The possible influence of local factors, such as pulmonary surfactant, on the activation process has not been studied until now. The aim of the present study was to investigate the effects of surfactant on the activation of monocytes by recombinant (r)IFN-gamma. The results revealed that human surfactant significantly inhibited both the increase in the expression of the high-affinity receptor for IgG, i.e. Fc gamma RI, and the production of H2O2 by rIFN-gamma-activated monocytes. Since our surfactant preparation stimulated the basal production of tumour necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) by monocytes, the effect of Survanta, a surfactant extract, on the rIFN-gamma-induced production of these cytokines by monocytes was studied. The results revealed that Survanta caused 80-90% inhibition of the rIFN-gamma-induced production of TNF-alpha and IL-1 beta by these cells. Together, these results could mean that surfactant is involved in the protection of the alveolar epithelium against injury caused by reactive oxygen intermediates (ROI) and TNF-alpha, and in the down-regulation of the production of inflammatory mediators. In view of these considerations, surfactant therapy may not only improve lung compliance and gas exchange but may also be beneficial in reducing the inflammatory reaction in the lungs. PMID:7959882

  7. Combined exposure to nano-silica and lead induced potentiation of oxidative stress and DNA damage in human lung epithelial cells.

    PubMed

    Lu, Chun-Feng; Yuan, Xiao-Yan; Li, Li-Zhong; Zhou, Wei; Zhao, Jun; Wang, Yi-Mei; Peng, Shuang-Qing

    2015-12-01

    Growing evidence has confirmed that exposure to ambient particulate matters (PM) is associated with increased morbidity and mortality of cardiovascular and pulmonary diseases. Ambient PM is a complex mixture of particles and air pollutants. Harmful effects of PM are specifically associated with ultrafine particles (UFPs) that can adsorb high concentrations of toxic air pollutants and are easily inhaled into the lungs. However, combined effects of UFPs and air pollutants on human health remain unclear. In the present study, we elucidated the combined toxicity of silica nanoparticles (nano-SiO2), a typical UFP, and lead acetate (Pb), a typical air pollutant. Lung adenocarcinoma A549 cells were exposed to nano-SiO2 and Pb alone or their combination, and their combined toxicity was investigated by focusing on cellular oxidative stress and DNA damage. Factorial analyses were performed to determine the potential interactions between nano-SiO2 and Pb. Our results showed that exposure of A549 cells to a modest cytotoxic concentration of Pb alone induced oxidative stress, as evidenced by elevated reactive oxygen species generation and lipid peroxidation, and reduced glutathione content and superoxide dismutase and glutathione peroxidase activities. In addition, exposure of A549 cells to Pb alone induced DNA damage, as evaluated by alkaline comet assay. Exposure of A549 cells to non-cytotoxic concentration of nano-SiO2 did not induce cellular oxidative stress and DNA damage. However, exposure to the combination of nano-SiO2 and Pb potentiated oxidative stress and DNA damage in A549 cells. Factorial analyses indicated that the potentiation of combined toxicity of nano-SiO2 and Pb was induced by additive or synergistic interactions. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. Effects of TRPC1 on epithelial mesenchymal transition in human airway in chronic obstructive pulmonary disease

    PubMed Central

    Xu, Feng; Liu, Xiao-Chun; Li, Li; Ma, Chao-Nan; Zhang, Ya-Jun

    2017-01-01

    Abstract Background: We investigated the effects of TRPC1 on epithelial mesenchymal transition (EMT) in human airway in chronic obstructive pulmonary disease (COPD). Methods: A total of 94 patients who underwent lobectomy were selected and divided into COPD (49 cases) and control (45 cases) groups. Immunohistochemistry was applied to detect expression of E-cadherin and vimentin and TRPC1. Correlation of TRPC1 expression with E-cadherin and vimentin expression, and correlations of lung function indicators in COPD patients with expression of TRPC1, E-cadherin, and vimentin were analyzed. Human airway epithelial cells (16HBE) were used for cell experiments; and cigarette smoking extract (CSE) was adopted to establish the COPD model using TRPC1 recombinant plasmids and siRNA. Cells were assigned into the control, CSE, CSE + vector, CSE + TRPC1, CSE + si-NC, and CSE + si-TRPC1 groups. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot were implemented to detect expression of TRPC1, E-cadherin, and vimentin. Results: Compared with the control group, expression of TRPC1 and vimentin significantly increased while expression of E-cadherin decreased in the COPD group, and protein expression of TRPC1 was positively correlated with the protein expression of vimentin but negatively correlated with the protein expression of E-cadherin. Patients exhibiting positive expression of TRPC1 had lower FEV1, FEV1%Pred, and FEV1/FVC, compared with the patients exhibiting negative expression of TRPC1. Compared with the control group, expression of TRPC1 and vimentin increased, whereas expression of E-cadherin decreased in the CSE, CSE + vector, CSE + TRPC1, and CSE + si-NC groups. Compared with the CSE and CSE + vector groups, the expression of TRPC1 and vimentin increased but the expression of E-cadherin decreased in the CSE + TRPC1 group. Compared with the CSE and CSE + si-NC groups, the expression of TRPC1 and vimentin

  9. Involvement of p42/p44 MAPK, JNK, and NF-kappaB in IL-1beta-induced ICAM-1 expression in human pulmonary epithelial cells.

    PubMed

    Lin, Feng-Shu; Lin, Chih-Chung; Chien, Chin-Sung; Luo, Shu-Feng; Yang, Chuen-Mao

    2005-02-01

    Interleukin-1beta (IL-1beta) has been shown to induce the expression of intercellular adhesion molecule-1 (ICAM-1) on airway epithelial cells and contributes to inflammatory responses. However, the mechanisms regulating ICAM-1 expression by IL-1beta in human A549 cells was not completely understood. Here, the roles of mitogen-activated protein kinases (MAPKs) and NF-kappaB pathways for IL-1beta-induced ICAM-1 expression were investigated in A549 cells. IL-1beta induced expression of ICAM-1 protein and mRNA in a time- and concentration-dependent manner. The IL-1beta induction of ICAM-1 mRNA and protein were partially inhibited by U0126 and PD98059 (specific inhibitors of MEK1/2) and SP600125 [a specific inhibitor of c-Jun-N-terminal kinase (JNK)]. U0126 was more potent than other inhibitors to attenuate IL-1beta-induced ICAM-1 expression. Consistently, IL-1beta stimulated phosphorylation of p42/p44 MAPK and JNK which was attenuated by pretreatment with U0126 or SP600125, respectively. Moreover, transfection with dominant negative mutants of MEK1/2 (MEK K97R) or ERK2 (ERK2 K52R) also attenuated IL-1beta-induced ICAM-1 expression. The combination of PD98059 and SP600125 displayed an additive effect on IL-1beta-induced ICAM-1 gene expression. IL-1beta-induced ICAM-1 expression was almost completely blocked by a specific NF-kappaB inhibitor helenalin. Consistently, IL-1beta stimulated translocation of NF-kappaB into the nucleus and degradation of IkappaB-alpha which was blocked by helenalin, U0126, or SP600125. Taken together, these results suggest that activation of p42/p44 MAPK and JNK cascades, at least in part, mediated through NF-kappaB pathway is essential for IL-1beta-induced ICAM-1 gene expression in A549 cells. These results provide new insight into the mechanisms of IL-1beta action that cytokines may promote inflammatory responses in the airway disease. 2004 Wiley-Liss, Inc.

  10. Silencing the Snail-dependent RNA splice regulator ESRP1 drives malignant transformation of human pulmonary epithelial cells. | Office of Cancer Genomics

    Cancer.gov

    Epithelial-to-mesenchymal transition (EMT) is organized in cancer cells by a set of key transcription factors, but the significance of this process is still debated including in non-small cell lung cancer (NSCLC). Here we report increased expression of the EMT-inducing transcription factor Snail in premalignant pulmonary lesions, relative to histologically normal pulmonary epithelium. In immortalized human pulmonary epithelial cells and isogenic derivatives, we documented Snail-dependent anchorage-independent growth in vitro and primary tumor growth and metastatic behavior in vivo.

  11. Grainyhead-like 2 (GRHL2) distribution reveals novel pathophysiological differences between human idiopathic pulmonary fibrosis and mouse models of pulmonary fibrosis

    PubMed Central

    Mahavadi, Poornima; Sasikumar, Satish; Cushing, Leah; Hyland, Tessa; Rosser, Ann E.; Riccardi, Daniela; Lu, Jining; Kalin, Tanya V.; Kalinichenko, Vladimir V.; Guenther, Andreas; Ramirez, Maria I.; Pardo, Annie; Selman, Moisés; Warburton, David

    2013-01-01

    Chronic injury of alveolar lung epithelium leads to epithelial disintegrity in idiopathic pulmonary fibrosis (IPF). We had reported earlier that Grhl2, a transcriptional factor, maintains alveolar epithelial cell integrity by directly regulating components of adherens and tight junctions and thus hypothesized an important role of GRHL2 in pathogenesis of IPF. Comparison of GRHL2 distribution at different stages of human lung development showed its abundance in developing lung epithelium and in adult lung epithelium. However, GRHL2 is detected in normal human lung mesenchyme only at early fetal stage (week 9). Similar mesenchymal reexpression of GRHL2 was also observed in IPF. Immunofluorescence analysis in serial sections from three IPF patients revealed at least two subsets of alveolar epithelial cells (AEC), based on differential GRHL2 expression and the converse fluorescence intensities for epithelial vs. mesenchymal markers. Grhl2 was not detected in mesenchyme in intraperitoneal bleomycin-induced injury as well as in spontaneously occurring fibrosis in double-mutant HPS1 and HPS2 mice, whereas in contrast in a radiation-induced fibrosis model, with forced Forkhead box M1 (Foxm1) expression, an overlap of Grhl2 with a mesenchymal marker was observed in fibrotic regions. Grhl2's role in alveolar epithelial cell plasticity was confirmed by altered Grhl2 gene expression analysis in IPF and further validated by in vitro manipulation of its expression in alveolar epithelial cell lines. Our findings reveal important pathophysiological differences between human IPF and specific mouse models of fibrosis and support a crucial role of GRHL2 in epithelial activation in lung fibrosis and perhaps also in epithelial plasticity. PMID:24375798

  12. Evaluation of pulmonary perfusion by SPECT imaging using an endothelial cell tracer in supine humans and dogs.

    PubMed

    Levac, Xavier; Harel, François; Finnerty, Vincent; Nguyen, Quang T; Letourneau, Myriam; Marcil, Sophie; Fournier, Alain; Dupuis, Jocelyn

    2016-12-01

    Pulmonary perfusion is not spatially homogeneously distributed, and its variations could be of diagnostic value in lung vascular disease. PulmoBind is a ligand of the adrenomedullin receptor densely expressed in endothelial cells of lung capillaries. The aim of this study was to evaluate spatial distribution of human lung perfusion by using this novel molecular tracer of the pulmonary vascular endothelium. Normal humans (n = 19) enrolled into the PulmoBind phase I trial were studied (Clinicaltrials.gov. NCT01539889 ). They were injected with (99m)Tc-PulmoBind for SPECT imaging. Results were compared with (99m)Tc-PulmoBind in quadruped mammals (dogs, n = 5). Imaging was performed in the supine position and distribution of activity was determined as a function of cumulative voxels along the different anatomical planes. PulmoBind uptake in humans was 58 ± 1 % (mean ± SEM) of the injected dose. Dorsal activity was 18.1 ± 2.1 % greater than ventral, and caudal activity was 25.7 ± 1.6 % greater than cranial. Lateral activity was only mildly higher than medial by 7.0 ± 1.0 %. In supine dogs, similar but higher PulmoBind gradients were present: dorsal 28.6 ± 2.5 %, caudal 34.1 ± 5.0 % and lateral 18.1 ± 2.0 %. The perfused pulmonary circulation of supine humans, assessed by an adrenomedullin receptor ligand, is not homogeneously distributed with more prominent distribution in dorsal and caudal regions. It is qualitatively similar to a supine quadruped mammal confirming the presence of a microcirculatory gravitational perfusion gradient detectable with this tracer. Future studies are needed to determine if this novel endothelial cell tracer could be used to detect physiologic and pathologic variations of lung perfusion such as in pulmonary hypertension. ClinicalTrial.gov, NCT01539889.

  13. Electrospinning of PVA/sericin nanofiber and the effect on epithelial-mesenchymal transition of A549 cells.

    PubMed

    Yan, Shanshan; Li, Xiuchun; Dai, Jing; Wang, Yiqun; Wang, Binbin; Lu, Yi; Shi, Jianlin; Huang, Pengyu; Gong, Jinkang; Yao, Yuan

    2017-10-01

    This research aims to investigate the cell-nanomaterial interaction between epithelial-mesenchymal transition of A549 cell and electrospinning nanofibers composed of polyvinyl alcohol (PVA)/silk sericin (SS). The electrospinning of regenerated nanofiber was performed with water as a spinning solvent and glutaraldehyde as a chemical cross-linker. Solution concentration, applied voltage and spin distances as well as other parameters were optimized to generate fine nanofibers with smooth surface in good homogeneity. From the scanning electron microscopy (SEM) analysis, the nanofibers had an average diameter of 200nm. Epithelial-mesenchymal transition (EMT) is a process by which epithelial cells lose their cell polarity to become mesenchymal stem cells. This transition is affected by multiple biochemical and physical factors in cell metabolism cascade. Herein, we investigate the biophysical effect on A549 EMT by culturing cells on nanofibrous mats with different topography and composition. The cell viability was evaluated by biochemical assay and its morphology was observed with SEM. The results demonstrate that cells appropriately attached to the surface of the nanofibrous mats with extended morphology by their filopodia. Gene expression analysis was conducted by real-time PCR using multiple markers for detecting EMT: N-cadherin (NCad), Vimentin (Vim), Fibronectin (Fib) and Matrix metallopeptidase (MMP9). An increasing expression pattern was observed on NCad, Vim, Fib, with respect to a negative control as cell cultured on polystyrene dish. This result indicates the 200nm PVA/SS nanofibers may induce A549 cells to process epithelial-mesenchymal transition during the culturing. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. The reservoir-wave approach to characterize pulmonary vascular-right ventricular interactions in humans.

    PubMed

    Ghimire, Anukul; Andersen, Mads J; Burrowes, Lindsay M; Bouwmeester, J Christopher; Grant, Andrew D; Belenkie, Israel; Fine, Nowell M; Borlaug, Barry A; Tyberg, John V

    2016-12-01

    Using the reservoir-wave approach (RWA) we previously characterized pulmonary vasculature mechanics in a normal canine model. We found reflected backward-traveling waves that decrease pressure and increase flow in the proximal pulmonary artery (PA). These waves decrease right ventricular (RV) afterload and facilitate RV ejection. With pathological alterations to the pulmonary vasculature, these waves may change and impact RV performance. Our objective in this study was to characterize PA wave reflection and the alterations in RV performance in cardiac patients, using the RWA. PA pressure, Doppler-flow velocity, and pulmonary arterial wedge pressure were measured in 11 patients with exertional dyspnea. The RWA was employed to analyze PA pressure and flow; wave intensity analysis characterized PA waves. Wave-related pressure was partitioned into two components: pressures due to forward-traveling and to backward-traveling waves. RV performance was assessed by examining the work done in raising reservoir pressure and that associated with the wave components of systolic PA pressure. Wave-related work, the mostly nonrecoverable energy expended by the RV to eject blood, tended to vary directly with mean PA pressure. Where PA pressures were lower, there were pressure-decreasing/flow-increasing backward waves that aided RV ejection. Where PA pressures were higher, there were pressure-increasing/flow-decreasing backward waves that impeded RV ejection. Pressure-increasing/flow-decreasing backward waves were responsible for systolic notches in the Doppler flow velocity profiles in patients with the highest PA pressure. Pulmonary hypertension is characterized by reflected waves that impede RV ejection and an increase in wave-related work. The RWA may facilitate the development of therapeutic strategies. Copyright © 2016 the American Physiological Society.

  15. Theranostic-PDT with the antibody anti isoform 4 SOD mitocondrial labeled with PpIX in the lung cancer cell line A-549.

    PubMed

    Martínez-García, Claudia; Medina-Flores, Yolanda; de la Rosa-Vázquez, José Manuel; Soriano-Pérez, Eva Elda; Villalobos-Hernández, Juan Ramón; Ramón-Gallegos, Eva

    2018-03-27

    In this work, a drug product composed of an IgM antibody derived from a hybridoma subclone 4C1F6D5G7B8 was prepared and further labeled with PpIX to be used in cell lines A-549 and MRC-5. The aim of this work was to evaluate the potential theranostic activity of the obtained product together with photodynamic therapy (PDT). The IgM antibody labeled with PpIX was used in different concentrations to perform theranostics with PDT in cell lines A-549 and MRC-5 in order to identify the specificity of IgM antibody in lung cancer cells by means of a LED-irradiation system set at 630 nm. The location of the conjugate was further determined by confocal microscopy. The theranostic with conjugate Ab-PpIX in the A-549 cell lines showed fluorescence by confocal microscopy, whereas the MRC-5 cell line showed no reactivity. The PDT with the conjugate in the cell line A-549 decreased its viability 70 % compared to the control. On the contrary, with the MRC-5 cell line no viability diference was shown. The confocal microscopy applied to the cell line A-549 showed that the Ab-PpIX was majorly located at the cytoplasm. Ab-PpIX showed therapeutical potential in lung cancer cells A-549 and had no activity in non-cancerous lung cells (MCR-5). Copyright © 2018. Published by Elsevier B.V.

  16. The Therapeutic Effects of Human Mesenchymal Stem Cells Primed with Sphingosine-1 Phosphate on Pulmonary Artery Hypertension.

    PubMed

    Kang, Hyunsook; Kim, Kang-Hyun; Lim, Jisun; Kim, You-Sun; Heo, Jinbeom; Choi, Jongjin; Jeong, Jaeho; Kim, YongHwan; Kim, Seong Who; Oh, Yeon-Mok; Choo, Myung-Soo; Son, Jaekyoung; Kim, Su Jung; Yoo, Hyun Ju; Oh, Wonil; Choi, Soo Jin; Lee, Sei Won; Shin, Dong-Myung

    2015-07-15

    Stem cell (SC) therapy has become a potential treatment modality for pulmonary artery hypertension (PAH), but the efficacy of human SC and priming effects have not yet been established. The mobilization and homing of hematopoietic stem cells (HSCs) are modulated by priming factors that include a bioactive lipid, sphingosine-1-phosphate (S1P), which stimulates CXCR4 receptor kinase signaling. Here, we show that priming human mesenchymal stem cells (MSCs) with S1P enhances their therapeutic efficacy in PAH. Human MSCs, similar to HSCs, showed stronger chemoattraction to S1P in transwell assays. Concomitantly, MSCs treated with 0.2 μM S1P showed increased phosphorylation of both MAPKp42/44 and AKT protein compared with nonprimed MSCs. Furthermore, S1P-primed MSCs potentiated colony forming unit-fibroblast, anti-inflammatory, and angiogenic activities of MSCs in culture. In a PAH animal model induced by subcutaneously injected monocrotaline, administration of human cord blood-derived MSCs (hCB-MSCs) or S1P-primed cells significantly attenuated the elevated right ventricular systolic pressure. Notably, S1P-primed CB-MSCs, but not unprimed hCB-MSCs, also elicited a significant reduction in the right ventricular weight ratio and pulmonary vascular wall thickness. S1P-primed MSCs enhanced the expression of several genes responsible for stem cell trafficking and angiogenesis, increasing the density of blood vessels in the damaged lungs. Thus, this study demonstrates that human MSCs have potential utility for the treatment of PAH, and that S1P priming increases the effects of SC therapy by enhancing cardiac and vascular remodeling. By optimizing this protocol in future studies, SC therapy might form a basis for clinical trials to treat human PAH.

  17. Cis-acting sequences from a human surfactant protein gene confer pulmonary-specific gene expression in transgenic mice

    SciTech Connect

    Korfhagen, T.R.; Glasser, S.W.; Wert, S.E.

    1990-08-01

    Pulmonary surfactant is produced in late gestation by developing type II epithelial cells lining the alveolar epithelium of the lung. Lack of surfactant at birth is associated with respiratory distress syndrome in premature infants. Surfactant protein C (SP-C) is a highly hydrophobic peptide isolated from pulmonary tissue that enhances the biophysical activity of surfactant phospholipids. Like surfactant phospholipid, SP-C is produced by epithelial cells in the distal respiratory epithelium, and its expression increases during the latter part of gestation. A chimeric gene containing 3.6 kilobases of the promoter and 5{prime}-flanking sequences of the human SP-C gene was used to expressmore » diphtheria toxin A. The SP-C-diphtheria toxin A fusion gene was injected into fertilized mouse eggs to produce transgenic mice. Affected mice developed respiratory failure in the immediate postnatal period. Morphologic analysis of lungs from affected pups showed variable but severe cellular injury confined to pulmonary tissues. Ultrastructural changes consistent with cell death and injury were prominent in the distal respiratory epithelium. Proximal components of the tracheobronchial tree were not severely affected. Transgenic animals were of normal size at birth, and structural abnormalities were not detected in nonpulmonary tissues. Lung-specific diphtheria toxin A expression controlled by the human SP-C gene injured type II epithelial cells and caused extensive necrosis of the distal respiratory epithelium. The absence of type I epithelial cells in the most severely affected transgenic animals supports the concept that developing type II cells serve as precursors to type I epithelial cells.« less

  18. SAHA treatment reveals the link between histone lysine acetylation and proteome in nonsmall cell lung cancer A549 Cells.

    PubMed

    Wu, Quan; Xu, Weiqing; Cao, Lejie; Li, Xin; He, Tieming; Wu, Zhiwei; Li, Wenting

    2013-09-06

    Suberoylanilide hydroxamic acid (SAHA) is a well-known pan HDAC inhibitor, and its clinical application (Vorinostat) has been demonstrated to treat nonsmall cell lung cancer (NSCLS). Nevertheless, the impact of SAHA treatment on histone lysine acetylation and proteome in NSCLS cells still need further elucidate. In NSCLS A549 cells, by using stable isotope labeling for cell culture (SILAC)-based quantitative proteomics, biochemistry assay, and bioinformatic analysis, here we for the first time comprehensively identified and quantified histone lysine acetylation in A549 cells toward SAHA treatment. Despite the fact that SAHA treatment significantly increased histone lysine acetylation in specific sites, unexpectedly, some important "histone markers" showed markedly decreased acetylation level. Further quantitative proteome studies showed that among totally quantifiable 2818 nonredundant proteins, 1355 proteins were with increased level and 1463 with decreased level in response to SAHA treatment. Bioinformatic analysis further revealed that those quantifiable proteins were mainly involved in multiple biological functions and metabolic and enzyme-regulated pathways as well as protein complexes. By establishing the link between histone modification and whole proteome in response to SAHA treatment in NSCLS cells, this study therefore may deepen our understanding of HDAC inhibitor-mediated cancer therapeutics.

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

    PubMed

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

    2017-03-01

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

  20. Telmisartan inhibits NSCLC A549 cell proliferation and migration by regulating the PI3K/AKT signaling pathway.

    PubMed

    Zhang, Suolin; Wang, Yayan

    2018-04-01

    Expression of angiotensin II (Ang II), a key biological peptide in the renin-angiotensin system, is closely associated with the occurrence and development of cancer. Ang II binds two receptor subtypes, the Ang II type 1 receptor (AT1R) and the AT2R, to mediate a series of biological effects. Telmisartan, a specific AT1R blocker, has been reported to have potential as an anticancer drug for treating renal cancer. In the present study, whether telmisartan had an effect on non-small cell lung cancer (NSCLC) cell proliferation and migration was investigated. The Cell Counting kit-8 assay revealed that telmisartan significantly inhibited the growth of the NSCLC A549 cell line in a time- and dose-dependent manner. In a transwell assay, telmisartan significantly inhibited cellular invasion and migration. Furthermore, it was determined that the expression of the anti-apoptotic protein B-cell lymphoma was decreased, and that of the pro-apoptotic proteins caspase-3 and Bcl-associated X increased in the A549 cells treated with telmisartan. Additionally, levels of phosphorylated RAC serine/threonine-protein kinase (p-AKT), p-mechanistic target of rapamycin, p70-S6 kinase and cyclin D1 was decreased in the telmisartan-treated group. Therefore, the current study reveals that telmisartan-induced NSCLC apoptosis may be regulated via the phosphoinositide 3-kinase/AKT signaling pathway, which indicates that it may be a potential novel drug for clinical NSCLC treatment.

  1. MicroRNA-1 targets Slug and endows lung cancer A549 cells with epithelial and anti-tumorigenic properties.

    PubMed

    Tominaga, Eiji; Yuasa, Katsutoshi; Shimazaki, Sho; Hijikata, Takao

    2013-02-01

    MicroRNA-1 (miR-1) has recently been suggested to function as a tumor suppressor. Its functional relevance was assessed by exploring structural and tumorigenic properties of lung cancer A549 cells stably transduced with retrovirus containing pre-miR-1. A549 cells overexpressing miR-1 exhibited a significant morphological change from a mesenchymal to an epithelial phenotype characterized by cell polarization and intercellular junctions. The cells showed increased expression of E-cadherin, which colocalized with cortical actin filaments and vinculin to form typical adherens junction at the apical regions of intercellular borders. Additionally, they exhibited occludin-positive tight junctions at similar apical regions. Moreover, their migratory and invasive activities were inhibited, and their sensitivity to doxorubicin was increased slightly compared to control mock-infected cells. These structural and tumorigenic properties induced by miR-1 were associated with the reduced expression of Slug, which was a transcriptional repressor of E-cadherin or an inducer of epithelial-to-mesenchymal transition. Consistently, Slug was identified as a miR-1 target by bioinformatics and a luciferase reporter assay with plasmids containing luciferase-Slug 3'UTR. Collectively, the data presented here suggest that re-expression of miR-1 may be an effective therapy that prevents cancer malignancy by converting cells from a mesenchymal phenotype to an epithelial phenotype via the downregulation of Slug. Copyright © 2012 Elsevier Inc. All rights reserved.

  2. Human metapneumovirus infection activates the TSLP pathway that drives excessive pulmonary inflammation and viral replication in mice.

    PubMed

    Lay, Margarita K; Céspedes, Pablo F; Palavecino, Christian E; León, Miguel A; Díaz, Rodrigo A; Salazar, Francisco J; Méndez, Gonzalo P; Bueno, Susan M; Kalergis, Alexis M

    2015-06-01

    Human metapneumovirus (hMPV) is a leading cause of acute respiratory tract infections in children and the elderly. The mechanism by which this virus triggers an inflammatory response still remains unknown. Here, we evaluated whether the thymic stromal lymphopoietin (TSLP) pathway contributes to lung inflammation upon hMPV infection. We found that hMPV infection promotes TSLP expression both in human airway epithelial cells and in the mouse lung. hMPV infection induced lung infiltration of OX40L(+) CD11b(+) DCs. Mice lacking the TSLP receptor deficient mice (tslpr(-/-) ) showed reduced lung inflammation and hMPV replication. These mice displayed a decreased number of neutrophils as well a reduction in levels of thymus and activation-regulated chemokine/CCL17, IL-5, IL-13, and TNF-α in the airways upon hMPV infection. Furthermore, a higher frequency of CD4(+) and CD8(+) T cells was found in tslpr(-/-) mice compared to WT mice, which could contribute to controlling viral spread. Depletion of neutrophils in WT and tslpr(-/-) mice decreased inflammation and hMPV replication. Remarkably, blockage of TSLP or OX40L with specific Abs reduced lung inflammation and viral replication following hMPV challenge in mice. Altogether, these results suggest that activation of the TSLP pathway is pivotal in the development of pulmonary pathology and pulmonary hMPV replication. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Pulmonary Thromboembolism: Evaluation By Intravenous Angiography

    NASA Astrophysics Data System (ADS)

    Pond, Gerald D.; Cook, Glenn C.; Woolfenden, James M.; Dodge, Russell R.

    1981-11-01

    Using perfusion lung scans as a guide, digital video subtraction angiography of the pulmonary arteries was performed in human subjects suspected of having pulmonary embolism. Dogs were employed as a pulmonary embolism model and both routine pulmonary angiography and intravenous pulmonary angiograms were obtained for comparison purposes. We have shown by our preliminary results that the technique is extremely promising as a safe and accurate alternative to routine pulmonary angiography in selected patients.

  4. Exercise-induced interstitial pulmonary edema at sea-level in young and old healthy humans

    PubMed Central

    Taylor, Bryan J.; Carlson, Alex R.; Miller, Andrew D.; Johnson, Bruce D.

    2014-01-01

    We asked whether aged adults are more susceptible to exercise-induced pulmonary edema relative to younger individuals. Lung diffusing capacity for carbon monoxide (DLCO), alveolar-capillary membrane conductance (Dm) and pulmonary-capillary blood volume (Vc) were measured before and after exhaustive discontinuous incremental exercise in 10 young (YNG; 27±3 yr) and 10 old (OLD; 69±5 yr) males. In YNG subjects, Dm increased (11±7%, P=0.031), Vc decreased (−10±9%, P=0.01) and DLCO was unchanged (30.5±4.1 vs. 29.7±2.9 ml/min/mmHg, P=0.44) pre- to post-exercise. In OLD subjects, DLCO and Dm increased (11±14%, P=0.042; 16±14%, P=0.025) but Vc was unchanged (58±23 vs. 56±23 ml, P=0.570) pre- to post-exercise. Group-mean Dm/Vc was greater after vs. before exercise in the YNG and OLD subjects. However, Dm/Vc was lower post-exercise in 2 of the 10 YNG (−7±4%) and 2 of the 10 OLD subjects (−10±5%). These data suggest that exercise decreases interstitial lung fluid in most YNG and OLD subjects, with a small number exhibiting evidence for exercise-induced pulmonary edema. PMID:24200644

  5. Pulmonary tuberculosis

    MedlinePlus

    TB; Tuberculosis - pulmonary; Mycobacterium - pulmonary ... Pulmonary TB is caused by the bacterium Mycobacterium tuberculosis (M tuberculosis) . TB is contagious. This means the bacteria is easily spread from an infected person to someone else. You can get ...

  6. Molecular Mode of Action and Role of TP53 in the Sensitivity to the Novel Epothilone Sagopilone (ZK-EPO) in A549 Non-Small Cell Lung Cancer Cells

    PubMed Central

    Winsel, Sebastian; Sommer, Anette; Eschenbrenner, Julia; Mittelstaedt, Kevin; Klar, Ulrich; Hammer, Stefanie; Hoffmann, Jens

    2011-01-01

    Sagopilone, an optimized fully synthetic epothilone, is a microtubule-stabilizing compound that has shown high in vitro and in vivo activity against a broad range of human tumor models. We analyzed the differential mechanism of action of sagopilone in non-small cell lung cancer cell lines in vitro. Sagopilone inhibited proliferation of non-small cell lung cancer cell lines at lower nanomolar concentration. The treatment with sagopilone caused strong disturbances of cellular cytoskeletal organization. Two concentration-dependent phenotypes were observed. At 2.5 nM sagopilone or 4 nM paclitaxel an aneuploid phenotype occur whereas a mitotic arrest phenotype was induced by 40 nM sagopilone or paclitaxel. Interestingly, treatment with 2.5 nM of sagopilone effectively inhibited cell proliferation, but - compared to high concentrations (40 nM) - only marginally induced apoptosis. Treatment with a high versus a low concentration of sagopilone or paclitaxel regulates a non-overlapping set of genes, indicating that both phenotypes substantially differ from each other. Genes involved in G2/M phase transition and the spindle assembly checkpoint, like Cyclin B1 and BUBR1 were upregulated by treatment with 40 nM sagopilone. Unexpectedly, also genes involved in DNA damage response were upregulated under that treatment. In contrast, treatment of A549 cells with a low concentration of sagopilone revealed an upregulation of direct transcriptional target genes of TP53, like CDKN1A, MDM2, GADD45A, FAS. Knockdown of TP53, which inhibited the transcriptional induction of TP53 target genes, led to a significant increase in apoptosis induction in A549 cells when treated with a low concentration of sagopilone. The results indicate that activation of TP53 and its downstream effectors like CDKN1A by low concentrations of sagopilone is responsible for the relative apoptosis resistance of A549 cells and might represent a mechanism of resistance to sagopilone. PMID:21559393

  7. D-4F, an apolipoprotein A-I mimetic, inhibits TGF-β1 induced epithelial-mesenchymal transition in human alveolar epithelial cell.

    PubMed

    You, Jia; Wang, Jintao; Xie, Linshen; Zhu, Chengwen; Xiong, Jingyuan

    2016-10-01

    Emerging evidences support that transforming growth factor β1 (TGF-β1) induced epithelial-mesenchymal transition (EMT) participates in the pathogenesis of pulmonary fibrosis and asthmatic airway remodeling. Recent studies demonstrated that apolipoprotein A-I (Apo A-I) is the only known substance that can resolve established pulmonary fibrotic nodules, and Apo A-I mimetic D-4F (a synthetic polypeptide consisting of 18 amino acids) plays an inhibitory role in murine asthmatic model. However, cellular mechanisms for such therapeutic effects of Apo A-I and D-4F remain to be elucidated. This study evaluated the effects of D-4F on TGF-β1 induced EMT in human type II alveolar epithelial cell line A549. A549 cells treated with 10ng/ml of TGF-β1 manifested distinct EMT, including fibroblastic morphological changes, down-regulation of epithelial marker E-cadherin and up-regulation of mesenchymal marker vimentin. These EMT related changes were all inhibited by D-4F in a concentration dependent manner. Transcriptional investigation demonstrated clearly that D-4F dose-dependently compensated for the reduced E-cadherin mRNA level and the increased vimentin mRNA level in TGF-β1 treated A549 cells. Translational analysis revealed that D-4F significantly reversed the TGF-β1 induced changes of E-cadherin and vimentin levels. These results suggested that D-4F inhibits TGF-β1 induced EMT in human alveolar epithelial cell. Given the functional similarities between D-4F and Apo A-I, it is speculated that D-4F and Apo A-I are able to exert possible anti-fibrotic and anti-asthmatic effects via inhibiting alveolar EMT, and D-4F may possess beneficial clinical potential for patients suffering from pulmonary fibrosis and asthma. Copyright © 2016 Elsevier GmbH. All rights reserved.

  8. In vitro chemoresistance profile and expression/function of MDR associated proteins in resistant cell lines derived from CCRF-CEM, K562, A549 and MDA MB 231 parental cells.

    PubMed

    Noskova, V; Dzubak, P; Kuzmina, G; Ludkova, A; Stehlik, D; Trojanec, R; Janostakova, A; Korinkova, G; Mihal, V; Hajduch, M

    2002-01-01

    Although cellular experiments have elucidated a number of active principles in the study of the multidrug resistance (MDR) phenomena, most of the drug resistant tumor cells were derived from different parental cell lines. This fact limits generalization of some experimental data and conclusions, and therefore we selected and characterized cell lines resistant to various anti-cancer agents derived from four parental cell lines: CEM (human T-lymphoblastic leukemia), K562 (human myeloid leukemia), A549 (human lung adenocarcinoma) and MDAMB 231 (human breast adenocarcinoma). In total we obtained a set of 42 resistant sublines, which is an excellent tool for the future studies of different aspects of MDR. In this study we report on some basic characteristics of these sublines, namely, cross-resistance to other anti-cancer drugs investigated by in vitro MTT assay, expression of MDR associated proteins (Pgp, MRP1, LRP, GST-pi and Topo IIalpha) as well as the functional activity of Pgp and MRP.

  9. Activation of p53/miR-34a Tumor Suppressor Axis by Chinese Herbal Formula JP-1 in A549 Lung Adenocarcinoma Cells

    PubMed Central

    Chow, Jyh-Ming; Lin, Pei-Chun; Hu, Tsai-Shu; Kuo, Hui-Ching; Huang, Jhy-Shrian

    2016-01-01

    Lung cancer is the leading cause of cancer death worldwide; the most common pathologic type is lung adenocarcinoma (LADC). In spite of the recent progress in targeted therapy, most LADC patients eventually expired due to the inevitable recurrence and drug resistance. New complementary agent with evidence-based molecular mechanism is urgently needed. MiR-34a is an important p53 downstream tumor suppressor, which regulates apoptosis, cell-cycle, EMT (epithelial mesenchymal transition), and so forth. Its expression is deficient in many types of cancers including LADC. Here, we show that a Chinese herbal formula JP-1 activates p53/miR-34a axis in A549 human LADC cells (p53 wild-type). Treatment with JP-1 induces p53 and its downstream p21 and BAX proteins as well as the miR-34a, resulting in growth inhibition, colony formation reduction, migration repression, and apoptosis induction. Accordingly, the decreases of miR-34a downstream targets such as CDK6, SIRT1, c-Myc, survivin, Snail, and AXL were observed. Moreover, JP-1 activates AMPKα and reduces mTOR activity, implying its inhibitory effect on the energy-sensitive protein synthesis and cell proliferation signaling. Our results show that JP-1 activates p53/miR-34a tumor suppressor axis and decreases proteins related to proliferation, apoptosis resistance, and metastasis, suggesting its potential as a complementary medicine for LADC treatment. PMID:28074102

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

    PubMed

    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. Copyright © 2012 Elsevier B.V. All rights reserved.

  11. A549 and PLC/PRF/5 cells can support the efficient propagation of swine and wild boar hepatitis E virus (HEV) strains: demonstration of HEV infectivity of porcine liver sold as food.

    PubMed

    Takahashi, Hideyuki; Tanaka, Toshinori; Jirintai, Suljid; Nagashima, Shigeo; Takahashi, Masaharu; Nishizawa, Tsutomu; Mizuo, Hitoshi; Yazaki, Yasuyuki; Okamoto, Hiroaki

    2012-02-01

    Recent evidence has indicated the cross-species transmission of hepatitis E virus (HEV) from pigs and wild boars to humans, causing zoonosis, mostly via consumption of uncooked or undercooked animal meat/viscera. However, no efficient cell culture system for swine and boar HEV strains has been established. We inoculated A549 cells with 12 swine and boar HEV strains of liver, feces, or serum origin at an HEV load of ≥2.0 × 10(4) copies per well and found that the HEV progeny replicated as efficiently as human HEV strains, with a maximum load of ~10(8) copies/ml. However, the HEV load in the culture medium at 30 days post-inoculation differed markedly by inoculum, ranging from 1.0 × 10(2) to 1.1 × 10(7) copies/ml upon inoculation at a lower load of approximately 10(5) copies per well. All progeny were passaged successfully onto A549 and PLC/PRF/5 cells. In sharp contrast, no progeny viruses were detectable in the culture supernatant upon inoculation with 13 swine and boar HEV strains at an HEV load of <1.8 × 10(4) copies per well. The present study also demonstrates that swine liver sold as food can be infectious, supporting the risk of zoonotic food-borne HEV infection.

  12. Natural antioxidant dihydroxybenzyl alcohol blocks ritonavir-induced endothelial dysfunction in porcine pulmonary arteries and human endothelial cells.

    PubMed

    Weakley, Sarah M; Jiang, Jun; Lü, Jianming; Wang, Xinwen; Lin, Peter H; Yao, Qizhi; Chen, Changyi

    2011-09-01

    Patients with HIV have an increased incidence of pulmonary artery hypertension. This study was designed to determine if the naturally occurring antioxidant dihydroxybenzyl alcohol (DHBA) could counteract the deleterious effects of ritonavir (RTV), an HIV-protease inhibitor known to impair endothelial function and increase oxidative stress. Antioxidant assays were performed on DHBA in a cell free system. Glutathione (GSH) levels were measured in human pulmonary artery endothelial cells (HPAEC) to determine the effect of DHBA on the level of oxidative stress in cells treated with RTV. Myograph analysis was performed on porcine pulmonary artery (PA) rings after treatment with RTV and/or DHBA. Likewise, reactive oxygen species (ROS) production was assessed in porcine PA rings after RTV +/- DHBA using a lucigenin reaction. Immunohistochemical staining for endothelial nitric oxide synthase (eNOS) was also performed in porcine PAs treated as above. DHBA demonstrated significant antioxidant activity in a cell free system that surpassed that of vitamin C. Also, treatment with DHBA reduced RTV-induced reduction in endothelium-dependent vasorelaxation and eNOS staining and increased superoxide anion levels. Meanwhile, there was a reversal in RTV-induced oxidative stress leading to reduced GSH levels in HPAECs after treatment with DHBA. These findings suggest that the naturally occurring antioxidant DHBA reduces the impairment of vasomotor functions caused by RTV in porcine PAs and reduces oxidative stress caused by RTV in HPAEC and porcine PA rings. This study indicates that DHBA may have clinical applications in the prevention or treatment of antiretroviral drugs-associated vascular complications in patients with HIV.

  13. Comparative characterization of pulmonary surfactant aggregates and alkaline phosphatase isozymes in human lung carcinoma tissue.

    PubMed

    Iino, Nozomi; Matsunaga, Toshiyuki; Harada, Tsuyoshi; Igarashi, Seiji; Koyama, Iwao; Komoda, Tsugikazu

    2007-05-01

    Alkaline phosphatase (AP) isozymes are surfactant-associated proteins (SPs). Since several different AP isozymes have been detected in the pneumocytes of lung cancer patients, we attempted to identify the relationship between pulmonary surfactant aggregate subtypes and AP isozymes. Pulmonary surfactant aggregates were isolated from carcinoma and non-carcinoma tissues of patients with non-small cell carcinoma of the lung. Upon analysis, ultraheavy, heavy, and light surfactant aggregates were detected in the non-carcinoma tissues, but no ultraheavy surfactant aggregates were found in the carcinoma tissues. Surfactant-associated protein A (SP-A) was detected as two bands (a 27-kDa band and a 54-kDa band) in the ultraheavy, heavy, and light surfactant aggregates found in the non-carcinoma tissues. Although both SP-A bands were detected in the heavy and light surfactant aggregates from adenocarcinoma tissues, the 54-kDa band was not detected in squamous cell carcinoma tissues. Liver AP (LAP) was detected in the heavy and light surfactant aggregates from both non-carcinoma and squamous carcinoma tissues, but not in heavy surfactant aggregates from adenocarcinoma tissues. A larger amount of bone type AP (BAP) was found in light surfactant aggregate fractions from squamous cell carcinomas than those from adenocarcinoma tissues or non-carcinoma tissues from patients with either type of cancer. LAP, BAP, and SP-A were identified immunohistochemically in type II pneumocytes from non-carcinoma tissues and adenocarcinoma cells, but no distinct SP-A staining was observed in squamous cell carcinoma tissues. The present study has thus revealed several differences in pulmonary surfactant aggregates and AP isozymes between adenocarcinoma tissue and squamous cell carcinoma tissue.

  14. Pulmonary hemodynamics responses to hypoxia and/or CO2 inhalation during moderate exercise in humans.

    PubMed

    Doutreleau, Stéphane; Enache, Irina; Pistea, Cristina; Geny, Bernard; Charloux, Anne

    2018-03-03

    In this study, we hypothesized that adding CO 2 to an inhaled hypoxic gas mixture will limit the rise of pulmonary artery pressure (PAP) induced by a moderate exercise. Eight 20-year-old males performed four constant-load exercise tests on cycle at 40% of maximal oxygen consumption in four conditions: ambient air, normobaric hypoxia (12.5% O 2 ), inhaled CO 2 (4.5% CO 2 ), and combination of hypoxia and inhaled CO 2 . Doppler echocardiography was used to measure systolic (s)PAP, cardiac output (CO). Total pulmonary resistance (TPR) was calculated. Arterialized blood pH was 7.40 at exercise in ambient and hypoxia conditions, whereas CO 2 inhalation and combined conditions showed acidosis. sPAP increases from rest in ambient air to exercise ranged as follows: ambient + 110%, CO 2 inhalation + 135%, combined + 184%, hypoxia + 217% (p < 0.001). CO was higher when inhaling O 2 -poor gas mixtures with or without CO 2 (~ 17 L min -1 ) than in the other conditions (~ 14 L min -1 , p < 0.001). Exercise induced a significant decrease in TPR in the four conditions (p < 0.05) but less marked in hypoxia (- 19% of the resting value in ambient air) than in ambient (- 33%) and in both CO 2 inhalation and combined condition (- 29%). We conclude that (1) acute CO 2 inhalation did not significantly modify pulmonary hemodynamics during moderate exercise. (2) CO 2 adjunction to hypoxic gas mixture did not modify CO, despite a higher CaO 2 in combined condition than in hypoxia. (3) TPR was lower in combined than in hypoxia condition, limiting sPAP increase in combined condition.

  15. The Evaluation of a Pulmonary Display to Detect Adverse Respiratory Events Using High Resolution Human Simulator

    PubMed Central

    Wachter, S. Blake; Johnson, Ken; Albert, Robert; Syroid, Noah; Drews, Frank; Westenskow, Dwayne

    2006-01-01

    Objective Authors developed a picture-graphics display for pulmonary function to present typical respiratory data used in perioperative and intensive care environments. The display utilizes color, shape and emergent alerting to highlight abnormal pulmonary physiology. The display serves as an adjunct to traditional operating room displays and monitors. Design To evaluate the prototype, nineteen clinician volunteers each managed four adverse respiratory events and one normal event using a high-resolution patient simulator which included the new displays (intervention subjects) and traditional displays (control subjects). Between-group comparisons included (i) time to diagnosis and treatment for each adverse respiratory event; (ii) the number of unnecessary treatments during the normal scenario; and (iii) self-reported workload estimates while managing study events. Measurements Two expert anesthesiologists reviewed video-taped transcriptions of the volunteers to determine time to treat and time to diagnosis. Time values were then compared between groups using a Mann-Whitney-U Test. Estimated workload for both groups was assessed using the NASA-TLX and compared between groups using an ANOVA. P-values < 0.05 were considered significant. Results Clinician volunteers detected and treated obstructed endotracheal tubes and intrinsic PEEP problems faster with graphical rather than conventional displays (p < 0.05). During the normal scenario simulation, 3 clinicians using the graphical display, and 5 clinicians using the conventional display gave unnecessary treatments. Clinician-volunteers reported significantly lower subjective workloads using the graphical display for the obstructed endotracheal tube scenario (p < 0.001) and the intrinsic PEEP scenario (p < 0.03). Conclusion Authors conclude that the graphical pulmonary display may serve as a useful adjunct to traditional displays in identifying adverse respiratory events. PMID:16929038

  16. [Pulmonary hypertension in patients infected with human immunodeficiency virus: current situation].

    PubMed

    Soto-Abánades, Clara Itzíar; Alcolea-Batres, Sergio; Ríos-Blanco, Juan José

    2013-01-01

    The increase in survival that has been achieved with the new treatments in the era of highly active antiretroviral therapy, has enabled clinicians and researchers to analyze issues that emerge in the long term in patients with HIV infection. Although the majority of cardiovascular complications have been widely described, the pathogenesis of pulmonary arterial hypertension is still poorly understood, and is one of the more complex and feared complications as it worsens the prognosis and quality of life of these patients This article reviews newer aspects related to the aetiology, symptoms, diagnosis and treatment of this disease. Copyright © 2011 Elsevier España, S.L. All rights reserved.

  17. Effect of pulmonary surfactant on the dissolution, stability and uptake of zinc oxide nanowires by human respiratory epithelial cells

    PubMed Central

    Theodorou, Ioannis G.; Ruenraroengsak, Pakatip; Gow, Andrew; Schwander, Stephan; Zhang, Junfeng (Jim); Chung, Kian Fan; Tetley, Teresa D.; Ryan, Mary P.; Porter, Alexandra E.

    2017-01-01

    Inhaled nanoparticles have high deposition rates in the alveolar region of the lung but the effects of pulmonary surfactant (PS) on nanoparticle bioreactivity are unclear. Here, the impact of PS on the stability and dissolution of ZnO nanowires (ZnONWs) was investigated, and linked with their bioreactivity in vitro with human alveolar epithelial type 1-like cells (TT1). Pre-incubation of ZnONWs with Curosurf® (a natural porcine PS) decreased their dissolution at acidic pH, through the formation of a phospholipid corona. Confocal live cell microscopy confirmed that Curosurf® lowered intracellular dissolution, thus delaying the onset of cell death compared to bare ZnONWs. Despite reducing dissolution, Curosurf® significantly increased the uptake of ZnONWs within TT1 cells, ultimately increasing their toxicity after 24h. Although serum, improved ZnONW dispersion in suspension similar to Curosurf®, it had no effect on ZnONW internalization and toxicity, indicating a unique role of PS in promoting particle uptake. In the absence of PS, ZnONW length had no effect on dissolution kinetics or degree of cellular toxicity, indicating a less important role of length in determining ZnONW bioreactivity. This work provides unique findings on the effects of PS on the stability and toxicity of ZnONWs, which could be important in the study of pulmonary toxicity and epithelial-endothelial translocation of nanoparticles in general. PMID:27441789

  18. Upregulation of Human Endogenous Retrovirus-K Is Linked to Immunity and Inflammation in Pulmonary Arterial Hypertension.

    PubMed

    Saito, Toshie; Miyagawa, Kazuya; Chen, Shih-Yu; Tamosiuniene, Rasa; Wang, Lingli; Sharpe, Orr; Samayoa, Erik; Harada, Daisuke; Moonen, Jan-Renier A J; Cao, Aiqin; Chen, Pin-I; Hennigs, Jan K; Gu, Mingxia; Li, Caiyun G; Leib, Ryan D; Li, Dan; Adams, Christopher M; Del Rosario, Patricia A; Bill, Matthew; Haddad, Francois; Montoya, Jose G; Robinson, William H; Fantl, Wendy J; Nolan, Garry P; Zamanian, Roham T; Nicolls, Mark R; Chiu, Charles Y; Ariza, Maria E; Rabinovitch, Marlene

    2017-11-14

    Immune dysregulation has been linked to occlusive vascular remodeling in pulmonary arterial hypertension (PAH) that is hereditary, idiopathic, or associated with other conditions. Circulating autoantibodies, lung perivascular lymphoid tissue, and elevated cytokines have been related to PAH pathogenesis but without a clear understanding of how these abnormalities are initiated, perpetuated, and connected in the progression of disease. We therefore set out to identify specific target antigens in PAH lung immune complexes as a starting point toward resolving these issues to better inform future application of immunomodulatory therapies. Lung immune complexes were isolated and PAH target antigens were identified by liquid chromatography tandem mass spectrometry, confirmed by enzyme-linked immunosorbent assay, and localized by confocal microscopy. One PAH antigen linked to immunity and inflammation was pursued and a link to PAH pathophysiology was investigated by next-generation sequencing, functional studies in cultured monocytes and endothelial cells, and hemodynamic and lung studies in a rat. SAM domain and HD domain-containing protein 1 (SAMHD1), an innate immune factor that suppresses HIV replication, was identified and confirmed as highly expressed in immune complexes from 16 hereditary and idiopathic PAH versus 12 control lungs. Elevated SAMHD1 was localized to endothelial cells, perivascular dendritic cells, and macrophages, and SAMHD1 antibodies were prevalent in tertiary lymphoid tissue. An unbiased screen using metagenomic sequencing related SAMHD1 to increased expression of human endogenous retrovirus K (HERV-K) in PAH versus control lungs (n=4). HERV-K envelope and deoxyuridine triphosphate nucleotidohydrolase mRNAs were elevated in PAH versus control lungs (n=10), and proteins were localized to macrophages. HERV-K deoxyuridine triphosphate nucleotidohydrolase induced SAMHD1 and proinflammatory cytokines (eg, interleukin 6, interleukin 1β, and tumor

  19. Assessing the utility of autofluorescence-based pulmonary optical endomicroscopy to predict the malignant potential of solitary pulmonary nodules in humans

    NASA Astrophysics Data System (ADS)

    Seth, Sohan; Akram, Ahsan R.; McCool, Paul; Westerfeld, Jody; Wilson, David; McLaughlin, Stephen; Dhaliwal, Kevin; Williams, Christopher K. I.

    2016-08-01

    Solitary pulmonary nodules are common, often incidental findings on chest CT scans. The investigation of pulmonary nodules is time-consuming and often leads to protracted follow-up with ongoing radiological surveillance, however, clinical calculators that assess the risk of the nodule being malignant exist to help in the stratification of patients. Furthermore recent advances in interventional pulmonology include the ability to both navigate to nodules and also to perform autofluorescence endomicroscopy. In this study we assessed the efficacy of incorporating additional information from label-free fibre-based optical endomicrosopy of the nodule on assessing risk of malignancy. Using image analysis and machine learning approaches, we find that this information does not yield any gain in predictive performance in a cohort of patients. Further advances with pulmonary endomicroscopy will require the addition of molecular tracers to improve information from this procedure.

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

    PubMed

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

    2018-03-01

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

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

    SciTech Connect

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

    2013-09-27

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

  2. Effect of Avastin on the number and structure of tumor blood vessels of nude mice with A549 lung adenocarcinoma.

    PubMed

    Zhang, Nali; Zhang, Guojun; Zheng, Youguang; Wang, Tongbing; Wang, Honglei

    2014-12-01

    The aim of the present study was to investigate the effect of Avastin on the number and structure of tumor blood vessels of nude mice with A549 lung adenocarcinoma. A total of 30 nude mice were randomly divided into three groups, namely the control, the Avastin I (Avastin 3 mg/kg) and the Avastin II (Avastin 6 mg/kg) groups. Following treatment, ELISA was used to detect the expression level of vascular endothelial growth factor (VEGF) in tumor tissues. The microvascular density in tumor tissues and tumor vascular pericyte coverage was detected by immunofluorescence. The tumor growth and survival rate of mice in the three groups were also analyzed. Compared with the control group, the Avastin I and II groups exhibited significantly decreased VEGF levels and microvascular density in the tumor tissues, with the decrease in the Avastin II group being more prominent (P<0.05). After 7 days of treatment, the vascular pericyte coverage in the tumor tissues of mice in the Avastin I and II groups was significantly increased compared with that in the control group mice (P<0.05). Compared with the control group, the mice in the Avastin I and II groups exhibited a significantly decreased tumor growth rate and this effect was dose-dependent. The survival rate of mice in the Avastin I and II groups was significantly increased compared with that of the mice in the control group (P<0.05). In conclusion, Avastin significantly decreased the microvascular density of the tumor in nude mice with A549 lung adenocarcinoma and also significantly increased tumor vascular pericyte coverage, inhibited tumor growth and increased the survival rate of the mice, through its potent antiangiogenic activity.

  3. Glutamine drives glutathione synthesis and contributes to radiation sensitivity of A549 and H460 lung cancer cell lines

    PubMed Central

    Sappington, Daniel R.; Siegel, Eric R.; Hiatt, Gloria; Desai, Abhishek; Penney, Rosalind B.; Jamshidi-Parsian, Azemat; Griffin, Robert J.; Boysen, Gunnar

    2016-01-01

    Background Increased glutamine uptake is known to drive cancer cell proliferation, making tumor cells glutamine-dependent. Glutamine provides additional carbon and nitrogen sources for cell growth. The first step in glutamine utilization is its conversion to glutamate by glutaminase (GLS). Glutamate is a precursor for glutathione synthesis, and we investigated the hypothesis that glutamine drives glutathione synthesis and thereby contributes to cellular defense systems. Methods The importance of glutamine for glutathione synthesis was studied in H460 and A549 lung cancer cell lines using glutamine-free medium and Bis-2-(5-phenyl-acetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide (BPTES) a GLS inhibitor. Metabolic activities were determined by targeted mass spectrometry. Results A significant correlation between glutamine consumption and glutathione excretion was demonstrated in H460 and A549 tumor cells. Culturing in the presence of [13C5]glutamine demonstrated that by 12 hrs >50% of excreted glutathione is derived from glutamine. Culturing in glutamine-free medium or treatment with BPTES, a glutaminase (GLS)-specific inhibitor, reduced cell proliferation and viability, and abolished glutathione excretion. Treatment with glutathione-ester prevented BPTES induced cytotoxicity. Inhibition of GLS markedly radiosensitized the lung tumor cell lines, suggesting an important role of glutamine-derived glutathione in determining radiation sensitivity. Conclusions We demonstrate here for the first time that a significant amount of extracellular glutathione is directly derived from glutamine. This finding adds yet another important function to the already known glutamine dependence of tumor cells and probably tumors as well. General significance Glutamine is essential for synthesis and excretion of glutathione to promote cell growth and viability. PMID:26825773

  4. Repeated Aurora-A siRNA Transfection Results in Effective Apoptosis of A549 Cells Compared to Single Transfection.

    PubMed

    Wang, Zhonghua; Sun, Wenwu; Cao, Jianping; Cui, Haiyang; Ma, Zhuang

    2016-01-01

    Suppression of Aurora kinase A (Aurora-A, AURKA) by Aurora-A siRNA has been proposed for lung tumor treatment. However, protocols using single administration have shown little benefit in some types of lung tumor. Given that transfection efficiency of Aurora-A siRNA is low due to tightly packed cells in the tumor, we hypothesized that repeated administration would result in efficient cell apoptosis. We compared single vs. repeated transfection (thrice) in A549 cells by transfecting Aurora-A siRNA (siA) on the 1st or 1st, 2nd and 3rd day after cell seeding. A random sequence was used as the negative siRNA control (siC). Cells in the single transfection group received only transfection reagent without siRNAs on the 2nd and 3rd day. Two days after the third transfection, both single and repeated siA administration decreased mRNA expression of Aurora-A and cell viability compared to no administration and siC single administration. However, the decrease in these two indices with repeated transfection was more obvious than that following single administration: cell viability decreased to 72.8 ± 3.05% (p < 0.05) following siA single transfection and to 64.2 ± 1.99% (p < 0.05) following siA repeated transfection, compared with normal control cells, respectively. Gene expression decreased to 17 ± 16.6% (p < 0.05 vs. normal control) following siA repeated transfection and to 43.2 ± 13.0% (p < 0.05 vs. normal control) following siA single transfection. Compared to single transfection, repeated Aurora-A siRNA transfection decreased Aurora-A, which, in turn, resulted in effective apoptosis of A549 cells.

  5. Diesel exhaust particles increase IL-1β-induced human β-defensin expression via NF-κB-mediated pathway in human lung epithelial cells

    PubMed Central

    Nam, Hae Yun; Ahn, Eun-Kyung; Kim, Hyung Jung; Lim, Young; Lee, Chun Beoun; Lee, Kyo Young; Vallyathan, Val

    2006-01-01

    Background Human β-defensin (hBD)-2, antimicrobial peptide primarily induced in epithelial cells, is a key factor in the innate immune response of the respiratory tract. Several studies showed increased defensin levels in both inflammatory lung diseases, such as cystic fibrosis, diffuse panbronchiolitis, idiopathic pulmonary fibrosis and acute respiratory distress syndrome, and infectious diseases. Recently, epidemiologic studies have demonstrated acute and serious adverse effects of particulate air pollution on respiratory health, especially in people with pre-existing inflammatory lung disease. To elucidate the effect of diesel exhaust particles (DEP) on pulmonary innate immune response, we investigated the hBD-2 and interleukin-8 (IL-8) expression to DEP exposure in interleukin-1 beta (IL-1β)-stimulated A549 cells. Results IL-1β markedly up-regulated the hBD-2 promoter activity, and the subsequent DEP exposure increased dose-dependently the expression of hBD-2 and inflammatory cytokine IL-8 at the transcriptional level. In addition, DEP further induced the NF-κB activation in IL-1β-stimulated A549 cells more rapidly than in unstimulated control cells, which was showed by nuclear translocation of p65 NF-κB and degradation of IκB-α. The experiment using two NF-κB inhibitors, PDTC and MG132, confirmed that this increase of hBD-2 expression following DEP exposure was regulated through NF-κB-mediated pathway. Conclusion These results demonstrated that DEP exposure increases the expression of antimicrobial peptide and inflammatory cytokine at the transcriptional level in IL-1β-primed A549 epithelial cells and suggested that the increase is mediated at least partially through NF-κB activation. Therefore, DEP exposure may contribute to enhance the airway-responsiveness especially on the patients suffering from chronic respiratory disease. PMID:16723032

  6. VA/Q distribution during heavy exercise and recovery in humans: implications for pulmonary edema

    NASA Technical Reports Server (NTRS)

    Schaffartzik, W.; Poole, D. C.; Derion, T.; Tsukimoto, K.; Hogan, M. C.; Arcos, J. P.; Bebout, D. E.; Wagner, P. D.

    1992-01-01

    Ventilation-perfusion (VA/Q) inequality has been shown to increase with exercise. Potential mechanisms for this increase include nonuniform pulmonary vasoconstriction, ventilatory time constant inequality, reduced large airway gas mixing, and development of interstitial pulmonary edema. We hypothesized that persistence of VA/Q mismatch after ventilation and cardiac output subside during recovery would be consistent with edema; however, rapid resolution would suggest mechanisms related to changes in ventilation and blood flow per se. Thirteen healthy males performed near-maximal cycle ergometry at an inspiratory PO2 of 91 Torr (because hypoxia accentuates VA/Q mismatch on exercise). Cardiorespiratory variables and inert gas elimination patterns were measured at rest, during exercise, and between 2 and 30 min of recovery. Two profiles of VA/Q distribution behavior emerged during heavy exercise: in group 1 an increase in VA/Q mismatch (log SDQ of 0.35 +/- 0.02 at rest and 0.44 +/- 0.02 at exercise; P less than 0.05, n = 7) and in group 2 no change in VA/Q mismatch (n = 6). There were no differences in anthropometric data, work rate, O2 uptake, or ventilation during heavy exercise between groups. Group 1 demonstrated significantly greater VA/Q inequality, lower vital capacity, and higher forced expiratory flow at 25-75% of forced vital capacity for the first 20 min during recovery than group 2. Cardiac index was higher in group 1 both during heavy exercise and 4 and 6 min postexercise. However, both ventilation and cardiac output returned toward baseline values more rapidly than did VA/Q relationships. Arterial pH was lower in group 1 during exercise and recovery. We conclude that greater VA/Q inequality in group 1 and its persistence during recovery are consistent with the hypothesis that edema occurs and contributes to the increase in VA/Q inequality during exercise. This is supported by observation of greater blood flows and acidosis and, presumably therefore

  7. 4-methoxychalcone enhances cisplatin-induced oxidative stress and cytotoxicity by inhibiting the Nrf2/ARE-mediated defense mechanism in A549 lung cancer cells.

    PubMed

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

    2013-10-01

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

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

    PubMed Central

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

    2013-01-01

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

  9. Targeted interfering DEP domain containing 1 protein induces apoptosis in A549 lung adenocarcinoma cells through the NF-κB signaling pathway.

    PubMed

    Wang, Qingqing; Li, Aili; Jin, Junfei; Huang, Guojin

    2017-01-01

    Ectopic expression of DEP domain containing 1 (DEPDC1) in lung adenocarcinomas is associated with poor prognosis, but its role and the underlying mechanism remain unknown. In this study, DEPDC1 expression in lung cancer cell lines was examined with Western blot assay, and DEPDC1-positive cell A549 was selected for further experiments. DEPDC1 inhibitor miR-130a was overexpressed in A549 cells, and the proliferation and apoptosis of these cells were analyzed with cell counting and flow cytometry assay. Interfering peptide 11R-DEP:611-628 and JNK inhibitor SP600125 were used alone or in combination to treat A549 cells, and the cell proliferation and apoptosis were assessed by flow cytometry assay; caspase 3 and cleaved caspase 3, phosphor-JNK, and total JNK were detected by Western blotting; and nuclear factor kappa B (NF-κB) localization was determined by immunofluorescence staining. We found that miR-130a and 11R-DEP:611-628 peptides (5 μM) both inhibited A549 proliferation and induced apoptosis. We observed that 11R-DEP:611-628 peptide treatment resulted in elevated A20 expression, dramatically reduced nuclear NF-κB, and increased phosphor-JNK. These findings indicate that DEPDC1 inhibits apoptosis of A549 cell by suppressing A20 expression to regulate NF-κB activity, and that JNK plays a protective role upon 11R-DEP:611-628 peptide treatment. In conclusion, DEPDC1 might be a novel therapeutic target for lung cancer, and the 11R-DEP:611-628 peptide is a potent apoptosis inducer in A549 cells.

  10. Targeted interfering DEP domain containing 1 protein induces apoptosis in A549 lung adenocarcinoma cells through the NF-κB signaling pathway

    PubMed Central

    Wang, Qingqing; Li, Aili; Jin, Junfei; Huang, Guojin

    2017-01-01

    Ectopic expression of DEP domain containing 1 (DEPDC1) in lung adenocarcinomas is associated with poor prognosis, but its role and the underlying mechanism remain unknown. In this study, DEPDC1 expression in lung cancer cell lines was examined with Western blot assay, and DEPDC1-positive cell A549 was selected for further experiments. DEPDC1 inhibitor miR-130a was overexpressed in A549 cells, and the proliferation and apoptosis of these cells were analyzed with cell counting and flow cytometry assay. Interfering peptide 11R-DEP:611–628 and JNK inhibitor SP600125 were used alone or in combination to treat A549 cells, and the cell proliferation and apoptosis were assessed by flow cytometry assay; caspase 3 and cleaved caspase 3, phosphor-JNK, and total JNK were detected by Western blotting; and nuclear factor kappa B (NF-κB) localization was determined by immunofluorescence staining. We found that miR-130a and 11R-DEP:611–628 peptides (5 μM) both inhibited A549 proliferation and induced apoptosis. We observed that 11R-DEP:611–628 peptide treatment resulted in elevated A20 expression, dramatically reduced nuclear NF-κB, and increased phosphor-JNK. These findings indicate that DEPDC1 inhibits apoptosis of A549 cell by suppressing A20 expression to regulate NF-κB activity, and that JNK plays a protective role upon 11R-DEP:611–628 peptide treatment. In conclusion, DEPDC1 might be a novel therapeutic target for lung cancer, and the 11R-DEP:611–628 peptide is a potent apoptosis inducer in A549 cells. PMID:28979136

  11. Combined metformin and resveratrol confers protection against UVC-induced DNA damage in A549 lung cancer cells via modulation of cell cycle checkpoints and DNA repair.

    PubMed

    Lee, Yong-Syu; Doonan, Barbara B; Wu, Joseph M; Hsieh, Tze-Chen

    2016-06-01

    Aging in humans is a multi-factorial cellular process that is associated with an increase in the risk of numerous diseases including diabetes, coronary heart disease and cancer. Aging is linked to DNA damage, and a persistent source of DNA damage is exposure to ultraviolet (UV) radiation. As such, identifying agents that confer protection against DNA damage is an approach that could reduce the public health burden of age-related disorders. Metformin and resveratrol have both shown effectiveness in preventing several age-related diseases; using human A549 cells, we investigated whether metformin or resveratrol, alone or combined, prevent UVC-induced DNA damage. We found that metformin inhibited UVC-induced upregulation of p53, as well as downregulated the expression of two DNA damage markers: γH2AX and p-chk2. Metformin also upregulated DNA repair as evidenced by the increase in expression of p53R2. Treatment with metformin also induced cell cycle arrest in UVC-induced cells, in correlation with a reduction in the levels of cyclin E/cdk2/Rb and cyclin B1/cdk1. Compared to metformin, resveratrol as a single agent showed less effectiveness in counteracting UVC-elicited cellular responses. However, resveratrol displayed synergism when combined with metformin as shown by the downregulation of p53/γH2AX/p-chk2. In conclusion, the results of the present study validate the effectiveness of metformin, alone or with the addition of resveratrol, in reducing the risk of aging by conferring protection against UV-induced DNA damage.

  12. Acetazolamide fails to decrease pulmonary artery pressure at high altitude in partially acclimatized humans.

    PubMed

    Basnyat, Buddha; Hargrove, Jenny; Holck, Peter S; Srivastav, Soni; Alekh, Kshitiz; Ghimire, Laxmi V; Pandey, Kaushal; Griffiths, Anna; Shankar, Ravi; Kaul, Komal; Paudyal, Asmita; Stasiuk, David; Basnyat, Rose; Davis, Christopher; Southard, Andrew; Robinson, Cathleen; Shandley, Thomas; Johnson, Dan W; Zafren, Ken; Williams, Sarah; Weiss, Eric A; Farrar, Jeremy J; Swenson, Erik R

    2008-01-01

    In this randomized, double-blind placebo controlled trial our objectives were to determine if acetazolamide is capable of preventing high altitude pulmonary edema (HAPE) in trekkers traveling between 4250 m (Pheriche)\\4350 m (Dingboche) and 5000 m (Lobuje) in Nepal; to determine if acetazolamide decreases pulmonary artery systolic pressures (PASP) at high altitude; and to determine if there is an association with PASP and signs and symptoms of HAPE. Participants received either acetazolamide 250 mg PO BID or placebo at Pheriche\\Dingboche and were reassessed in Lobuje. The Lake Louise Consensus Criteria were used for the diagnosis of HAPE, and cardiac ultrasonography was used to measure the velocity of tricuspid regurgitation and estimate PASP. Complete measurements were performed on 339 of the 364 subjects (164 in the placebo group, 175 in the acetazolamide group). No cases of HAPE were observed in either study group nor were differences in the signs and symptoms of HAPE found between the two groups. Mean PASP values did not differ significantly between the acetazolamide and placebo groups (31.3 and 32.6 mmHg, respectively). An increasing number of signs and symptoms of HAPE was associated with elevated PASP (p < 0.01). The efficacy of acetazolamide against acute mountain sickness, however, was significant with a 21.9% incidence in the placebo group compared to 10.2 % in the acetazolamide group (p < 0.01). Given the lack of cases of HAPE in either group, we can draw no conclusions about the efficacy of acetazolamide in preventing HAPE, but the absence of effect on PASP suggests that any effect may be minor possibly owing to partial acclimatization during the trek up to 4200 m.

  13. Lung inflammation and genotoxicity in mice lungs after pulmonary exposure to candle light combustion particles.

    PubMed

    Skovmand, Astrid; Damiao Gouveia, Ana Cecilia; Koponen, Ismo Kalevi; Møller, Peter; Loft, Steffen; Roursgaard, Martin

    2017-07-05

    Candle burning produces a large amount of particles that contribute substantially to the exposure to indoor particulate matter. The exposures to various types of combustion particles, such as diesel exhaust particles, have been associated with increased risk of lung cancer by mechanisms that involve oxidative stress, inflammation and genotoxicity. The aim of this study was to compare pulmonary effects of candle light combustion particles (CP) with two benchmark diesel exhaust particles (A-DEP and SRM2975). Intratracheal (i.t.) instillation of CP (5mg/kg bodyweight) in C57BL/6n mice produced a significant influx of alveolar macrophages and polymorphonuclear leukocytes and increased concentrations of proteins and lactate dehydrogenase activity in bronchoalveolar fluid. Lower levels of these markers of inflammation and cytotoxicity were observed after i.t. instillation of the same dose of A-DEP or SRM2975. The i.t. instillation of CP did not generate oxidative damage to DNA in lung tissue, measured as DNA strand breaks and human 8-oxoguanine glycosylase-sensitive sites by the comet assay. The lack of genotoxic response was confirmed in lung epithelial (A549) cells, although the exposure to CP increased intracellular levels of reactive oxygen species. In conclusion, pulmonary exposure to particles from burning candles is associated with inflammation and cytotoxicity in the lungs. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Deletion of the transmembrane transporter ABCG1 results in progressive pulmonary lipidosis.

    PubMed

    Baldán, Angel; Tarr, Paul; Vales, Charisse S; Frank, Joy; Shimotake, Thomas K; Hawgood, Sam; Edwards, Peter A

    2006-09-29

    We show that mice lacking the ATP-binding cassette transmembrane transporter ABCG1 show progressive and age-dependent severe pulmonary lipidosis that recapitulates the phenotypes of different respiratory syndromes in both humans and mice. The lungs of chow-fed Abcg1(-/-) mice, >6-months old, exhibit extensive subpleural cellular accumulation, macrophage, and pneumocyte type 2 hypertrophy, massive lipid deposition in both macrophages and pneumocytes and increased levels of surfactant. No such abnormalities are observed at 3 months of age. However, gene expression profiling reveals significant changes in the levels of mRNAs encoding key genes involved in lipid metabolism in both 3- and 8-month-old Abcg1(-/-) mice. These data suggest that the lungs of young Abcg1(-/-) mice maintain normal lipid levels by repressing lipid biosynthetic pathways and that such compensation is inadequate as the mice mature. Studies with A-549 cells, a model for pneumocytes type 2, demonstrate that overexpression of ABCG1 specifically stimulates the efflux of cellular cholesterol by a process that is dependent upon phospholipid secretion. In addition, we demonstrate that Abcg1(-/-), but not wild-type macrophages, accumulate cholesterol ester droplets when incubated with surfactant. Together, these data provide a mechanism to explain the lipid accumulation in the lungs of Abcg1(-/-)mice. In summary, our results demonstrate that ABCG1 plays essential roles in pulmonary lipid homeostasis.

  15. Growth factor independence-1 is expressed in primary human neuroendocrine lung carcinomas and mediates the differentiation of murine pulmonary neuroendocrine cells.

    PubMed

    Kazanjian, Avedis; Wallis, Deeann; Au, Nicholas; Nigam, Rupesh; Venken, Koen J T; Cagle, Philip T; Dickey, Burton F; Bellen, Hugo J; Gilks, C Blake; Grimes, H Leighton

    2004-10-01

    Human small cell lung cancers might be derived from pulmonary cells with a neuroendocrine phenotype. They are driven to proliferate by autocrine and paracrine neuropeptide growth factor stimulation. The molecular basis of the neuroendocrine phenotype of lung carcinomas is relatively unknown. The Achaete-Scute Homologue-1 (ASH1) transcription factor is critically required for the formation of pulmonary neuroendocrine cells and is a marker for human small cell lung cancers. The Drosophila orthologues of ASH1 (Achaete and Scute) and the growth factor independence-1 (GFI1) oncoprotein (Senseless) genetically interact to inhibit Notch signaling and specify fly sensory organ development. Here, we show that GFI1, as with ASH1, is expressed in neuroendocrine lung cancer cell lines and that GFI1 in lung cancer cell lines functions as a DNA-binding transcriptional repressor protein. Forced expression of GFI1 potentiates tumor formation of small-cell lung carcinoma cells. In primary human lung cancer specimens, GFI1 expression strongly correlates with expression of ASH1, the neuroendocrine growth factor gastrin-releasing peptide, and neuroendocrine markers synaptophysin and chromogranin A (P < 0.0000001). GFI1 colocalizes with chromogranin A and calcitonin-gene-related peptide in embryonic and adult murine pulmonary neuroendocrine cells. In addition, mice with a mutation in GFI1 display abnormal development of pulmonary neuroendocrine cells, indicating that GFI1 is important for neuroendocrine differentiation.

  16. 129 Xe chemical shift in human blood and pulmonary blood oxygenation measurement in humans using hyperpolarized 129 Xe NMR.

    PubMed

    Norquay, Graham; Leung, General; Stewart, Neil J; Wolber, Jan; Wild, Jim M

    2017-04-01

    To evaluate the dependency of the 129 Xe-red blood cell (RBC) chemical shift on blood oxygenation, and to use this relation for noninvasive measurement of pulmonary blood oxygenation in vivo with hyperpolarized 129 Xe NMR. Hyperpolarized 129 Xe was equilibrated with blood samples of varying oxygenation in vitro, and NMR was performed at 1.5 T and 3 T. Dynamic in vivo NMR during breath hold apnea was performed at 3 T on two healthy volunteers following inhalation of hyperpolarized 129 Xe. The 129 Xe chemical shift in RBCs was found to increase nonlinearly with blood oxygenation at 1.5 T and 3 T. During breath hold apnea, the 129 Xe chemical shift in RBCs exhibited a periodic time modulation and showed a net decrease in chemical shift of ∼1 ppm over a 35 s breath hold, corresponding to a decrease of 7-10 % in RBC oxygenation. The 129 Xe-RBC signal amplitude showed a modulation with the same frequency as the 129 Xe-RBC chemical shift. The feasibility of using the 129 Xe-RBC chemical shift to measure pulmonary blood oxygenation in vivo has been demonstrated. Correlation between 129 Xe-RBC signal and 129 Xe-RBC chemical shift modulations in the lung warrants further investigation, with the aim to better quantify temporal blood oxygenation changes in the cardiopulmonary vascular circuit. Magn Reson Med 77:1399-1408, 2017. © 2016 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2016 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine.

  17. Preceding human metapneumovirus infection increases adherence of Streptococcus pneumoniae and severity of murine pneumococcal pneumonia.

    PubMed

    Lai, Shen-Hao; Liao, Sui-Ling; Wong, Kin-Sun; Lin, Tzou-Yien

    2016-04-01

    Coinfection with respiratory virus and Streptococcus pneumoniae has been frequently reported in several epidemiologic studies. The aim of this study was to explore the effect of preceding human metapneumovirus (hMPV) inoculation on subsequent pneumococcal infection. Hep-2 and A549 cells were infected with hMPV then inoculated with S. pneumoniae. Bacterial adhesion was measured using colony forming unit and cytometric-fluorescence assays. In vivo bacterial adhesion was examined in hMPV-infected mice after inoculation of fluorescence-conjugated S. pneumoniae. Pulmonary inflammation (bacterial titers, cytokine levels, and histopathology) of hMPV-infected mice was investigated after inoculation with S. pneumoniae. In vitro results of bacterial infection with S. pneumoniae on A549 and Hep-2 monolayer cells showed that even though cellular adherence was variable among different serotypes, there was significantly enhanced bacterial adherence in A549 cells with preceding hMPV infection. In addition, in vivo study of hMPV-infected mice showed increased adhesion of S. pneumoniae on the bronchial epithelium with delayed bacterial clearance and exacerbated histopathology. Furthermore, mice with preceding hMPV infection showed repressed recruitment of airway neutrophils with decreased expression of neutrophil chemoattractants during pneumococcal infection. These results suggest that hMPV-infected airway cells, especially the lower airway epithelium, express increased adherence with S. pneumoniae. Furthermore, hMPV-infected mice showed impaired recruitment of airway neutrophils, possibly leading to delayed bacterial clearance and exacerbated pulmonary inflammation, after secondary infection with pneumococcal isolates. Copyright © 2014. Published by Elsevier B.V.

  18. Human γ-herpesviruses Epstein-Barr virus and human herpesvirus-8 are not detected in the lungs of patients with severe pulmonary arterial hypertension.

    PubMed

    Valmary, Séverine; Dorfmüller, Peter; Montani, David; Humbert, Marc; Brousset, Pierre; Degano, Bruno

    2011-06-01

    In susceptible individuals, multiple events may trigger pulmonary vascular remodeling and pulmonary arterial hypertension (PAH). Human herpesvirus-8 (HHV-8), a γ-herpesvirus homologous with Epstein-Barr virus (EBV), was suggested to act as a "second hit" in the development of PAH in susceptible patients. Although there is indirect evidence from in vitro and animal studies in favor of a link between γ-herpesviruses and the pathophysiology of PAH, results remain controversial. Therefore, we investigated the presence of EBV and HHV-8 in the lungs of patients with PAH. Thirty-four lungs explanted from French patients with end-stage PAH (mean age, 38 ± 14 years; 19 women) were studied. Tissue samples were incorporated into tissue microarrays. Normal lung tissues served as negative controls. Kaposi sarcoma tissue served as a positive control for HHV-8, and EBV-associated lymphoma served as a positive control for EBV. The presence of HHV-8 was investigated with immunohistochemistry and polymerase chain reaction. The presence of EBV was investigated with immunohistochemistry and in situ hybridization. For HHV-8, none of PAH lung samples showed a "stippling" nuclear pattern classically observed in HHV-8-positive Kaposi sarcoma lesions. When studied by polymerase chain reaction, all cases remained negative. For EBV, none of the PAH lung samples showed positive staining, whatever the technique applied. HHV-8 and EBV cannot be detected in the lungs of patients with end-stage PAH. The role of these γ-herpesviruses in the pathophysiology of PAH is, therefore, unlikely.

  19. Persistent Pneumocystis colonization leads to the development of chronic obstructive pulmonary disease (COPD) in a non-human primate model of AIDS

    PubMed Central

    Shipley, Timothy W.; Kling, Heather M.; Morris, Alison; Patil, Sangita; Kristoff, Jan; Guyach, Siobhan E.; Murphy, Jessica M.; Shao, Xiuping; Sciurba, Frank C.; Rogers, Robert M.; Richards, Thomas; Thompson, Paul; Montelaro, Ronald C.; Coxson, Harvey O.; Hogg, James C.; Norris, Karen A.

    2010-01-01

    HIV-infected patients are at increased risk for development of pulmonary complications, including chronic obstructive pulmonary disease (COPD). Inflammation associated with sub-clinical infection has been postulated to promote COPD. Persistence of Pneumocystis (Pc) is associated with HIV and COPD, although a causal relationship has not been established. We used a simian/human immunodeficiency virus (SHIV) model of HIV infection to study pulmonary effects of Pc colonization. SHIV-infected/Pc-colonized monkeys developed progressive obstructive pulmonary disease characterized by increased emphysematous tissue and bronchial-associated lymphoid tissue. Elevated Th2 cytokines and pro-inflammatory mediators in bronchoalveolar lavage fluid coincided with Pc colonization and pulmonary function decline. These results support the concept that an infectious agent contributes to development of HIV-associated lung disease and suggests that Pc colonization may be a risk factor for the development of HIV-associated COPD. Furthermore, this model allows examination of early host responses important to disease progression thus identifying potential therapeutic targets for COPD. PMID:20533880

  20. Functional ion channels in human pulmonary artery smooth muscle cells: Voltage-dependent cation channels

    PubMed Central

    Firth, Amy L.; Remillard, Carmelle V.; Platoshyn, Oleksandr; Fantozzi, Ivana; Ko, Eun A.; Yuan, Jason X.-J.

    2011-01-01

    The activity of voltage-gated ion channels is critical for the maintenance of cellular membrane potential and generation of action potentials. In turn, membrane potential regulates cellular ion homeostasis, triggering the opening and closing of ion channels in the plasma membrane and, thus, enabling ion transport across the membrane. Such transmembrane ion fluxes are important for excitation–contraction coupling in pulmonary artery smooth muscle cells (PASMC). Families of voltage-dependent cation channels known to be present in PASMC include voltage-gated K+ (Kv) channels, voltage-dependent Ca2+-activated K+ (Kca) channels, L- and T- type voltage-dependent Ca2+ channels, voltage-gated Na+ channels and voltage-gated proton channels. When cells are dialyzed with Ca2+-free K+- solutions, depolarization elicits four components of 4-aminopyridine (4-AP)-sensitive Kvcurrents based on the kinetics of current activation and inactivation. In cell-attached membrane patches, depolarization elicits a wide range of single-channel K+ currents, with conductances ranging between 6 and 290 pS. Macroscopic 4-AP-sensitive Kv currents and iberiotoxin-sensitive Kca currents are also observed. Transcripts of (a) two Na+ channel α-subunit genes (SCN5A and SCN6A), (b) six Ca2+ channel α–subunit genes (α1A, α1B, α1X, α1D, α1Eand α1G) and many regulatory subunits (α2δ1, β1-4, and γ6), (c) 22 Kv channel α–subunit genes (Kv1.1 - Kv1.7, Kv1.10, Kv2.1, Kv3.1, Kv3.3, Kv3.4, Kv4.1, Kv4.2, Kv5.1, Kv 6.1-Kv6.3, Kv9.1, Kv9.3, Kv10.1 and Kv11.1) and three Kv channel β-subunit genes (Kvβ1-3) and (d) four Kca channel α–subunit genes (Sloα1 and SK2-SK4) and four Kca channel β-subunit genes (Kcaβ1-4) have been detected in PASMC. Tetrodotoxin-sensitive and rapidly inactivating Na+ currents have been recorded with properties similar to those in cardiac myocytes. In the presence of 20 mM external Ca2+, membrane depolarization from a holding potential of -100 mV elicits a rapidly

  1. A fully human antimelanoma cellular adhesion molecule/MUC18 antibody inhibits spontaneous pulmonary metastasis of osteosarcoma cells in vivo.

    PubMed

    McGary, Eric C; Heimberger, Amy; Mills, Lisa; Weber, Kristy; Thomas, Gary W; Shtivelband, Mikhail; Lev, Dina Chelouche; Bar-Eli, Menashe

    2003-12-15

    The melanoma cellular adhesion molecule, also known as MUC18, is highly expressed on several tumors, including bone sarcomas. The level of MUC18 expression has been found to correlate directly with tumor progression and metastatic potential. These observations have established MUC18 as a candidate mediator of tumor growth and metastasis, and suggest that blockade of MUC18 might be a potential target for immunotherapy against several MUC18-expressing tumors, including human bone sarcomas. To investigate whether blockade of MUC18 might be a potential target for immunotherapy against osteosarcoma, we have recently developed a fully human anti-MUC18 antibody, ABX-MA1. We studied the effect of ABX-MA1 on growth, adhesion, invasion, and metastasis of human osteosaroma cells both in vitro and in vivo. MUC18 was widely expressed on both osteosarcoma and Ewing's sarcoma cells. ABX-MA1 had no effect on the proliferation of osteosarcoma cells in vitro, nor did it significantly inhibit the growth of KRIB human osteosarcoma cells when they were orthotopically implanted into the tibias of nude mice. However, after 6 weeks, significantly fewer ABX-MA1-treated mice developed spontaneous pulmonary metastases than did IgG-treated control mice. Additionally, ABX-MA1 decreased the invasion of osteosarcoma cells through Matrigel-coated filters and disrupted homotypic adhesion between osteosarcoma cells and their heterotypic interaction with human vascular endothelial cells. Our findings demonstrate that MUC18 plays a central role in the metastasis of osteosarcoma and suggest that targeted inhibition of this antigen by ABX-MA1 may be a novel immunotherapeutic approach in the management of this tumor.

  2. Role of platelet-derived growth factor-BB (PDGF-BB) in human pulmonary artery smooth muscle cell proliferation.

    PubMed

    Zhao, Yan; Lv, Wentao; Piao, Hongying; Chu, Xiaojie; Wang, Hao

    2014-08-01

    Pulmonary arterial hypertension (PAH) is a vascular remodeling disease characterized by enhanced proliferation of pulmonary artery smooth muscle cells (PASMCs) and suppressed apoptosis. Platelet-derived growth factor (PDGF) is a potent mitogen involved in cell proliferation and migration. PDGF-BB induces the proliferation and migration of PASMCs and has been proposed to be a key mediator in the progression of PAH. Previous studies have shown that PDGF and its receptor are substantially elevated in lung tissues and PASMCs isolated from patients and animals with PAH, but the underlying mechanisms are still poorly manifested. MAP kinases, including extracellular signal-regulated kinase1/2 (ERK1/2), c-Jun NH2-terminal kinase1/2 (JNK1/2), and p38 are the key intracellular signals for stimuli-induced cell proliferation, survival, and apoptosis. Therefore, the purpose of this study is to determine whether PDGF-BB on cell proliferation process is mediated through the MAP kinases pathway in human PASMCs (HPASMCs). Our results showed PDGF-BB-induced proliferating cell nuclear antigen (PCNA), Cyclin A and Cyclin E expression in a concentration-dependent manner. The expression levels of phosphorylated JNK (p-JNK) was upregulated with 20 ng/ml PDGF-BB treatment, while PDGF-BB could not increase phosphorylated ERK1/2 (p-ERK1/2) and p-38 (p-p38) expression. The effects of PDGF-BB on cell proliferation and survival were weakened after the administration of antagonist of the JNK pathway or si-JNK. In addition, PDGF-BB protected against the loss of mitochondrial membrane potentials evoked by serum deprivation (SD) in a JNK-dependent manner. These results suggest that PDGF-BB promotes HPASMCs proliferation and survival, which is likely to be mediated via the JNK pathway.

  3. Lipid-restricted recognition of mycobacterial lipoglycans by human pulmonary surfactant protein A: a surface-plasmon-resonance study.

    PubMed Central

    Sidobre, Stéphane; Puzo, Germain; Rivière, Michel

    2002-01-01

    The human pulmonary surfactant protein A (hSP-A), a member of the mammalian collectin family, is thought to play a key defensive role against airborne invading pulmonary pathogens, among which is Mycobacterium tuberculosis, the aetiologic agent of tuberculosis. hSP-A has been shown to promote the uptake and the phagocytosis of pathogenic bacilli through the recognition and the binding of carbohydrate motifs on the invading pathogen surface. Recently we identified lipomannan and mannosylated lipoarabinomannan (ManLAM), two major mycobacterial cell-wall lipoglycans, as potential ligands for binding of hSP-A. We demonstrated that both the terminal mannose residues and the fatty acids are critical for binding, whereas the inner arabinosyl and mannosyl domains do not participate. In the present study we developed a surface-plasmon-resonance assay to analyse the molecular basis for the recognition of ManLAM by hSP-A and to try to define further the role of the lipidic aglycone moiety. Binding of ManLAM to immobilized hSP-A was consistent with the simplest one-to-one interaction model involving a single class of carbohydrate-binding site. This observation strongly suggests that the lipid moiety of ManLAM does not directly interact with hSP-A, but is rather responsible for the macromolecular organization of the lipoglycan, which may be necessary for efficient recognition of the terminal mannosyl epitopes. The indirect, structural role of the lipoglycan lipidic component is further supported by the complete lack of interaction with hSP-A in the presence of a low concentration of mild detergent. PMID:12071842

  4. A critical role of nicotinamide phosphoribosyltransferase in human telomerase reverse transcriptase induction by resveratrol in aortic smooth muscle cells

    PubMed Central

    Huang, Peixin; Riordan, Sean M.; Heruth, Daniel P.; Grigoryev, Dmitry N.; Zhang, Li Qin; Ye, Shui Qing

    2015-01-01

    Aging is the predominant risk factor for cardiovascular diseases and contributes to a considerably more severe outcome in patients with acute myocardial infarction. Resveratrol, a polyphenol found in red wine, is a caloric restriction mimetic with potential anti-aging properties which has emerged as a beneficial nutraceutical for patients with cardiovascular disease. Although resveratrol is widely consumed as a nutritional supplement, its mechanism of action remains to be elucidated fully. Here, we report that resveratrol activates human nicotinamide phosphoribosyltransferase (NAMPT), SIRT4 and telomerase reverse transcriptase (hTERT) in human aortic smooth muscle cells. Similar observations were obtained in resveratrol treated C57BL/6J mouse heart and liver tissues. Resverotrol can also augment telomerase activity in both human pulmonary microvascular endothelial cells and A549 cells. Blocking NAMPT and SIRT4 expression prevents induction of hTERT in human aortic smooth muscle cells while overexpression of NAMPT elevates the telomerase activity induced by resveratrol in A549 cells. Together, these results identify a NAMPT-SIRT4-hTERT axis as a novel mechanism by which resveratrol may affect the anti-aging process in human aortic smooth muscle cells, mouse hearts and other cells. These findings enrich our understanding of the positive effects of resveratrol in human cardiovascular diseases. PMID:25926556

  5. Pulmonary Inflammatory Responses to Acute Meteorite Dust Exposures - Implications for Human Space Exploration

    NASA Technical Reports Server (NTRS)

    Harrington, A. D.; McCubbin, F. M.; Vander Kaaden, K. E.; Kaur, J.; Smirnov, A.; Galdanes, K.; Schoonen, M. A. A.; Chen, L. C.; Tsirka, S. E.; Gordon, T.

    2018-01-01

    New initiatives to send humans to Mars within the next few decades are illustrative of the resurgence of interest in space travel. However, as with all exploration, there are risks. The Human Research Roadmap developed by NASA identifies the Risk of Adverse Health and Performance Effects of Celestial Dust Exposure as an area of concern. Extended human exploration will further increase the probability of inadvertent and repeated exposures to celestial dusts.

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

    PubMed

    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. Copyright © 2016 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

  7. MMP-2 siRNA Induced Fas/CD95 Mediated Extrinsic II Apoptotic Pathway in the A549 Lung Adenocarcinoma Cell Line

    PubMed Central

    Chetty, Chandramu; Bhoopathi, Praveen; Lakka, Sajani S.; Rao, Jasti S.

    2007-01-01

    We have previously reported that the downregulation of MMP-2 by adenovirus-mediated delivery of MMP-2 siRNA (Ad-MMP-2) reduced spheroid invasion and angiogenesis in vitro, and, metastasis and tumor growth in vivo. In this study, we investigated the mechanism of Ad-MMP-2-mediated growth inhibition in vitro and in vivo. Ad-MMP-2 infection led to the induction of apoptosis as determined by TUNEL assay, Annexin-V staining and PARP-1 cleavage in a dose-dependent manner in A549 cells. Ad-MMP-2 decreased the content of the anti-apoptotic members of the Bcl-2 family proteins (Bcl-2 and Bcl-xL) and increased the content of the pro-apoptotic members of the Bcl-2 family (Bax and Bcl-xS) as determined by immunoblotting analysis. Furthermore, Ad-MMP-2-mediated apoptosis was accompanied by increase in truncated Bid, release of cytochrome-c, and the activation of caspases-8, -9 and -3. Immunoblot analysis showed that Ad-MMP-2 infection caused upregulation of Fas/Fas-L and FADD. And Anti-Fas-L antibody reversed Ad-MMP-2-induced apoptosis. TIMP-3, an endogenous inhibitor of MMP-2, which cleaves Fas-L and activates the Fas/Fas-L inducing apoptotic pathway, was increased in Ad-MMP-2-treated cells. Adenovirus-mediated expression of MMP-2 siRNA in human lung xenografts in vivo resulted in increased immunostaining of Fas, Fas-L, cleaved Bid and TIMP-3. This is the first report, to our knowledge, showing that MMP-2 inhibition upregulates TIMP-3 levels, which in turn, promotes apoptosis in lung cancer. PMID:17599056

  8. MMP-2 siRNA induced Fas/CD95-mediated extrinsic II apoptotic pathway in the A549 lung adenocarcinoma cell line.

    PubMed

    Chetty, C; Bhoopathi, P; Lakka, S S; Rao, J S

    2007-12-06

    We have previously reported that the downregulation of MMP-2 by adenovirus-mediated delivery of MMP-2 siRNA (Ad-MMP-2) reduced spheroid invasion and angiogenesis in vitro, and, metastasis and tumor growth in vivo. In this study, we investigated the mechanism of Ad-MMP-2-mediated growth inhibition in vitro and in vivo. Ad-MMP-2 infection led to the induction of apoptosis as determined by TUNEL assay, Annexin-V staining and PARP-1 cleavage in a dose-dependent manner in A549 cells. Ad-MMP-2 decreased the content of the antiapoptotic members of the Bcl-2 family proteins (Bcl-2 and Bcl-xL) and increased the content of the pro-apoptotic members of the Bcl-2 family (Bax and Bcl-xS) as determined by immunoblotting analysis. Furthermore, Ad-MMP-2-mediated apoptosis was accompanied by increase in truncated Bid, release of cytochrome c and the activation of caspase-8, -9 and -3. Immunoblot analysis showed that Ad-MMP-2 infection caused upregulation of Fas/Fas-L and FADD, and Anti-Fas-L antibody reversed Ad-MMP-2-induced apoptosis. Tissue inhibitor of metalloproteinases (TIMP)-3, an endogenous inhibitor of MMP-2, which cleaves Fas-L and activates the Fas/Fas-L inducing apoptotic pathway, was increased in Ad-MMP-2-treated cells. Adenovirus-mediated expression of MMP-2 siRNA in human lung xenografts in vivo resulted in increased immunostaining of Fas, Fas-L, cleaved Bid and TIMP-3. This is the first report, to our knowledge, showing that MMP-2 inhibition upregulates TIMP-3 levels, which in turn, promotes apoptosis in lung cancer.

  9. New Biochemical Insights into the Mechanisms of Pulmonary Arterial Hypertension in Humans

    PubMed Central

    Blanco, Isabel; Izquierdo-García, José Luis; Dudzik, Danuta; Markuszewski, Michał J.; Peinado, Victor Ivo; Laclaustra, Martín; Barberá, Joan Albert; Barbas, Coral

    2016-01-01

    Diagnosis of pulmonary arterial hypertension (PAH) is difficult due to the lack of specific clinical symptoms and biomarkers, especially at early stages. We compared plasma metabolic fingerprints of PAH patients (n = 20) with matched healthy volunteers (n = 20) using, for the first time, untargeted multiplatform metabolomics approach consisting of high-performance liquid and gas chromatography coupled with mass spectrometry. Multivariate statistical analyses were performed to select metabolites that contribute most to groups’ classification (21 from liquid in both ionization modes and 9 from gas chromatography-mass spectrometry). We found metabolites related to energy imbalance, such as glycolysis-derived metabolites, as well as metabolites involved in fatty acid, lipid and amino acid metabolism. We observed statistically significant changes in threitol and aminomalonic acid in PAH patients, which could provide new biochemical insights into the pathogenesis of the disease. The results were externally validated on independent case and control cohorts, confirming up to 16 metabolites as statistically significant in the validation study. Multiplatform metabolomics, followed by multivariate chemometric data analysis has a huge potential for explaining pathogenesis of PAH and for searching potential and new more specific and less invasive markers of the disease. PMID:27486806

  10. Hypoxia-inducible factor 1 promoter-induced JAB1 overexpression enhances chemotherapeutic sensitivity of lung cancer cell line A549 in an anoxic environment.

    PubMed

    Hu, Ming-Dong; Xu, Jian-Cheng; Fan, Ye; Xie, Qi-Chao; Li, Qi; Zhou, Chang-Xi; Mao, Mei; Yang, Yu

    2012-01-01

    The presence of lung cancer cells in anoxic zones is a key cause od chemotherapeutic resistance. Thus, it is necessary to enhance the sensitivity of such lung cancer cells. However, loss of efficient gene therapeutic targeting and inefficient objective gene expression in the anoxic zone in lung cancer are dilemmas. In the present study, a eukaryotic expression plasmid pUC57-HRE-JAB1 driven by a hypoxia response elements promoter was constructed and introduced into lung cancer cell line A549. The cells were then exposed to a chemotherapeutic drug cis-diamminedichloroplatinum (C-DDP). qRT-PCR and western blotting were used to determine the mRNA and protein level and flow cytometry to examine the cell cycle and apoptosis of A549 transfected pUC57-HRE-JAB1. The results showed that JAB1 gene in the A549 was overexpressed after the transfection, cell proliferation being arrested in G1 phase and the apoptosis ratio significantly increased. Importantly, introduction of pUC57-HRE-JAB1 significantly increased the chemotherapeutic sensitivity of A549 in an anoxic environment. In conclusion, JAB1 overexpression might provide a novel strategy to overcome chemotherapeutic resistance in lung cancer.

  11. Oxidative stress and cell cycle arrest induced by short-term exposure to dustfall PM2.5 in A549 cells.

    PubMed

    Yang, Jie; Huo, Tingting; Zhang, Xu; Ma, Jie; Wang, Yulin; Dong, Faqin; Deng, Jianjun

    2017-11-02

    It was reported that in vitro short-term exposure to PM 2.5 caused different lung diseases through inflammatory response, immune toxicity, oxidative stress, and genetic mutations. However, the complex molecular biological mechanism for its toxicity had not been fully elucidated. Therefore, the present study investigated the cytotoxicity, oxidative damage, mitochondria damage, apoptosis, and cell cycle arrest of NX and QH PM 2.5 in A549 cells. Further, cell cycle arrest-related gene levels in PM 2.5 -induced A549 cells were also detected. Our results suggested that PM 2.5 reduced the cell viability in A549 cells. Simultaneously, excessive ROS decreased MMP levels and damaged mitochondrial membrane integrity and induced mitochondrial oxidative damage through the oxygen-dependent killer route, resulting in mitochondrial damage and cell apoptosis. Besides, the results also showed that PM 2.5 induced A549 cell cycle alteration in G2/M phase after co-culture for 24 h. G2/M phase arrest was induced by upregulation of p53 and p21 and downregulation of CDK1 mRNA expression. In addition, lncRNA Sox2ot might play an important role as the specific oncogenes and it participated in G2/M phase arrest by regulating the expression of EZH 2 .

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

    PubMed

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

    2018-03-10

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

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

    PubMed

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

    2016-09-01

    Considerable interest has been gained by withasteroids because of their structural uniqueness and wide spectrum of biological activities. However, limited systematic studies for proving their cytotoxic potential have so far been reported. Hence, an attempt was made to test the cytotoxicity of six withasteroids viz., withametelin (WM), withaphysalin D, withaphysalin E, 12-deoxywithastramonolide, Withaperuvin B, and physalolactone against A549, HT-29, and MDA-MB-231 cancer cell lines. Significant cytotoxic effect of WM against A549 cells (IC 50 value of 6.0 μM), MDA-MB-231 cells (IC 50 value of 7.6 μM), and HT-29 cells (IC 50 value of 8.2 μM) was observed. Withaperuvin B and physalolactone were found to be effective against MDA-MB-231 cells. The significantly active WM arrested the A549 cells at G2/M phase and downregulated the expression of G2/M regulatory proteins such as cdc2, cyclin B1, and cdc25C. Apoptosis induced by WM in A549 cells was associated with the generation of ROS and depletion of MMP. Furthermore, WM treatment resulted in Bax upregulation, Bcl-2 downregulation, translocation of cytochrome c to mitochondria, activation of caspase-9 and -3, and PARP cleavage corroborating the apoptosis induction through intrinsic apoptotic pathway. Thus, WM possessing broader cytotoxic effect is a promising lead molecule which has the potential to be developed as a new therapeutic agent for NSCLC.

  14. Interaction of New-Developed TiO₂-Based Photocatalytic Nanoparticles with Pathogenic Microorganisms and Human Dermal and Pulmonary Fibroblasts.

    PubMed

    Nica, Ionela Cristina; Stan, Miruna Silvia; Popa, Marcela; Chifiriuc, Mariana Carmen; Lazar, Veronica; Pircalabioru, Gratiela G; Dumitrescu, Iuliana; Ignat, Madalina; Feder, Marcel; Tanase, Liviu Cristian; Mercioniu, Ionel; Diamandescu, Lucian; Dinischiotu, Anca

    2017-01-25

    TiO₂-based photocatalysts were obtained during previous years in order to limit pollution and to ease human daily living conditions due to their special properties. However, obtaining biocompatible photocatalysts is still a key problem, and the mechanism of their toxicity recently received increased attention. Two types of TiO₂ nanoparticles co-doped with 1% of iron and nitrogen (TiO₂-1% Fe-N) atoms were synthesized in hydrothermal conditions at pH of 8.5 (HT1) and 5.5 (HT2), and their antimicrobial activity and cytotoxic effects exerted on human pulmonary and dermal fibroblasts were assessed. These particles exhibited significant microbicidal and anti-biofilm activity, suggesting their potential application for microbial decontamination of different environments. In addition, our results demonstrated the biocompatibility of TiO₂-1% Fe-N nanoparticles at low doses on lung and dermal cells, which may initiate oxidative stress through dose accumulation. Although no significant changes were observed between the two tested photocatalysts, the biological response was cell type specific and time- and dose-dependent; the lung cells proved to be more sensitive to nanoparticle exposure. Taken together, these experimental data provide useful information for future photocatalytic applications in the industrial, food, pharmaceutical, and medical fields.

  15. Levofloxacin reduces inflammatory cytokine levels in human bronchial epithelia cells: implications for aerosol MP-376 (levofloxacin solution for inhalation) treatment of chronic pulmonary infections.

    PubMed

    Tsivkovskii, Ruslan; Sabet, Mojgan; Tarazi, Ziad; Griffith, David C; Lomovskaya, Olga; Dudley, Michael N

    2011-03-01

    Inflammation resulting from chronic bacterial infection in the lung contributes to long-term pulmonary complications in chronic pulmonary infections such as cystic fibrosis. Aerosol administration of levofloxacin as in the form of the investigational formulation MP-376 results in higher concentrations in lung tissues that are higher than those that can be attained with oral or intravenous dosing of levofloxacin. The objective of this study was to evaluate the effect of high concentrations of levofloxacin achieved with aerosol administration of MP-376 on proinflammatory cytokine secretion by immortalized human bronchial epithelia cells in vitro. Additionally, we investigated the potential mechanisms of the immunomodulatory effect of levofloxacin. In vitro studies in human lung epithelial cell lines showed that levofloxacin led to a dose-related reduction in IL-6 and IL-8 concentrations, with 300 μg mL(-1) resulting in the reduction of levels of IL-6 by fourfold and IL-8 by twofold (P<0.05); in contrast, tobramycin increased IL-6 levels by 50%, but had no effect on IL-8. Levofloxacin treatment did not affect the cytokine mRNA level and nuclear factor-κB-dependent promoter activity. These findings suggest that high concentrations of levofloxacin obtained in pulmonary tissues following the administration of aerosol MP-376 may provide additional benefits in patients with chronic pulmonary infections that are independent of its antibacterial properties. © 2010 MPEX Pharmaceuticals, Inc. FEMS Immunology & Medical Microbiology © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd.

  16. Pulmonary Hypertension and Pulmonary Vasodilators.

    PubMed

    Keller, Roberta L

    2016-03-01

    Pulmonary hypertension in the perinatal period can present acutely (persistent pulmonary hypertension of the newborn) or chronically. Clinical and echocardiographic diagnosis of acute pulmonary hypertension is well accepted but there are no broadly validated criteria for echocardiographic diagnosis of pulmonary hypertension later in the clinical course, although there are significant populations of infants with lung disease at risk for this diagnosis. Contributing cardiovascular comorbidities are common in infants with pulmonary hypertension and lung disease. It is not clear who should be treated without confirmation of pulmonary vascular disease by cardiac catheterization, with concurrent evaluation of any contributing cardiovascular comorbidities. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Pulmonary Embolism

    MedlinePlus

    A pulmonary embolism is a sudden blockage in a lung artery. The cause is usually a blood clot in the ... and travels through the bloodstream to the lung. Pulmonary embolism is a serious condition that can cause Permanent ...

  18. Pulmonary Fibrosis

    MedlinePlus

    Pulmonary fibrosis is a condition in which the tissue deep in your lungs becomes scarred over time. This tissue gets thick ... blood may not get enough oxygen. Causes of pulmonary fibrosis include environmental pollutants, some medicines, some connective ...

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

    PubMed

    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

  20. Aqueous extract of Sapindus mukorossi induced cell death of A549 cells and exhibited antitumor property in vivo.

    PubMed

    Liu, Min; Chen, Yen-Lin; Kuo, Yao-Haur; Lu, Mei-Kuang; Liao, Chia-Ching

    2018-03-19

    Sapindus mukorossi is a deciduous plant and has recently been recognized to have anticancer property. In the present study, we discovered that S. mukorossi leaf and stem aqueous extract (SaM) contained two polysaccharides mainly made of myo-inositol, galactose, glucose, and fructose and the aim of this study was to investigate the antitumor property the aqueous extract SaM. In vitro treatment of SaM diminished proliferative potential of lung adenocarcinomic cells and induced intracellular oxidative stress, as well as necrotic cell death. Moreover, exposure to SaM attenuated cell migration, demonstrating the effectiveness at reducing invasive property of malignant lung cells. Gene and protein expression studies indicated that SaM treatment altered the expression of proliferation/survival modulator NF-κB, tumor growth modulator ERK2, metastasis-associated molecules MMP9/12, and tumor suppressor p53 in A549 cells. Using model animals bearing Lewis lung cancer cell LL/2, we demonstrated that SaM was antitumoral and did not induce any undesired organ damage, immunotoxicity, and off-target inflammation. This work, to our knowledge, is the first study documents the antitumor bioactivity of aqueous extract riched in polysaccharides from S. mukorossi and provides insights into the potential pharmacological application of SaM as antitumor agent against lung cancer.

  1. Estradiol and nicotine exposure enhances A549 bronchioloalveolar carcinoma xenograft growth in mice through the stimulation of angiogenesis.

    PubMed

    Jarzynka, Michael J; Guo, Ping; Bar-Joseph, Ifat; Hu, Bo; Cheng, Shi-Yuan

    2006-02-01

    Angiogenesis is required for lung cancer growth, which is mediated by various growth factors such as vascular endothelial growth factor (VEGF). Increases in VEGF and angiogenesis have been correlated with poor prognosis and survival in patients with lung cancer. In addition, recent reports show that estradiol and nicotine play important roles in lung tumor initiation and progression. In this report, we demonstrate that estradiol and nicotine exposure enhances the growth of A549 bronchioloalveolar carcinoma xenografts in mice through the stimulation of cell proliferation, VEGF secretion and angiogenesis. We detect a four-fold increase in microvascular density in tumors from mice exposed to estradiol and nicotine compared to control tumors resulting in an increase in tumor growth. Intriguingly, the effects on angiogenesis and tumor growth by the combination of agents were additive when compared to either agent alone. Furthermore, estradiol promotes VEGF secretion from various non-small cell lung carcinoma (NSCLC) cells and this effect is augmented by nicotine in a tumor xenograft model. These results indicate that aside from their roles in promoting cell proliferation, estradiol and nicotine appear to have additive effects on the induction of angiogenesis through the stimulation of VEGF secretion during NSCLC progression.

  2. Study on the effect of BMSCs-EGFP-tk as mediator of HSV1-tk/GCV suicide gene therapy directed against A549 in vitro.

    PubMed

    Yang, Kun; Xu, Wen-Gui; Liu, Yong-Zhe; Meng, Xiang-Rui; Chen, Peng; Wu, Li-Chuan

    2014-01-01

    This study aims to observe the expression of HSV1-tk in mouse bone marrow mesenchymal stem cells (BMSCs-EGFP-tk) and detect the inhibition and killing effects of BMSCs as mediator of HSV1-tk/GCV on A549 cells in vitro, which can provide the experimental basis for gene therapy of lung cancer. We constructed the recombinant plasmid Vector pDON-AI-2 Neo-HSV1-tk-IRES2-EGFP with genetic engineering methods. Then we obtained the virus-like particles with infection ability after packaging the virus. The recombinant plasmid was transfected into mouse bone marrow mesenchymal stem cells in vitro. The expressions of EGFP in cells were observed by fluorescence microscopy and HSV1-tk gene was detected with RT-PCR. At last, the A549 cells and BMSCs-EGFP-tk cells were co-cultured with in vitro contact method, and the effect of BMSCs-EGFP-tk/GCV system was determined by MTT. Results indicated that the biological characteristics of BMSCs-EGFP-tk were consistent with those of BMSCs and fluorescent light expression and HSV1-tk gene expression can persist at least 15 days. The A549 cells and BMSCs-EGFP-tk cells were co-cultured and BMSCs-EGFP-tk:A549 = 2:1, adding 1 μg/mL GCV, the theory mortality is 58.44%, but actually the mortality is 90%. There is almost no difference between BMSCs-EGFP-tk and BMSCs cells in biological characteristics. The growth of A549 cells have an obviously inhibition and the bystander effect is outstanding in vitro after co-culture and this experiment lays solid foundation for the future research.

  3. Nur77 attenuates endothelin-1 expression via downregulation of NF-κB and p38 MAPK in A549 cells and in an ARDS rat model.

    PubMed

    Jiang, Yujie; Zeng, Yi; Huang, Xia; Qin, Yueqiu; Luo, Weigui; Xiang, Shulin; Sooranna, Suren R; Pinhu, Liao

    2016-12-01

    Acute respiratory distress syndrome (ARDS) is characterized by inflammatory injury to the alveolar and capillary barriers that results in impaired gas exchange and severe acute respiratory failure. Nuclear orphan receptor Nur77 has emerged as a regulator of gene expression in inflammation, and its role in the pathogenesis of ARDS is not clear. The objective of this study is to investigate the potential role of Nur77 and its underlying mechanism in the regulation of endothelin-1 (ET-1) expression in lipopolysaccharide (LPS)-induced A549 cells and an ARDS rat model. We demonstrate that LPS induced Nur77 expression and nuclear export in A549 cells. Overexpression of Nur77 markedly decreased basal and LPS-induced ET-1 expression in A549 cells, whereas knockdown of Nur77 increased the ET-1 expression. LPS-induced phosphorylation and nuclear translocation of NF-κB and p38 MAPK were blocked by Nur77 overexpression and augmented by Nur77 knockdown in A549 cells. In vivo, LPS induced Nur77 expression in lung in ARDS rats. Pharmacological activation of Nur77 by cytosporone B (CsnB) inhibited ET-1 expression in ARDS rats, decreased LPS-induced phosphorylation of NF-κB and p38 MAPK, and relieved lung, liver, and kidney injury. Pharmacological deactivation of Nur77 by 1,1-bis-(3'-indolyl)-1-(p-hydroxyphenyl)methane (DIM-C-pPhOH, C-DIM8) had no effect on ET-1 expression and lung injury. These results indicated that Nur77 decreases ET-1 expression by suppressing NF-κB and p38 MAPK in LPS-stimulated A549 cells in vitro, and, in an LPS-induced ARDS rat model, CsnB reduced ET-1 expression and lung injury in ARDS rats. Copyright © 2016 the American Physiological Society.

  4. Pulmonary edema

    MedlinePlus

    Lung congestion; Lung water; Pulmonary congestion; Heart failure - pulmonary edema ... Pulmonary edema is often caused by congestive heart failure . When the heart is not able to pump efficiently, blood can back up into the veins that take blood through the lungs. As ...

  5. Specific inhibition of AQP1 water channels in human pulmonary microvascular endothelial cells by small interfering RNAs.

    PubMed

    Gao, Chengjin; Tang, Jiajun; Li, Rongrong; Huan, Jingning

    2012-01-01

    Aquaporin (AQP)-1 is expressed in most microvasculature endothelial cells forming water channels that play major roles in a variety of physiologic processes. Our aim was to investigate the regulatory functions of AQP1 on trancellular and paracellular permeability. We designed, synthesized, and used small interfering RNAs (siRNAs) selective for AQP1 and investigated their effectiveness in altering AQP1-mediated permeability in human pulmonary microvascular endothelial cells. Twenty-four hours after transfection of ECs with siRNAs targeting two different regions of the AQP1 transcript, AQP1 protein was inhibited by 47.8% to 74.6%. siRNAs containing the same percent of base pairs as the AQP1-siRNAs but in random sequence (i.e., scrambled siRNAs) had no effect. Suppression of AQP1 expression in ECs resulted in decreases in epithelial Na+ channel (ENaC) and Na-K ATPase of ECs, and the suppression ENaC α, β, γ, and Na-K ATPase were 43.1% to 48.2%,70.0% to 76.0%, 52.6% to 55.0%, and 72.7% to 79.3%, respectively. The reduced AQP1expression also resulted in decreased cell-cell junction protein level of VE-cadherin, which was suppressed by 36.5% to 59.5% but had no effect on occludin protein. Tube formation assay and tranwell assay showed AQP1 siRNAs induced high permeability of human pulmonary microvascular endothelial cells. Rho-kinase (ROCK) I and ROCK II were increased by 46.0% to 50.0% and 59% to 81%, respectively, AQP1 siRNA treatment accelerated the formation of F-actin bundles, demonstrating the activation of Rho/ROCK signaling pathway, and decreased mitochondrial membrane potential after AQP1 siRNA treatment, showing an important event of apoptosis process. The data demonstrate that AQP1 is a critical participate in regulating endothelial permeability and barrier function and provide direct evidence of the contribution of AQP1 to blood vessel formation.

  6. Learn About Pulmonary Fibrosis

    MedlinePlus

    ... What Is Pulmonary Fibrosis? Types, Causes and Risk Factors of Pulmonary Fibrosis Symptoms of Pulmonary Fibrosis How Is Pulmonary Fibrosis Diagnosed? Stages of Pulmonary Fibrosis What Is the Life Expectancy of Someone with Pulmonary Fibrosis? Patients Overview How ...

  7. Combination therapy of advanced invasive pulmonary aspergillosis in transiently neutropenic rats using human pharmacokinetic equivalent doses of voriconazole and anidulafungin.

    PubMed

    van de Sande, Wendy W J; Mathot, Ron A A; ten Kate, Marian T; van Vianen, Wim; Tavakol, Mehri; Rijnders, Bart J A; Bakker-Woudenberg, Irma A J M

    2009-05-01

    At present, voriconazole (VOR) is the drug of first choice for treating invasive pulmonary aspergillosis (IPA). However, particularly in advanced stages of disease and in the severely immunocompromised host, the mortality remains substantial. The combination of VOR with an echinocandin may improve the therapeutic outcome. We investigate here whether combining VOR and anidulafungin (ANI) in advanced IPA in transiently neutropenic rats results in a higher therapeutic efficacy. Since VOR is metabolized more rapidly in rodents than in humans, dosage adjustment for VOR is necessary to obtain an area under the plasma concentration-time curve (AUC) in rodents that is equivalent to that of humans. In this study, the pharmacokinetics of VOR and ANI in rats were elucidated, and dosage schedules were applied that produced AUCs similar to those of humans. The developed dose schedules were well tolerated by the rats, without effects on renal and hepatic functions. VOR showed excellent efficacy in early IPA (100% rat survival). In advanced IPA, VOR was less efficacious (50% rat survival), whereas a significant decrease in galactomannan concentrations in lungs and sera was found in surviving rats. ANI administered in advanced IPA resulted in 22% rat survival, and the serum concentrations of fungal galactomannan were slightly but not significantly decreased. The addition of ANI to VOR did not result in significantly increased therapeutic efficacy in advanced IPA, resulting in 67% rat survival and a significant decrease in galactomannan concentration in serum. In conclusion, VOR monotherapy is therapeutically effective in the treatment of advanced-stage IPA and superior to the use of ANI. Combining both agents does not significantly improve the therapeutic outcome.

  8. Signal Transducers and Activators of Transcription-3/Pim1 Axis Plays a Critical Role in the Pathogenesis of Human Pulmonary Arterial Hypertension

    PubMed Central

    Paulin, Roxane; Courboulin, Audrey; Meloche, Jolyane; Mainguy, Vincent; de la Roque, Eric Dumas; Saksouk, Nehmé; Côté, Jacques; Provencher, Steeve; Sussman, Mark A.; Bonnet, Sébastien

    2012-01-01

    Background Pulmonary artery hypertension (PAH) is a proliferative disorder associated with enhanced pulmonary artery smooth muscle cell proliferation and suppressed apoptosis. The sustainability of this phenotype required the activation of a prosurvival transcription factor like signal transducers and activators of transcription-3 (STAT3) and nuclear factor of activated T cell (NFAT). Because these factors are implicated in several physiological processes, their inhibition in PAH patients could be associated with detrimental effects. Therefore, a better understanding of the mechanism accounting for their expression/activation in PAH pulmonary artery smooth muscle cells is of great therapeutic interest. Methods and Results Using multidisciplinary and translational approaches, we demonstrated that STAT3 activation in both human and experimental models of PAH accounts for the expression of both NFATc2 and the oncoprotein kinase Pim1, which trigger NFATc2 activation. Because Pim1 expression correlates with the severity of PAH in humans and is confined to the PAH pulmonary artery smooth muscle cell, Pim1 was identified as an attractive therapeutic target for PAH. Indeed, specific Pim1 inhibition in vitro decreases pulmonary artery smooth muscle cell proliferation and promotes apoptosis, all of which are sustained by NFATc2 inhibition. In vivo, tissue-specific inhibition of Pim1 by nebulized siRNA reverses monocrotaline-induced PAH in rats, whereas Pim1 knockout mice are resistant to PAH development. Conclusion We demonstrated for the first time that inhibition of the inappropriate activation of STAT3/Pim1 axis is a novel, specific, and attractive therapeutic strategy to reverse PAH. PMID:21382889

  9. Pulmonary haptoglobin (pHp) is part of the surfactant system in the human lung

    PubMed Central

    2012-01-01

    Abstract Since the existence of pHp was demonstrated, it has been shown that this molecule and its receptor CD163 are regulated by different stimuli. Furthermore, a comparably fast secretion of pHp was described as well as the immuno-stimulatory effects. The intention of this study was to elucidate the role of pHp in the human lungs further. Here we show, by means of confocal microscopy and immune-electron-microscopy, a clear co-localization of pHp with Surfactant protein-B in lamellar bodies of Alveolar Epithelial Cells Type II. These results are underlined by immunohistochemical stainings in differently fixed human lung tissues, which show pHp in vesicular and released form. The images of the released form resemble the intended position of surfactant in the human alveolus. pHp is secreted by Alveolar epithelial cells type II as previously shown. Moreover, pHp is co-localized with Surfactant protein-B. We conclude that the presented data shows that pHp is a native part of the surfactant system in the human lung. Virtual slides http://www.diagnosticpathology.diagnomx.eu/vs/2563584738239912. PMID:23164167

  10. Pulmonary haptoglobin (pHp) is part of the surfactant system in the human lung.

    PubMed

    Abdullah, Mahdi; Goldmann, Torsten

    2012-11-20

    Since the existence of pHp was demonstrated, it has been shown that this molecule and its receptor CD163 are regulated by different stimuli. Furthermore, a comparably fast secretion of pHp was described as well as the immuno-stimulatory effects. The intention of this study was to elucidate the role of pHp in the human lungs further. Here we show, by means of confocal microscopy and immune-electron-microscopy, a clear co-localization of pHp with surfactant protein-B in lamellar bodies of alveolar epithelial cells type II. These results are underlined by immunohistochemical stainings in differently fixed human lung tissues, which show pHp in vesicular and released form. The images of the released form resemble the intended position of surfactant in the human alveolus. pHp is secreted by Alveolar epithelial cells type II as previously shown. Moreover, pHp is co-localized with Surfactant protein-B. We conclude that the presented data shows that pHp is a native part of the surfactant system in the human lung. http://www.diagnosticpathology.diagnomx.eu/vs/2563584738239912.

  11. Extracts of Magnoliae flos inhibit inducible nitric oxide synthase via ERK in human respiratory epithelial cells.

    PubMed

    Baek, Jin Ah; Lee, Yang Deok; Lee, Chan Bog; Go, Hyeon Kyu; Kim, Jin Pyo; Seo, Jeong Ju; Rhee, Yang Keun; Kim, A Mi; Na, Dong Jib

    2009-03-01

    Nitric oxide (NO) is a marker of pulmonary inflammation. In asthma, the levels of exhaled NO are elevated and the source of this increased NO is inducible nitric oxide synthase (iNOS) within airway epithelial cells. Epimagnolin and fargesin are compounds isolated from the ethanol extract of Magnoliae flos, the seed of the Magnolia plant and are used to treat nasal congestion, headache and sinusitis in Asian countries. This study investigated whether epimagnolin and fargesin inhibit extracellular signal-regulated kinase (ERK) activation and decrease iNOS expression and NO production in stimulated human respiratory epithelial cells. An immortal Type II alveolar cell line of human origin (A549) was stimulated by cytomix (CM), composed of IL-1beta, TNF-alpha and IFN-gamma, with or without concurrent exposure to M. flos extract (epimagnolin or fargesin). CM-induced levels of NO production, iNOS expression and ERK activation were evaluated. A549 cells stimulated with CM showed increases in iNOS mRNA and protein expression, and NO synthesis. However, treatment with epimagnolin or fargesin decreased levels of iNOS mRNA and protein expression, and NO synthesis. CM stimulated a rapid increase in the activity of ERK, whereas epimagnolin and fargesin inhibited ERK phosphorylation. Epimagnolin and fargesin inhibit iNOS expression and decrease production of NO via ERK pathway in cytokine-stimulated human respiratory epithelial cells.

  12. Effect of Acetazolamide and Gingko Biloba on the Human Pulmonary Vascular Response to an Acute Altitude Ascent

    PubMed Central

    Ke, Tao; Wang, Jiye; Swenson, Erik R.; Zhang, Xiangnan; Hu, Yunlong; Chen, Yaoming; Liu, Mingchao; Zhang, Wenbin; Zhao, Feng; Shen, Xuefeng; Yang, Qun

    2013-01-01

    Abstract Ke, Tao, Jiye Wang, Erik R. Swenson, Xiangnan Zhang, Yunlong Hu, Yaoming Chen, Mingchao Liu, Wenbin Zhang, Feng Zhao, Xuefeng Shen, Qun Yang, Jingyuan Chen, and Wenjing Luo. Effect of acetazolamide and gingko biloba on the human pulmonary vascular response to an acute altitude ascent. High Alt Med Biol 14:162–167, 2013.—Acetazolamide and gingko biloba are the two most investigated drugs for the prevention of acute mountain sickness (AMS). Evidence suggests that they may also reduce pulmonary artery systolic pressure (PASP). To investigate whether these two drugs for AMS prevention also reduce PASP with rapid airlift ascent to high altitude, a randomized controlled trial was conducted on 28 healthy young men with acetazolamide (125 mg bid), gingko biloba (120 mg bid), or placebo for 3 days prior to airlift ascent (397 m) and for the first 3 days at high altitude (3658 m). PASP, AMS, arterial oxygen saturation (Sao2), mean arterial pressure (MAP), heart rate (HR), forced vital capacity (FVC), forced expiratory volume in the first second (FEV1), and peak expiratory flow (PEF) were assessed both at 397 m and 3658 m. HR, PEF, and PASP increased with altitude exposure (p<0.05), and SaO2 decreased (p<0.05). PASP with acetazolamide (mean at 3658 m, 26.2 mm Hg; incremental change, 4.7 mm Hg, 95% CI., 2.6–6.9 mm Hg) was lower than that with ginkgo biloba (mean at 3658 m, 33.7 mm Hg, p=0.001; incremental change, 13.1 mm Hg, 95%CI., 9.6–16.5 mm Hg, p=0.002), and with placebo (mean at 3658 m, 34.7 mm Hg, p<0.001; 14.4 mm Hg, 95% CI., 8.8–20.0 mm Hg, p=0.001). The data show that a low prophylactic dosage of acetazolamide, but not gingko biloba, mitigates the early increase of PASP in a quick ascent profile. PMID:23795737

  13. Gemcitabine-(5'-phosphoramidate)-[anti-IGF-1R]: molecular design, synthetic organic chemistry reactions, and antineoplastic cytotoxic potency in populations of pulmonary adenocarcinoma (A549).

    PubMed

    Coyne, Cody P; Narayanan, Lakshmi

    2017-03-01

    One molecular-based approach that increases potency and reduces dose-limited sequela is the implementation of selective 'targeted' delivery strategies for conventional small molecular weight chemotherapeutic agents. Descriptions of the molecular design and organic chemistry reactions that are applicable for synthesis of covalent gemcitabine-monophosphate immunochemotherapeutics have to date not been reported. The covalent immunopharmaceutical, gemcitabine-(5'-phosphoramidate)-[anti-IGF-1R] was synthesized by reacting gemcitabine with a carbodiimide reagent to form a gemcitabine carbodiimide phosphate ester intermediate which was subsequently reacted with imidazole to create amine-reactive gemcitabine-(5'-phosphorylimidazolide) intermediate. Monoclonal anti-IGF-1R immunoglobulin was combined with gemcitabine-(5'-phosphorylimidazolide) resulting in the synthetic formation of gemcitabine-(5'-phosphoramidate)-[anti-IGF-1R]. The gemcitabine molar incorporation index for gemcitabine-(5'-phosphoramidate)-[anti-IGF-R1] was 2.67:1. Cytotoxicity Analysis - dramatic increases in antineoplastic cytotoxicity were observed at and between the gemcitabine-equivalent concentrations of 10 -9  M and 10 -7  M where lethal cancer cell death increased from 0.0% to a 93.1% maximum (100.% to 6.93% residual survival), respectively. Advantages of the organic chemistry reactions in the multistage synthesis scheme for gemcitabine-(5'-phosphoramidate)-[anti-IGF-1R] include their capacity to achieve high chemotherapeutic molar incorporation ratios; option of producing an amine-reactive chemotherapeutic intermediate that can be preserved for future synthesis applications; and non-dedicated organic chemistry reaction scheme that allows substitutions of either or both therapeutic moieties, and molecular delivery platforms. © 2016 The Authors Chemical Biology & Drug Design Published by John Wiley & Sons Ltd.

  14. Cell division cycle 25 homolog c effects on low-dose hyper-radiosensitivity and induced radioresistance at elevated dosage in A549 cells.

    PubMed

    Zhao, Yanxia; Cui, Yingshan; Han, Jun; Ren, Jinghua; Wu, Gang; Cheng, Jing

    2012-09-01

    The underlying mechanisms behind both low-dose hyper-radiosensitivity (HRS) and induced radioresistance (IRR), generally occurring at elevated radiation levels, remain unclear; however, elucidation of the relationship between cell cycle division 25 homolog c (Cdc25c) phosphatase and HRS/IRR may provide important insights into this process. Two cell lines with disparate HRS status, A549 and SiHa cells, were selected as cell models for comparison of dose-dependent Cdc25c phosphatase expression subsequent to low-dose irradiation. Knockdown of Cdc25c in A549 cells was mediated by transfection with a pGCsi-RAN-U6neo vector containing hairpin siRNA sequences. S216-phosphorylated Cdc25c protein [p-Cdc25c (Ser216)], cell survival and mitotic ratio were measured by western blot, colony-forming assay and histone H3 phosphorylation analysis. Variant p-Cdc25c (Ser216) expression was observed in the two cell lines after irradiation. The p-Cdc25c (Ser216) expression noted in SiHa cells after administration of 0-1 Gy radiation was similar to the radioresistance model; however, in A549 cells, the dose response for the phosphorylation of the Cdc25c Ser216 residue overlapped the level required to overcome the HRS response. Furthermore, Cdc25c repression prior to low-dose radiation induced more distinct HRS and prevented the development of IRR. The dose required to overcome the HRS response coincided with the effect of early G2-phase checkpoint arrest in A549 cells (approximately 0.3 Gy), and Cdc25c knockdown in A549 cells (approximately 0.5 Gy) corresponded to the phosphorylation of the Cdc25c Ser216 residue. Resultant data confirmed that dose-dependent Cdc25c phosphatase does effectively act as an early G2-phase checkpoint, thus indicating mechanistic importance in the HRS to IRR transition in A549 cells.

  15. Enhanced cytotoxic activity of endophytic bacterial extracts fromAdhatoda beddomeileaves in A549 lung cancer cell lines.

    PubMed

    Swarnalatha, Y; Saha, Bhaswti

    2016-01-01

    The current study is aimed to isolate and study the efficacy of the anticancer activity of endophytic bacteria from adhatoda beddomei leaves. Endophytic bacteria, microorganisms can found in the plant tissues, like leaves, branches and roots and able to produce various novel secondary metabolites for the medicinal applications. Endophytic bacteria were isolated from the leaves of the adhatoda beddomei leaves. The extract from the culture was tested for the cytotoxicity in A549 cell lines using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)assay, dual staining and nuclear staining. The expression of the apoptotic and proliferative genes were assessed with the reverse transcriptase polymerase chain reaction (RT-PCR) comparing with the control gene. The inhibitory concentration (IC50) of the bacterial extract was found to be 43.97 μg/ml. With the dual staining the apoptotic cell percentage was significantly increased (P < 0.001) and with the 40μg/ml and 80μg/ml concentration the apoptotic percentages observed as 67% and 89% respectively. Similar concentrations were used for the nuclear fragmentation (PI) and the cell cycle analysis (FACS) using WinMDI 2.9 software. During cell cycle analysis the accumulation of the cells at G0-G1 stage was observed with increasing concentrations of the chi-alg encophytic bacterial extract nanoparticles. Finally the proapoptotic and proliferative gene expression for the bax, Bcl-2 and caspase was significantly regulated (P < 0.01; P < 0.05). The Bax and Caspase were up-regulated and Bcl-2 was down regulated. The results conclude that enophytic bacterial extract possess good cytotoxic activity.

  16. Activation of Notch signaling by short-term treatment with Jagged-1 enhances store-operated Ca(2+) entry in human pulmonary arterial smooth muscle cells.

    PubMed

    Yamamura, Hisao; Yamamura, Aya; Ko, Eun A; Pohl, Nicole M; Smith, Kimberly A; Zeifman, Amy; Powell, Frank L; Thistlethwaite, Patricia A; Yuan, Jason X-J

    2014-05-01

    Notch signaling plays a critical role in controlling proliferation and differentiation of pulmonary arterial smooth muscle cells (PASMC). Upregulated Notch ligands and Notch3 receptors in PASMC have been reported to promote the development of pulmonary vascular remodeling in patients with pulmonary arterial hypertension (PAH) and in animals with experimental pulmonary hypertension. Activation of Notch receptors by their ligands leads to the cleavage of the Notch intracellular domain (NICD) to the cytosol by γ-secretase; NICD then translocates into the nucleus to regulate gene transcription. In this study, we examined whether short-term activation of Notch functionally regulates store-operated Ca(2+) entry (SOCE) in human PASMC. Treatment of PASMC with the active fragment of human Jagged-1 protein (Jag-1) for 15-60 min significantly increased the amplitude of SOCE induced by passive deletion of Ca(2+) from the intracellular stores, the sarcoplasmic reticulum (SR). The Jag-1-induced enhancement of SOCE was time dependent: the amplitude was maximized at 30 min of treatment with Jag-1, which was closely correlated with the time course of Jag-1-mediated increase in NICD protein level. The scrambled peptide of Jag-1 active fragment had no effect on SOCE. Inhibition of γ-secretase by N-[N-(3,5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester (DAPT) significantly attenuated the Jag-1-induced augmentation of SOCE. In addition to the short-term effect, prolonged treatment of PASMC with Jag-1 for 48 h also markedly enhanced the amplitude of SOCE. These data demonstrate that short-term activation of Notch signaling enhances SOCE in PASMC; the NICD-mediated functional interaction with store-operated Ca(2+) channels (SOC) may be involved in the Jag-1-mediated enhancement of SOCE in human PASMC.

  17. The Psa Fimbriae of Yersinia pestis Interact with Phosphatidylcholine on Alveolar Epithelial Cells and Pulmonary Surfactant▿

    PubMed Central

    Galván, Estela M.; Chen, Huaiqing; Schifferli, Dieter M.

    2007-01-01

    The pH 6 antigen (Psa) of Yersinia pestis consists of fimbriae with adhesive properties of potential importance for the pathogenesis of plague, including pneumonic plague. The Psa fimbriae mediate bacterial binding to human alveolar epithelial cells. The Psa fimbriae bound mostly to one component present in the total lipid extract from type II alveolar epithelial cells of the cell line A549 separated by thin-layer chromatography (TLC). The Psa receptor was identified as phosphatidylcholine (PC) by TLC using alkali treatment, molybdenum blue staining, and Psa overlays. The Psa fimbriae bound to PC in a dose-dependent manner, and binding was inhibited by phosphorylcholine (ChoP) and choline. Binding inhibition was dose dependent, although only high concentrations of ChoP completely blocked Psa binding to PC. In contrast, less than 1 μM of a ChoP-polylysine polymer inhibited specifically the adhesion of Psa-fimbriated Escherichia coli to PC, and type I (WI-26 VA4) and type II alveolar epithelial cells. These results indicated that the homopolymeric Psa fimbriae are multimeric adhesins. Psa also bound to pulmonary surfactant, which covers the alveolar surface as a product of type II alveolar epithelial cells and includes PC as the major component. The observed dose-dependent interaction of Psa with pulmonary surfactant was blocked by ChoP. Interestingly, surfactant did not inhibit Psa-mediated bacterial binding to alveolar cells, suggesting that both surfactant and cell membrane PC retain Psa-fimbriated bacteria on the alveolar surface. Altogether, the results indicate that Psa uses the ChoP moiety of PC as a receptor to mediate bacterial binding to pulmonary surfactant and alveolar epithelial cells. PMID:17178780

  18. The Psa fimbriae of Yersinia pestis interact with phosphatidylcholine on alveolar epithelial cells and pulmonary surfactant.

    PubMed

    Galván, Estela M; Chen, Huaiqing; Schifferli, Dieter M

    2007-03-01

    The pH 6 antigen (Psa) of Yersinia pestis consists of fimbriae with adhesive properties of potential importance for the pathogenesis of plague, including pneumonic plague. The Psa fimbriae mediate bacterial binding to human alveolar epithelial cells. The Psa fimbriae bound mostly to one component present in the total lipid extract from type II alveolar epithelial cells of the cell line A549 separated by thin-layer chromatography (TLC). The Psa receptor was identified as phosphatidylcholine (PC) by TLC using alkali treatment, molybdenum blue staining, and Psa overlays. The Psa fimbriae bound to PC in a dose-dependent manner, and binding was inhibited by phosphorylcholine (ChoP) and choline. Binding inhibition was dose dependent, although only high concentrations of ChoP completely blocked Psa binding to PC. In contrast, less than 1 muM of a ChoP-polylysine polymer inhibited specifically the adhesion of Psa-fimbriated Escherichia coli to PC, and type I (WI-26 VA4) and type II alveolar epithelial cells. These results indicated that the homopolymeric Psa fimbriae are multimeric adhesins. Psa also bound to pulmonary surfactant, which covers the alveolar surface as a product of type II alveolar epithelial cells and includes PC as the major component. The observed dose-dependent interaction of Psa with pulmonary surfactant was blocked by ChoP. Interestingly, surfactant did not inhibit Psa-mediated bacterial binding to alveolar cells, suggesting that both surfactant and cell membrane PC retain Psa-fimbriated bacteria on the alveolar surface. Altogether, the results indicate that Psa uses the ChoP moiety of PC as a receptor to mediate bacterial binding to pulmonary surfactant and alveolar epithelial cells.

  19. Regulation of deprotonation of 3,3-di(1H-tetrazol-5-yl)pentanedioic acid: Solvothermal synthesis of La(III) and heterometallic La(III)/Cu(II) compounds for ablation of A549 cells

    NASA Astrophysics Data System (ADS)

    Guo, Meng Yue; Zhang, Xin; Zhao, Liang; Li, Yue Kang; Chen, Dian Yu; Yang, Gao Wen; Li, Qiao Yun

    2018-03-01

    3,3-di(1H-tetrazol-5-yl)pentanedioic acid (H4dtzpda) can display tunable valence when reacted with different metal ions because it has four acidic hydrogen atoms of both tetrazole rings and carboxylate groups. Solvothermal reactions of H4dtzpda with La(NO3)2·6H2O or La(NO3)3·6H2O/Cu(NO3)2·6H2O afforded a one dimensional [La(Hdtzpda)(H2O)4]·4H2O (1) and a three dimensional [La2Cu(dtzpda)2(H2O)10]·4H2O (2), respectively, where only three acidic hydrogen atoms of H4dtzpda are deprotonated in compound 1 while all the four acidic ones are deprotonated in compound 2. In compound 1, Hdtzpda3- is a penta-dentate ligand to bridge La(III) centers via only oxygen atoms of the carboxylate group while in compound 2, dtzpda4- is a hepta-dentate one via not only the oxygen atoms of the carboxylate group but also the nitrogen atoms of the tetrazole rings. PEG-5000 (poly(ethyleneglycol-5000)) coated compound 1 or 2 nanoparticles (NPs) have good dispersity in water. Cytotoxicity study on A549 cells (human caucasian lung carcinoma) shows that compound 2 (half-maximal inhibitory concentration, IC50~18 μg/mL) is superior to compound 1 (IC50~38 μg/mL). Furthermore, both NPs of the two compounds are able to inhibit the migration of A549 cells, indicating their potential to inhibit the transfer of tumors in vivo.

  20. The influence of Hurricanes Katrina and Rita on the inflammatory cytokine response and protein expression in A549 cells exposed to PM2.5 collected in the Baton Rouge-Port Allen industrial corridor of Southeastern Louisiana in 2005.

    PubMed

    Bourgeois, Brian; Owens, John Wesley

    2014-03-01

    Hurricanes Katrina and Rita hit the coast of Louisiana in 2005 and killed more than 2000 people. The two storms resulted in a significant spike in particulate matter (PM2.5) levels across the state of Louisiana. This report focuses on PM2.5 samples collected in 2005 from two monitoring sites in the neighboring cities of Baton Rouge and Port Allen, Louisiana. Inductively coupled plasma (ICP) revealed the presence of PM2.5-adsorbed representative and Fenton-active transition metals. Gas chromatography/mass spectrometry (GC-MS) analyses revealed the presence of 23 PAH compounds. Endotoxins were also detected. Metals and endotoxins were extracted with water. PAH were extracted with dichloromethane. In order to assess cytotoxicity, aqueous PM2.5 extracts were introduced to A549 Human Epithelial Lung Carcinoma Cells. Results indicated decreased cell viability in a dose-dependent manner, with an LC50 of 235 µg/ml and 250 µg/ml, respectively, for the two sites featured here. Endotoxins alone were not cytotoxic. The concentration of reactive oxygen species (ROS) and released LDH activity increased following exposure of A549 cells to aqueous PM2.5 extracts. Fluorescence microscopy revealed apoptotic and necrotic cell death mechanisms. ELISA revealed increased secretion of primary pro-inflammatory cytokines, IL-6, IL-8, and TNF-α. Global PCR gene expression revealed up-regulation of proteins associated with the cytokine storm; e.g. interleukins, chemokines, and TNF-α. Global antibody microarray was consistent with an inflammatory response, with up-regulation of cytokines involved in the down-field activation of the caspase cascade and kinase pathways. The up-regulation of metal-redox sensitive transcription factors, NF-κβ and AP-1, is consistent with a cell death mechanism initiated by Fenton-active transition metal redox catalysis.

  1. Assessment of Cellular Responses after Short- and Long-Term Exposure to Silver Nanoparticles in Human Neuroblastoma (SH-SY5Y) and Astrocytoma (D384) Cells

    PubMed Central

    Manzo, Luigi; Bellotti, Vittorio

    2014-01-01

    Silver nanoparticle (AgNP, 20 nm) neurotoxicity was evaluated by an integrated in vitro testing protocol employing human cerebral (SH-SY5Y and D384) cell lines. Cellular response after short-term (4–48 h, 1–100 μg/ml) and prolonged exposure (up to 10 days, 0.5–50 μg/ml) to AgNP was assessed by MTT, calcein-AM/PI, clonogenic tests. Pulmonary A549 cells were employed for data comparison along with silver nitrate as metal ionic form. Short-term data: (i) AgNP produced dose- and time-dependent mitochondrial metabolism changes and cell membrane damage (effects starting at 25 μg/ml after 4 h: EC50s were 40.7 ± 2.0 and 49.5 ± 2.1 μg/ml for SH-SY5Y and D384, respectively). A549 were less vulnerable; (ii) AgNP doses of ≤ 18 μg/ml were noncytotoxic; (iii) AgNO3 induced more pronounced effects compared to AgNP on cerebral cells. Long-term data: (i) low AgNP doses (≤1 μg/ml) compromised proliferative capacity of all cell types (cell sensibility: SHSY5Y > A549 > D384). Colony number decrease in SH-SY5Y and D384 was 50% and 25%, respectively, at 1 μg/ml, and lower dose (0.5 μg/ml) was significantly effective towards SH-SY5Y and pulmonary cells; (ii) cell proliferation activity was more affected by AgNO3 than AgNPs. In summary, AgNP-induced cytotoxic effects after short-term and prolonged exposure (even at low doses) were evidenced regardless of cell model types. PMID:24693232

  2. miR-4632 mediates PDGF-BB-induced proliferation and antiapoptosis of human pulmonary artery smooth muscle cells via targeting cJUN.

    PubMed

    Qian, Zhengjiang; Li, Yanjiao; Chen, Jidong; Li, Xiang; Gou, Deming

    2017-10-01

    MicroRNAs (miRNAs) can regulate the proliferative status of pulmonary artery smooth muscle cells (PASMCs), which is a core factor modulating pulmonary vascular remodeling diseases, such as atherosclerosis and pulmonary arterial hypertension (PAH). Our previous work has shown that miR-4632, a rarely reported miRNA, is significantly downregulated in platelet-derived growth factor (PDGF)-BB-stimulated human pulmonary artery smooth muscle cells (HPASMCs), yet its cell function and the underlying molecular mechanisms remain to be elucidated. Here, we find that miR-4632 is highly expressed in HPASMCs and its expression significantly decreased in response to different stimuli. Functional studies revealed that miR-4632 inhibited proliferation and promoted apoptosis of HPASMCs but had no effects on cell contraction and migration. Furthermore, the cJUN was identified as a direct target gene of miR-4632, while knockdown of cJUN was necessary for miR-4632-mediated HPASMC proliferation and apoptosis. In addition, the downregulation of miR-4632 by PDGF-BB was found to associate with histone deacetylation through the activation of PDGF receptor/phosphatidylinositol 3'-kinase/histone deacetylase 4 signaling. Finally, the expression of miR-4632 was reduced in the serum of patients with PAH. Overall, our results suggest that miR-4632 plays an important role in regulating HPASMC proliferation and apoptosis by suppression of cJUN, providing a novel therapeutic miRNA candidate for the treatment of pulmonary vascular remodeling diseases. It also implies that serum miR-4632 has the potential to serve as a circulating biomarker for PAH diagnosis. Copyright © 2017 the American Physiological Society.

  3. Regulation of Pulmonary and Systemic Bacterial Lipopolysaccharide Responses in Transgenic Mice Expressing Human Elafin

    PubMed Central

    Sallenave, J.-M.; Cunningham, G. A.; James, R. M.; McLachlan, G.; Haslett, C.

    2003-01-01

    The control of lung inflammation is of paramount importance in a variety of acute pathologies, such as pneumonia, the acute respiratory distress syndrome, and sepsis. It is becoming increasingly apparent that local innate immune responses in the lung are negatively influenced by systemic inflammation. This is thought to be due to a local deficit in cytokine responses by alveolar macrophages and neutrophils following systemic bacterial infection and the development of a septic response. Recently, using an adenovirus-based strategy which overexpresses the human elastase inhibitor elafin locally in the lung, we showed that elafin is able to prime lung innate immune responses. In this study, we generated a novel transgenic mouse strain expressing human elafin and studied its response to bacterial lipopolysaccharide (LPS) when the LPS was administered locally in the lungs and systemically. When LPS was delivered to the lungs, we found that mice expressing elafin had lower serum-to-bronchoalveolar lavage ratios of proinflammatory cytokines, including tumor necrosis factor alpha (TNF-α), macrophage inflammatory protein 2, and monocyte chemoattractant protein 1, than wild-type mice. There was a concomitant increase in inflammatory cell influx, showing that there was potential priming of innate responses in the lungs. When LPS was given systemically, the mice expressing elafin had reduced levels of serum TNF-α compared to the levels in wild-type mice. These results indicate that elafin may have a dual function, promoting up-regulation of local lung innate immunity while simultaneously down-regulating potentially unwanted systemic inflammatory responses in the circulation. PMID:12819058

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

    PubMed

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

    2017-01-01

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

  5. Cooking oil fume-derived PM2.5 induces apoptosis in A549 cells and MAPK/NF-кB/STAT1 pathway activation.

    PubMed

    Dou, Changming; Zhang, Jie; Qi, Cuicui

    2018-04-01

    Cooking oil fumes (COFs) are the major sources of indoor air pollution in Asia. It is well known that alveolar cells are key participants in the development of respiratory system; however, it still remains unknown whether alveolar cells are affected by COFs. Therefore, the present study investigated the effects of COFs on alveolar cells (A549 cells) and illuminated its apoptotic mechanism in response to COF-PM 2.5 exposure. When A549 cells were exposed to COF-PM 2.5 , cell viability was substantially decreased, while the generation of ROS increased, and LDH levels and CCK-8 levels gradually changed within a dose-dependent manner. The nitrite concentration in the supernatants was augmented, while the SOD activity and GSH recycling were decreased upon COF-PM 2.5 . Moreover, COF-PM 2.5 treatment increased mRNA levels of COX-2, inducible NO synthase, and TNF-α, and Elisa assay suggested that secretory proteins IL-6 and TNF-α were also increased. Furthermore, the Bax/Bcl-2 mRNA ratio was increased, and cleaved caspase-3 protein was activated in the A549 cells. Strikingly, COF-PM 2.5 induced the phosphorylation of STAT1 at Tyr701/Ser727 and activation of NF-кB and ERK1/2, p38, and JNK of the MAPK pathway. In short, our study suggested that COF-PM 2.5 resulted in inflammation, apoptosis, and cell damage in A549 cells, which might be modulated via the activation of MAPK/NF-кB/STAT1 pathway.

  6. Intestinal Parasite Co-infection among Pulmonary Tuberculosis Cases without Human Immunodeficiency Virus Infection in a Rural County in China

    PubMed Central

    Li, Xin-Xu; Chen, Jia-Xu; Wang, Li-Xia; Tian, Li-Guang; Zhang, Yu-Ping; Dong, Shuang-Pin; Hu, Xue-Guang; Liu, Jian; Wang, Feng-Feng; Wang, Yue; Yin, Xiao-Mei; He, Li-Jun; Yan, Qiu-Ye; Zhang, Hong-Wei; Xu, Bian-Li; Zhou, Xiao-Nong

    2014-01-01

    Epidemiologic studies of co-infection with tuberculosis (TB) and intestinal parasites in humans have not been extensively investigated in China. A cross-section study was conducted in a rural county of Henan Province, China. Pulmonary TB (PTB) case-patients receiving treatment for infection with Mycobacterium tuberculosis and healthy controls matched for geographic area, age, and sex were surveyed by using questionnaires. Fecal and blood specimens were collected for detection of intestinal parasites, routine blood examination, and infection with human immunodeficiency virus. The chi-square test was used for univariate analysis and multivariate logistic regression models were used to adjust for potential confounding factors. A total of 369 persons with PTB and 366 healthy controls were included; all participants were negative for human immunodeficiency virus. The overall prevalence of intestinal parasites in persons with PTB was 14.9%, including intestinal protozoa (7.9%) and helminthes (7.6%). The infection spectrum of intestinal parasites was Entamoeba spp. (1.4%), Blastocystis hominis (6.2%), Trichomonas hominis (0.3%), Clonorchis sinensis (0.3%), Ascaris lumbricoides (0.5%), Trichuris trichiura (2.2%), and hookworm (4.6%). The prevalence of intestinal parasites showed no significant difference between persons with PTB and healthy controls after adjusting for potential confounding factors. There was no factor that affected infection rates for intestinal parasites between the two groups. Infection with intestinal parasites of persons with PTB was associated with female sex (adjusted odds ratio [AOR] = 2.05, 95% confidence interval [CI] = 1.01–4.17), body mass index ≤ 19 (AOR = 3.02, 95% CI = 1.47–6.20), and anemia (AOR = 2.43, 95% CI = 1.17–5.03). Infection of healthy controls was only associated with an annual labor time in farmlands > 2 months (AOR = 4.50, 95% CI = 2.03–10.00). In addition, there was no significant trend between rates of infection with

  7. Low-dose carbon-based nanoparticle-induced effects in A549 lung cells determined by biospectroscopy are associated with increases in genomic methylation.

    PubMed

    Li, Junyi; Tian, Meiping; Cui, Li; Dwyer, John; Fullwood, Nigel J; Shen, Heqing; Martin, Francis L

    2016-02-02

    Nanotechnology has introduced many manufactured carbon-based nanoparticles (CNPs) into our environment, generating a debate into their risks and benefits. Numerous nanotoxicology investigations have been carried, and nanoparticle-induced toxic effects have been reported. However, there remain gaps in our knowledge, primarily regarding mechanism. Herein, we assessed the global alterations induced by CNPs in A549 lung cells using biospectroscopy techniques, including attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy and surface-enhanced Raman spectroscopy (SERS). A549 cells were treated with fullerene (C60), long or short multi-walled carbon nanotubes, or single-walled carbon nanotubes at concentrations of 0.1 mg/L, 0.01 mg/L and 0.001 mg/L. Exposed cells were then analysed by ATR-FTIR spectroscopy and SERS. Spectra were pre-processed via computational analysis, and information on biochemical alterations in exposed cells were identified. Additionally, global DNA methylation levels in cells exposed to CNPs at 0.1 mg/L were determined using HPLC-MS and genetic regulators (for DNA methylation) were checked by quantitative real-time RT-PCR. It was found that CNPs exert marked effects in A549 cells and also contribute to increases in global DNA methylation. For the first time, this study highlights that real-world levels of nanoparticles can alter the methylome of exposed cells; this could have enormous implications for their regulatory assessment.

  8. Heat-Modified Citrus Pectin Induces Apoptosis-Like Cell Death and Autophagy in HepG2 and A549 Cancer Cells

    PubMed Central

    Leclere, Lionel; Fransolet, Maude; Cote, Francois; Cambier, Pierre; Arnould, Thierry; Van Cutsem, Pierre; Michiels, Carine

    2015-01-01

    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. PMID:25794149

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

    PubMed

    Leclere, Lionel; Fransolet, Maude; Cote, Francois; Cambier, Pierre; Arnould, Thierry; Van Cutsem, Pierre; Michiels, Carine

    2015-01-01

    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.

  10. Silica nanoparticles and lead acetate co-exposure triggered synergistic cytotoxicity in A549 cells through potentiation of mitochondria-dependent apoptosis induction.

    PubMed

    Lu, Chun-Feng; Li, Li-Zhong; Zhou, Wei; Zhao, Jun; Wang, Yi-Mei; Peng, Shuang-Qing

    2017-06-01

    The adverse effects of PM2.5 are the results of combined toxicities of finer particles and their adsorbed toxic pollutants. Nevertheless,