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Sample records for lung fibroblasts isolated

  1. Caspase 3 activity in isolated fetal rat lung fibroblasts and rat periodontal ligament fibroblasts: cigarette smoke-induced alterations

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    James Elliot Scott

    2016-03-01

    Full Text Available Background Cigarette smoking is the leading cause of preventable death in the world. It has been implicated in the pathogenesis of pulmonary, oral and systemic diseases. Smoking during pregnancy is clearly a risk factor for the developing fetus and may be a major cause of infant mortality. Moreover, the oral cavity is the first site of exposure to cigarette smoke and may be a possible source for the spread of toxins to other organs of the body. Fibroblasts in general are morphologically heterogeneous connective tissue cells with diverse functions. Apoptosis or programmed cell death is a crucial process during embryogenesis and for the maintenance of homeostasis throughout life. Deregulation of apoptosis has been implicated in abnormal lung development in the fetus and disease progression in adults. Caspases, are proteases which belong to the family of cysteine aspartic acid proteases and are the key components for the downstream amplification of intra-cellular apoptotic signals. Of the 14 caspases known, caspase-3 is the key executioner of apoptosis. Fetal rat lung fibroblasts but not PDL viability is reduced by exposure to CSE. In addition Caspase 3 activity is elevated after CSE exposure in fetal lung fibroblasts but not in PDLs. Expression of caspase 3 is induced in CSE exposed lung fibroblasts but not in PDLs. Caspase 3 was localized to the cytoplasm in both cell types.

  2. Heparan sulfate proteoglycan from the extracellular matrix of human lung fibroblasts. Isolation, purification, and core protein characterization

    International Nuclear Information System (INIS)

    Heremans, A.; Cassiman, J.J.; Van den Berghe, H.; David, G.

    1988-01-01

    Confluent cultured human lung fibroblasts were labeled with 35SO4(2-). After 48 h of labeling, the pericellular matrix was prepared by Triton X-100 and deoxycholate extraction of the monolayers. Heparan sulfate proteoglycan (HSPG) accounted for nearly 80% of the total matrix [35S]proteoglycans. After solubilization in 6 M guanidinium HCl and cesium chloride density gradient centrifugation, the majority (78%) of these [35S] HSPG equilibrated at an average buoyant density of 1.35 g/ml. This major HSPG fraction was purified by ion-exchange chromatography on Mono Q and by gel filtration on Sepharose CL-4B, and further characterized by gel electrophoresis and immunoblotting. Intact [35S]HSPG eluted with Kav 0.1 from Sepharose CL-4B, whereas the protein-free [35S]heparan sulfate chains, obtained by alkaline borohydride treatment of the proteoglycan fractions, eluted with Kav 0.45 (Mr approximately 72,000). When analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography, core (protein) preparations, obtained by heparitinase digestion of 125I-labeled HSPG fractions, yielded one major labeled band with apparent molecular mass of approximately 300 kDa. Reduction with beta-mercaptoethanol slightly increased the apparent Mr of the labeled band, suggesting a single polypeptide structure and the presence of intrachain disulfide bonds. Immunoadsorption experiments and immunostaining of electrophoretically separated heparitinase-digested core proteins with monoclonal antibodies raised against matrix and cell surface-associated HSPG suggested that the major matrix-associated HSPG of cultured human lung fibroblasts is distinct from the HSPG that are anchored in the membranes of these cells. Binding studies suggested that this matrix HSPG interacts with several matrix components, both through its glycosaminoglycan chains and through its heparitinase-resistant core. (Abstract Truncated)

  3. Lung Fibroblasts, Aging, and Idiopathic Pulmonary Fibrosis.

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    Pardo, Annie; Selman, Moisés

    2016-12-01

    Idiopathic pulmonary fibrosis (IPF) is an aging-associated, progressive, and irreversible lung disease of unknown etiology, elusive pathogenesis, and very limited therapeutic options. The hallmarks of IPF are aberrant activation of alveolar epithelial cells and accumulation of fibroblasts and myofibroblasts along with excessive production of extracellular matrix. The linkage of aging with this disorder is uncertain, but a number of changes associated with aging, including telomere attrition, cell senescence, and mitochondrial dysfunction, have been revealed in IPF lungs. Also, aging seems to confer a profibrotic phenotype upon fibroblasts and to increase the severity of the fibrogenic response in non-IPF fibrotic lung disorders. Better knowledge of the pathophysiological mechanisms linking aging to IPF will advance understanding of its pathogenesis and may provide new therapeutic windows to treatment of this devastating disease.

  4. Effects of titanium dioxide nanoparticles isolated from confectionery products on the metabolic stress pathway in human lung fibroblast cells.

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    Periasamy, Vaiyapuri Subbarayan; Athinarayanan, Jegan; Al-Hadi, Ahmed M; Juhaimi, Fahad Al; Alshatwi, Ali A

    2015-04-01

    Titanium dioxide (TiO2) is a common additive in many foods, pigments, personal care products, and other consumer products used in daily life. Despite the widespread use of nanoscale TiO2 and composites of nanoscale TiO2 in the food industry, there is a serious lack of awareness of the toxicity of TiO2 nanoparticles (NPs) among consumers and manufacturers. There is an urgent need for toxicological studies of TiO2 NPs. TiO2 food additives separated from marketed foods were characterized by transmission electron microscopy. In addition, the effects of TiO2 NPs on metabolic stress in WI-38 cells were analyzed. Cell viability, total ROS, mitochondrial transmembrane potential (ΔψM), cell cycle, and metabolism-related gene expression were analyzed. The results indicate that TiO2 NPs have a significant concentration-dependent toxic effect in lung cells. The ΔψM, the intracellular ROS level, and the stages of the WI-38 cell cycle were altered by increasing TiO2 concentrations after exposure for 24 and 48 h relative to the control. Cytochrome P450 1A, GSTM3, and glutathione S-transferase A4 upregulation in response to the TiO2 NPs was observed. These findings suggest that the toxicity of TiO2 from confectionery products in WI-38 cells may be mediated through an increase in oxidative stress. The results of this study clearly demonstrate the nanotoxicological effects of TiO2 on WI-38 cells and will be useful for nanotoxicological indexing.

  5. CCN5 overexpression inhibits profibrotic phenotypes via the PI3K/Akt signaling pathway in lung fibroblasts isolated from patients with idiopathic pulmonary fibrosis and in an in vivo model of lung fibrosis.

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    Zhang, Lin; Li, Yingna; Liang, Chunlian; Yang, Weilin

    2014-02-01

    Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive interstitial lung disease with unknown etiology and undefined treatment modality. Fibroblasts are regarded as the major cell type that mediates the onset and progression of lung fibrosis by secreting large amounts of extracellular matrix (ECM) proteins, such as connective tissue growth factor (CTGF/CCN2). Current knowledge confers a crucial role of CCN2 in lung fibrosis. CCN5, another member of the CCN family, has been suggested to play an inhibitory role in some fibrotic diseases, such as cardiac fibrosis. However, the role of CCN5 in the process of IPF remains unknown. In the present study, using western blot analysis, we demonstrate that CCN2 is highly expressed in fibroblasts derived from IPF tissue, but is only slightly expressed in normal human lung fibroblasts. However, CCN5 was weakly expressed in all the above cells. qRT-PCR revealed that transforming growth factor (TGF)-β1 stimulation increased CCN2 expression in the IPF-derived cultures of primary human lung fibroblasts (PIFs) in a time- and concentration-dependent manner, but only slightly affected the expression of CCN5. The overexpression of CCN5 induced by the transfection of PIFs with recombinant plasmid did not affect cell viability, proliferation and apoptosis; however, it significantly suppressed the expression of CCN2, α-smooth muscle actin (α-SMA) and collagen type I. The TGF-β1-induced upregulation of the phosphorylation of Akt was reversed by CCN5 overexpression. Our results also demonstrated that adenovirus-mediated CCN5 overexpression in a mouse model of bleomycin-induced IPF significantly decreased the hydroxyproline content in the lungs, as well as TGF-β1 expression in bronchoalveolar lavage fluid. Taken together, our data demonstrate that CCN5 exerts an inhibitory effect on the fibrotic phenotypes of pulmonary fibroblasts in vitro and in vivo, and as such may be a promising target for the treatment of IPF.

  6. Different proliferative capacity of lung fibroblasts obtained from control subjects and patients with emphysema

    NARCIS (Netherlands)

    Noordhoek, JA; Postma, DS; Chong, LL; Vos, JTWM; Kauffman, HF; Timens, W; van Straaten, JFM

    2003-01-01

    To characterize the possible role of a dysregulated proliferative capacity of pulmonary fibroblasts in insufficient tissue repair in lungs from patients with pulmonary emphysema, the authors undertook in vitro proliferative studies with pulmonary fibroblasts obtained from lung tissue of patients

  7. Anti-fibrotic effects of theophylline on lung fibroblasts

    International Nuclear Information System (INIS)

    Yano, Yukihiro; Yoshida, Mitsuhiro; Hoshino, Shigenori; Inoue, Koji; Kida, Hiroshi; Yanagita, Masahiko; Takimoto, Takayuki; Hirata, Haruhiko; Kijima, Takashi; Kumagai, Toru; Osaki, Tadashi; Tachibana, Isao; Kawase, Ichiro

    2006-01-01

    Theophylline has been used in the management of bronchial asthma and chronic obstructive pulmonary disease for over 50 years. It has not only a bronchodilating effect, but also an anti-inflammatory one conducive to the inhibition of airway remodeling, including subepithelial fibrosis. To date however, whether theophylline has a direct inhibitory effect on airway fibrosis has not been established. To clarify this question, we examined whether theophylline affected the function of lung fibroblasts. Theophylline suppressed TGF-β-induced type I collagen (COL1) mRNA expression in lung fibroblasts and also inhibited fibroblast proliferation stimulated by FBS and TGF-β-induced α-SMA protein. A cAMP analog also inhibited TGF-β-induced COL1 mRNA expression in lung fibroblasts. A PKA inhibitor reduced the inhibitory effect of theophylline on TGF-β-induced COL1 mRNA expression. These results indicate that theophylline exerts anti-fibrotic effects, at least partly, through the cAMP-PKA pathway

  8. Radiation-Induced Differentiation in Human Lung Fibroblast

    International Nuclear Information System (INIS)

    Park, Sa-Rah; Ahn, Ji-Yeon; Han, Young-Soo; Shim, Jie-Young; Yun, Yeon-Sook; Song, Jie-Young

    2007-01-01

    One of the most common tumors in many countries is lung cancer and patients with lung cancer may take radiotherapy. Although radiotherapy may have its own advantages, it can also induce serious problems such as acute radiation pneumonitis and pulmonary fibrosis. Pulmonary fibrosis is characterized by excessive production of α-SMA and accumulation of extracellular matrix (ECM) such as collagen and fibronectin. There has been a great amount of research about fibrosis but the exact mechanism causing the reaction is not elucidated especially in radiation-induced fibrosis. Until now it has been known that several factors such as transforming growth factor (TGF-β), tumor necrosis factor (TNF), interleukin (IL)-1, IL-6, platelet-derived growth factor (PDGF) and fibroblast growth factor (FGF) are related to fibrosis. Among them TGF-β with Smad signaling is known to be the main stream and other signaling molecules such as MAPK, ERK and JNK (3) also participates in the process. In addition to those above factors, it is thought that more diverse and complicate mechanisms may involve in the radiationinduced fibrosis. Therefore, to investigate the underlying mechanisms in radiation induced fibrosis, first of all, we confirmed whether radiation induces trans differentiation in human normal lung fibroblasts. Here, we suggest that not only TGF-β but also radiation can induce trans differentiation in human lung fibroblast WI-38 and IMR-90

  9. Rosmarinic acid potentiates carnosic acid induced apoptosis in lung fibroblasts.

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    Sana Bahri

    Full Text Available Pulmonary fibrosis is characterized by over-population and excessive activation of fibroblasts and myofibroblasts disrupting normal lung structure and functioning. Rosemary extract rich in carnosic acid (CA and rosmarinic acid (RA was reported to cure bleomycin-(BLM-induced pulmonary fibrosis. We demonstrate that CA decreased human lung fibroblast (HLF viability with IC50 value of 17.13±1.06 μM, while RA had no cytotoxic effect. In the presence of 50 μM of RA, dose-response for CA shifted to IC50 value of 11.70±1.46 μM, indicating synergic action. TGFβ-transformed HLF, rat lung fibroblasts and L929 cells presented similar sensitivity to CA and CA+RA (20μM+100μM, respectively treatment. Rat alveolar epithelial cells died only under CA+RA treatment, while A549 cells were not affected. Annexin V staining and DNA quantification suggested that HLF are arrested in G0/G1 cell cycle phase and undergo apoptosis. CA caused sustained activation of phospho-Akt and phospho-p38 expression and inhibition of p21 protein.Addition of RA potentiated these effects, while RA added alone had no action.Only triple combination of inhibitors (MAPK-p38, pan-caspase, PI3K/Akt/autophagy partially attenuated apoptosis; this suggests that cytotoxicity of CA+RA treatment has a complex mechanism involving several parallel signaling pathways. The in vivo antifibrotic effect of CA and RA was compared with that of Vitamine-E in BLM-induced fibrosis model in rats. We found comparable reduction in fibrosis score by CA, RA and CA+RA, attenuation of collagen deposition and normalization of oxidative stress markers. In conclusion, antifibrotic effect of CA+RA is due to synergistic pro-apoptotic action on lung fibroblasts and myofibroblasts.

  10. Metabolic cooperation between co-cultured lung cancer cells and lung fibroblasts.

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    Koukourakis, Michael I; Kalamida, Dimitra; Mitrakas, Achilleas G; Liousia, Maria; Pouliliou, Stamatia; Sivridis, Efthimios; Giatromanolaki, Alexandra

    2017-11-01

    Cooperation of cancer cells with stromal cells, such as cancer-associated fibroblasts (CAFs), has been revealed as a mechanism sustaining cancer cell survival and growth. In the current study, we focus on the metabolic interactions of MRC5 lung fibroblasts with lung cancer cells (A549 and H1299) using co-culture experiments and studying changes of the metabolic protein expression profile and of their growth and migration abilities. Using western blotting, confocal microscopy and RT-PCR, we observed that in co-cultures MRC5 respond by upregulating pyruvate dehydrogenase (PDH) and the monocarboxylate transporter MCT1. In contrast, cancer cells increase the expression of glucose transporters (GLUT1), LDH5, PDH kinase and the levels of phosphorylated/inactivated pPDH. H1299 cells growing in the same culture medium with fibroblasts exhibit a 'metastasis-like' phenomenon by forming nests within the fibroblast area. LDH5 and pPDH were drastically upregulated in these nests. The growth rate of both MRC5 and cancer cells increased in co-cultures. Suppression of LDHA or PDK1 in cancer cells abrogates the stimulatory signal from cancer cells to fibroblasts. Incubation of MRC5 fibroblasts with lactate resulted in an increase of LDHB and of PDH expression. Silencing of PDH gene in fibroblasts, or silencing of PDK1 or LDHA gene in tumor cells, impedes cancer cell's migration ability. Overall, a metabolic cooperation between lung cancer cells and fibroblasts has been confirmed in the context of direct Warburg effect, thus the fibroblasts reinforce aerobic metabolism to support the intensified anaerobic glycolytic pathways exploited by cancer cells.

  11. [Isolation, purification and primary culture of adult mouse cardiac fibroblasts].

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    Li, Rujun; Gong, Kaizheng; Zhang, Zhengang

    2017-01-01

    Objective To establish a method for primary culture of adult mouse cardiac fibroblasts. Methods Myocardial tissues from adult mice were digested with 1 g/L trypsin and 0.8 g/L collagenase IV by oscillating water bath for a short time repeatedly. Cardiac fibroblasts and myocardial cells were isolated with differential adhesion method. Immunofluorescence staining was used to assess the purity of cardiac fibroblasts. The cell morphology was observed under an inverted phase contrast microscope. The proliferation of cardiac fibroblasts was analyzed by growth curve and CCK-8 assay. The Smad2/3 phosphorylation induced by TGF-β1 was detected by Western blotting. Results After 90 minutes of differential adhesion, adherent fibroblasts formed spherical cell mass and after 3 days, cells were spindle-shaped and proliferated rapidly. Cells were confluent after 5 days and the growth curve presented nearly "S" shape. The positive expression rate of vimentin was 95%. CCK-8 assay showed that the optimal cell proliferating activity was found from day 3 to day 5. The level of phosphorylated Smad2/3 obviously increased at the second passage induced by TGF-β1. Conclusion This method is economical and stable to isolate cardiac fibroblasts with high activity and high purity from adult mice.

  12. Age related changes in steroid receptors on cultured lung fibroblasts

    International Nuclear Information System (INIS)

    Barile, F.A.; Bienkowski, R.S.

    1986-01-01

    The number of high affinity glucocorticoid receptors (Ro) on human fetal lung fibroblasts decreases as the cells age in vitro, and it has been suggested that these cell systems may be useful models of age-related changes in vivo. They examined the relation between change in Ro with in vitro aging and donor age. Confluent monolayers of lung fibroblasts at various population doubling levels (PDL), were incubated with ( 3 H)-dexamethasone (( 3 H)Dex) either alone or with excess (.01 mM) Dex. Specific binding was calculated as the difference between radioactivity in cells incubated with and without unlabeled Dex; Scatchard plots were used to analyze the data. Ro, measured as fmol ( 3 H)Dex/10 6 cells, for two lines of human fetal cells (HFL-1 and MRC-5) decreased with increasing age in vitro. However, human newborn (CRL-1485) and adult (CCL-201) cells and fetal rabbit cells (FAB-290), showed increases in Ro with continuous passage. For each cell line, the affinity constant (K/sub d/) did not change significantly with passage. They conclude that the direction of changes in steroid receptor levels on cells aging in vitro is influenced by donor age and species. Caution should be used in applying results obtained from model systems to aging organisms

  13. Role of epithelial-mesenchymal transition (EMT) and fibroblast function in cerium oxide nanoparticles-induced lung fibrosis

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    Ma, Jane; Bishoff, Bridget; Mercer, R.R.; Barger, Mark; Schwegler-Berry, Diane; Castranova, Vincent

    2017-01-01

    The emission of cerium oxide nanoparticles (CeO 2 ) from diesel engines, using cerium compounds as a catalyst to lower the diesel exhaust particles, is a health concern. We have previously shown that CeO 2 induced pulmonary inflammation and lung fibrosis. The objective of the present study was to investigate the modification of fibroblast function and the role of epithelial-mesenchymal transition (EMT) in CeO 2 -induced fibrosis. Male Sprague-Dawley rats were exposed to CeO 2 (0.15 to 7 mg/kg) by a single intratracheal instillation and sacrificed at various times post-exposure. The results show that at 28 days after CeO 2 (3.5 mg/kg) exposure, lung fibrosis was evidenced by increased soluble collagen in bronchoalveolar lavage fluid, elevated hydroxyproline content in lung tissues, and enhanced sirius red staining for collagen in the lung tissue. Lung fibroblasts and alveolar type II (ATII) cells isolated from CeO 2 -exposed rats at 28 days post-exposure demonstrated decreasing proliferation rate when compare to the controls. CeO 2 exposure was cytotoxic and altered cell function as demonstrated by fibroblast apoptosis and aggregation, and ATII cell hypertrophy and hyperplasia with increased surfactant. The presence of stress fibers, expressed as α-smooth muscle actin (SMA), in CeO 2 -exposed fibroblasts and ATII cells was significantly increased compared to the control. Immunohistofluorescence analysis demonstrated co-localization of TGF-β or α-SMA with prosurfactant protein C (SPC)-stained ATII cells. These results demonstrate that CeO 2 exposure affects fibroblast function and induces EMT in ATII cells that play a role in lung fibrosis. These findings suggest potential adverse health effects in response to CeO 2 nanoparticle exposure. - Highlights: • CeO 2 exposure induced lung fibrosis. • CeO 2 were detected in lung tissue, alveolar type II (ATII) cells and fibroblasts. • CeO 2 caused ATII cell hypertrophy and hyperplasia and altered fibroblast function

  14. Role of epithelial-mesenchymal transition (EMT) and fibroblast function in cerium oxide nanoparticles-induced lung fibrosis

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    Ma, Jane [Health Effects Laboratory Division, NIOSH, Morgantown, WV (United States); Bishoff, Bridget [Mylan Pharmaceuticals, Morganntown, WV (United States); Mercer, R.R.; Barger, Mark; Schwegler-Berry, Diane [Health Effects Laboratory Division, NIOSH, Morgantown, WV (United States); Castranova, Vincent, E-mail: vcastran@hsc.wvu.edu [School of Pharmacy, West Virginia University, Morgantown, WV (United States)

    2017-05-15

    The emission of cerium oxide nanoparticles (CeO{sub 2}) from diesel engines, using cerium compounds as a catalyst to lower the diesel exhaust particles, is a health concern. We have previously shown that CeO{sub 2} induced pulmonary inflammation and lung fibrosis. The objective of the present study was to investigate the modification of fibroblast function and the role of epithelial-mesenchymal transition (EMT) in CeO{sub 2}-induced fibrosis. Male Sprague-Dawley rats were exposed to CeO{sub 2} (0.15 to 7 mg/kg) by a single intratracheal instillation and sacrificed at various times post-exposure. The results show that at 28 days after CeO{sub 2} (3.5 mg/kg) exposure, lung fibrosis was evidenced by increased soluble collagen in bronchoalveolar lavage fluid, elevated hydroxyproline content in lung tissues, and enhanced sirius red staining for collagen in the lung tissue. Lung fibroblasts and alveolar type II (ATII) cells isolated from CeO{sub 2}-exposed rats at 28 days post-exposure demonstrated decreasing proliferation rate when compare to the controls. CeO{sub 2} exposure was cytotoxic and altered cell function as demonstrated by fibroblast apoptosis and aggregation, and ATII cell hypertrophy and hyperplasia with increased surfactant. The presence of stress fibers, expressed as α-smooth muscle actin (SMA), in CeO{sub 2}-exposed fibroblasts and ATII cells was significantly increased compared to the control. Immunohistofluorescence analysis demonstrated co-localization of TGF-β or α-SMA with prosurfactant protein C (SPC)-stained ATII cells. These results demonstrate that CeO{sub 2} exposure affects fibroblast function and induces EMT in ATII cells that play a role in lung fibrosis. These findings suggest potential adverse health effects in response to CeO{sub 2} nanoparticle exposure. - Highlights: • CeO{sub 2} exposure induced lung fibrosis. • CeO{sub 2} were detected in lung tissue, alveolar type II (ATII) cells and fibroblasts. • CeO{sub 2} caused ATII

  15. Annexin A1 is elevated in patients with COPD and affects lung fibroblast function

    Directory of Open Access Journals (Sweden)

    Lai TW

    2018-02-01

    Full Text Available Tianwen Lai,1,* Yanyu Li,1,* Zongjiong Mai,2 Xiaoxia Wen,1 Yingying Lv,1 Zhanqing Xie,3 Quanchao Lv,1 Min Chen,1 Dong Wu,1 Bin Wu1 1Department of Respiratory and Critical Care Medicine, 2Department of Oncology, 3Department of Thoracic Surgery, The Affiliated Hospital of Guangdong Medical University, Zhanjiang, People’s Republic of China *These authors contributed equally to this work Purpose: Fibrosis in peripheral airways is responsible for airflow limitation in chronic obstructive pulmonary disease (COPD. Annexin A1 modulates several key biological events during inflammation. However, little is known about its role in airway fibrosis in COPD. We investigated whether levels of Annexin A1 were upregulated in patients with COPD, and whether it promoted airway fibrosis.Methods: We quantified serum Annexin A1 levels in never-smokers (n=12, smokers without COPD (n=11, and smokers with COPD (n=22. Correlations between Annexin A1 expression and clinical indicators (eg, lung function were assessed. In vitro, human bronchial epithelial (HBE cells were exposed to cigarette smoke extract (CSE and Annexin A1 expression was assessed. Primary human lung fibroblasts were isolated from patients with COPD and effects of Annexin A1 on fibrotic deposition of lung fibroblasts were evaluated.Results: Serum Annexin A1 was significantly higher in patients with Global Initiative for Chronic Obstructive Lung Disease (GOLD guidelines stage III or IV than in those with GOLD stages I or II (12.8±0.8 ng/mL versus 9.8±0.7 ng/mL; p=0.016. Annexin A1 expression was negatively associated with airflow obstruction (forced expiratory volume in one second % predicted; r=−0.72, p<0.001. In vitro, Annexin A1 was significantly increased in CSE-exposed HBE cells in a time- and concentration-dependent manner. Annexin A1 promoted lung fibroblasts proliferation, migration, differentiation, and collagen deposition via the ERK1/2 and p38 mitogen-activated protein kinase pathways

  16. ROS Mediates Radiation-Induced Differentiation in Human Lung Fibroblast

    International Nuclear Information System (INIS)

    Park, Sa Rah; Ahn, Ji Yeon; Kim, Mi Hyeung; Lim, Min Jin; Yun, Yeon Sook; Song, Jie Young

    2009-01-01

    One of the most common tumors worldwide is lung cancer and the number of patients with lung cancer received radiotherapy is increasing rapidly. Although radiotherapy may have lots of advantages, it can also induce serious adverse effects such as acute radiation pneumonitis and pulmonary fibrosis. Pulmonary fibrosis is characterized by excessive production of smooth muscle actin-alpha (a-SMA) and accumulation of extracellular matrix (ECM) such as collagen and fibronectin. There has been a great amount of research about fibrosis but the exact mechanism causing the reaction is not elucidated especially in radiation-induced fibrosis. Until now it has been known that several factors such as transforming growth factor (TGF-b), tumor necrosis factor (TNF), IL-6, platelet-derived growth factor (PDGF) and reactive oxygen species are related to fibrosis. It is also reported that reactive oxygen species (ROS) can be induced by radiation and can act as a second messenger in various signaling pathways. Therefore we focused on the role of ROS in radiation induced fibrosis. Here, we suggest that irradiation generate ROS mainly through NOX4, result in differentiation of lung fibroblast into myofibroblast

  17. Dihydrotestosterone Potentiates EGF-Induced ERK Activation by Inducing SRC in Fetal Lung Fibroblasts

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    Smith, Susan M.; Murray, Sandy; Pham, Lucia D.; Minoo, Parviz; Nielsen, Heber C.

    2014-01-01

    Lung maturation is regulated by interactions between mesenchymal and epithelial cells, and is delayed by androgens. Fibroblast–Type II cell communications are dependent on extracellular signal-regulated kinases (ERK) 1/2 activation by the ErbB receptor ligands epidermal growth factor (EGF), transforming growth factor (TGF)-α, and neuregulin (Nrg). In other tissues, dihydrotestosterone (DHT) has been shown to activate SRC by a novel nontranscriptional mechanism, which phosphorylates EGF receptors to potentiate EGF-induced ERK1/2 activation. This study sought to determine if DHT potentiates EGFR signaling by a nontranscriptional mechanism. Embryonic day (E)17 fetal lung cells were isolated from dams treated with or without DHT since E12. Cells were exposed to 30 ng/ml DHT for periods of 30 minutes to 3 days before being stimulated with 100 ng/ml EGF, TGF-α, or Nrg for up to 30 minutes. Lysates were immunoblotted for ErbB and SRC pathway signaling intermediates. DHT increased ERK1/2 activation by EGF, TGF-α, and Nrg in fibroblasts and Type II cells. Characterization in fibroblasts showed that potentiation of the EGF pathway was significant after 60 minutes of DHT exposure and persisted in the presence of the translational inhibitor cycloheximide. SRC and EGF receptor phosphorylation was increased by DHT, as was EGF-induced SHC1 phosphorylation and subsequent association with GRB2. Finally, SRC silencing, SRC inhibition with PP2, and overexpression of a dominant-negative SRC each prevented DHT from increasing EGF-induced ERK1/2 phosphorylation. These results suggest that DHT activates SRC to potentiate the signaling pathway leading from the EGF receptor to ERK activation in primary fetal lung fibroblasts. PMID:24484548

  18. Effect of primarily cultured human lung cancer-associated fibroblasts on radiosensitivity of lung cancer cells

    International Nuclear Information System (INIS)

    Ji Xiaoqin; Ji Jiang; Chen Yongbing; Shan Fang; Lu Xueguan

    2014-01-01

    Objective: To investigate the effect of human lung cancer-associated fibroblasts (CAF) on the radiosensitivity of lung cancer cells when CAF is placed in direct contact co-culture with lung cancer cells. Methods: Human lung CAF was obtained from fresh human lung adenocarcinoma tissue specimens by primary culture and subculture and was then identified by immunofluorescence staining. The CAF was placed in direct contact co-culture with lung cancer A 549 and H 1299 cells, and the effects of CAF on the radiosensitivity of A 549 and H 1299 cells were evaluated by colony-forming assay. Results: The human lung CAF obtained by adherent culture could stably grow and proliferate, and it had specific expression of α-smooth muscle actin, vimentin, and fibroblast activation protein,but without expression of cytokeratin-18. The plating efficiency (PE, %) of A 549 cells at 0 Gy irradiation was (20.0 ± 3.9)% when cultured alone versus (32.3 ± 5.5)% when co-cultured with CAF (t=3.16, P<0.05), and the PE of H 1299 cells at 0 Gy irradiation was (20.6 ± 3.1)% when cultured alone versus (35.2 ± 2.3)% when co-cultured with CAF (t=6.55, P<0.05). The cell survival rate at 2 Gy irradiation (SF 2 ) of A 549 cells was 0.727 ±0.061 when cultured alone versus 0.782 ± 0.089 when co-cultured with CAF (t=0.88, P>0.05), and the SF 2 of H 1299 cells was 0.692 ±0.065 when cultured alone versus 0.782 ± 0.037 when co-cultured with CAF (t=2.08, P>0.05). The protection enhancement ratios of human lung CAF for A 549 cells and H 1299 cells were 1.29 and 1.25, respectively. Conclusions: Human lung CAF reduces the radiosensitivity of lung cancer cells when placed in direct contact co-culture with them, and the radioprotective effect may be attributed to CAF promoting the proliferation of lung cancer cells. (authors)

  19. C-type natriuretic peptide ameliorates pulmonary fibrosis by acting on lung fibroblasts in mice.

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    Kimura, Toru; Nojiri, Takashi; Hino, Jun; Hosoda, Hiroshi; Miura, Koichi; Shintani, Yasushi; Inoue, Masayoshi; Zenitani, Masahiro; Takabatake, Hiroyuki; Miyazato, Mikiya; Okumura, Meinoshin; Kangawa, Kenji

    2016-02-19

    Pulmonary fibrosis has high rates of mortality and morbidity; however, no effective pharmacological therapy has been established. C-type natriuretic peptide (CNP), a member of the natriuretic peptide family, selectively binds to the transmembrane guanylyl cyclase (GC)-B receptor and exerts anti-inflammatory and anti-fibrotic effects in various organs through vascular endothelial cells and fibroblasts that have a cell-surface GC-B receptor. Given the pathophysiological importance of fibroblast activation in pulmonary fibrosis, we hypothesized that the anti-fibrotic and anti-inflammatory effects of exogenous CNP against bleomycin (BLM)-induced pulmonary fibrosis were exerted in part by the effect of CNP on pulmonary fibroblasts. C57BL/6 mice were divided into two groups, CNP-treated (2.5 μg/kg/min) and vehicle, to evaluate BLM-induced (1 mg/kg) pulmonary fibrosis and inflammation. A periostin-CNP transgenic mouse model exhibiting CNP overexpression in fibroblasts was generated and examined for the anti-inflammatory and anti-fibrotic effects of CNP via fibroblasts in vivo. Additionally, we assessed CNP attenuation of TGF-β-induced differentiation into myofibroblasts by using immortalized human lung fibroblasts stably expressing GC-B receptors. Furthermore, to investigate whether CNP acts on human lung fibroblasts in a clinical setting, we obtained primary-cultured fibroblasts from surgically resected lungs of patients with lung cancer and analyzed levels of GC-B mRNA transcription. CNP reduced mRNA levels of the profibrotic cytokines interleukin (IL)-1β and IL-6, as well as collagen deposition and the fibrotic area in lungs of mice with bleomycin-induced pulmonary fibrosis. Furthermore, similar CNP effects were observed in transgenic mice exhibiting fibroblast-specific CNP overexpression. In cultured-lung fibroblasts, CNP treatment attenuated TGF-β-induced phosphorylation of Smad2 and increased mRNA and protein expression of α-smooth muscle actin and SM22

  20. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) promotes lung fibroblast proliferation, survival and differentiation to myofibroblasts.

    Science.gov (United States)

    Hasaneen, Nadia A; Cao, Jian; Pulkoski-Gross, Ashleigh; Zucker, Stanley; Foda, Hussein D

    2016-02-17

    Idiopathic pulmonary fibrosis (IPF) is a chronic progressively fatal disease. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) is a glycosylated transmembrane protein that induces the expression of some matrix metalloproteinase (MMP) in neighboring stromal cells through direct epithelial-stromal interactions. EMMPRIN is highly expressed in type II alveolar epithelial cells at the edges of the fibrotic areas in IPF lung sections. However, the exact role of EMMPRIN in IPF is unknown. To determine if EMMPRIN contributes to lung fibroblast proliferation, resistance to apoptosis, and differentiation to myofibroblasts, normal Human lung fibroblasts (NHLF) transiently transfected with either EMMPRIN/GFP or GFP were treated with TGF- β1 from 0 to 10 ng/ml for 48 h and examined for cell proliferation (thymidine incorporation), apoptosis (FACS analysis and Cell Death Detection ELISA assay), cell migration (Modified Boyden chamber) and differentiation to myofibroblasts using Western blot for α-smooth actin of cell lysates. The effect of EMMPRIN inhibition on NHLF proliferation, apoptosis, migration and differentiation to myofibroblasts after TGF- β1 treatment was examined using EMMPRIN blocking antibody. We examined the mechanism by which EMMPRIN induces its effects on fibroblasts by studying the β-catenin/canonical Wnt signaling pathway using Wnt luciferase reporter assays and Western blot for total and phosphorylated β-catenin. Human lung fibroblasts overexpressing EMMPRIN had a significant increase in cell proliferation and migration compared to control fibroblasts. Furthermore, EMMPRIN promoted lung fibroblasts resistance to apoptosis. Lung fibroblasts overexpressing EMMPRIN showed a significantly increased expression of α- smooth muscle actin, a marker of differentiation to myofibroblasts compared to control cells. TGF-β1 increased the expression of EMMPRIN in lung fibroblasts in a dose-dependent manner. Attenuation of EMMPRIN expression with the use of an

  1. Congestive heart failure effects on atrial fibroblast phenotype: differences between freshly-isolated and cultured cells.

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    Kristin Dawson

    Full Text Available Fibroblasts are important in the atrial fibrillation (AF substrate resulting from congestive heart failure (CHF. We previously noted changes in in vivo indices of fibroblast function in a CHF dog model, but could not detect changes in isolated cells. This study assessed CHF-induced changes in the phenotype of fibroblasts freshly isolated from control versus CHF dogs, and examined effects of cell culture on these differences.Left-atrial fibroblasts were isolated from control and CHF dogs (ventricular tachypacing 240 bpm × 2 weeks. Freshly-isolated fibroblasts were compared to fibroblasts in primary culture. Extracellular-matrix (ECM gene-expression was assessed by qPCR, protein by Western blot, fibroblast morphology with immunocytochemistry, and K(+-current with patch-clamp. Freshly-isolated CHF fibroblasts had increased expression-levels of collagen-1 (10-fold, collagen-3 (5-fold, and fibronectin-1 (3-fold vs. control, along with increased cell diameter (13.4 ± 0.4 µm vs control 8.4 ± 0.3 µm and cell spreading (shape factor 0.81 ± 0.02 vs. control 0.87 ± 0.02, consistent with an activated phenotype. Freshly-isolated control fibroblasts displayed robust tetraethylammonium (TEA-sensitive K(+-currents that were strongly downregulated in CHF. The TEA-sensitive K(+-current differences between control and CHF fibroblasts were attenuated after 2-day culture and eliminated after 7 days. Similarly, cell-culture eliminated the ECM protein-expression and shape differences between control and CHF fibroblasts.Freshly-isolated CHF and control atrial fibroblasts display distinct ECM-gene and morphological differences consistent with in vivo pathology. Culture for as little as 48 hours activates fibroblasts and obscures the effects of CHF. These results demonstrate potentially-important atrial-fibroblast phenotype changes in CHF and emphasize the need for caution in relating properties of cultured fibroblasts to in vivo systems.

  2. CD14-negative isolation enhances chondrogenesis in synovial fibroblasts.

    Science.gov (United States)

    Bilgen, Bahar; Ren, Yuexin; Pei, Ming; Aaron, Roy K; Ciombor, Deborah McK

    2009-11-01

    Synovial membrane has been shown to contain mesenchymal stem cells. We hypothesized that an enriched population of synovial fibroblasts would undergo chondrogenic differentiation and secrete cartilage extracellular matrix to a greater extent than would a mixed synovial cell population (MSCP). The optimum doses of transforming growth factor beta 1 (TGF-beta1) and insulin-like growth factor 1 (IGF-1) for chondrogenesis were investigated. CD14-negative isolation was used to obtain a porcine cell population enriched in type-B synovial fibroblasts (SFB) from an MSCP. The positive cell surface markers in SFB were CD90, CD44, and cadherin-11. SFB and MSCP were cultured in the presence of 20 ng/mL TGF-beta1 for 7 days, and SFB were demonstrated to have higher chondrogenic potential. Further dose-response studies were carried out using the SFB cells and several doses of TGF-beta1 (2, 10, 20, and 40 ng/mL) and/or IGF-1 (1, 10, 100, and 500 ng/mL) for 14 days. TGF-beta1 supplementation was essential for chondrogenesis and prevention of cell death, whereas IGF-1 did not have a significant effect on the SFB cell number or glycosaminoglycan production. This study demonstrates that the CD14-negative isolation yields an enhanced cell population SFB that is more potent than MSCP as a cell source for cartilage tissue engineering.

  3. Expression of WNT5A in Idiopathic Pulmonary Fibrosis and Its Control by TGF-β and WNT7B in Human Lung Fibroblasts.

    Science.gov (United States)

    Newman, Donna R; Sills, W Shane; Hanrahan, Katherine; Ziegler, Amanda; Tidd, Kathleen McGinnis; Cook, Elizabeth; Sannes, Philip L

    2016-02-01

    The wingless (Wnt) family of signaling ligands contributes significantly to lung development and is highly expressed in patients with usual interstitial pneumonia (UIP). We sought to define the cellular distribution of Wnt5A in the lung tissue of patients with idiopathic pulmonary fibrosis (IPF) and the signaling ligands that control its expression in human lung fibroblasts and IPF myofibroblasts. Tissue sections from 40 patients diagnosed with IPF or UIP were probed for the immunolocalization of Wnt5A. Further, isolated lung fibroblasts from normal or IPF human lungs, adenovirally transduced for the overexpression or silencing of Wnt7B or treated with TGF-β1 or its inhibitor, were analyzed for Wnt5A protein expression. Wnt5A was expressed in IPF lungs by airway and alveolar epithelium, smooth muscle cells, endothelium, and myofibroblasts of fibroblastic foci and throughout the interstitium. Forced overexpression of Wnt7B with or without TGF-β1 treatment significantly increased Wnt5A protein expression in normal human smooth muscle cells and fibroblasts but not in IPF myofibroblasts where Wnt5A was already highly expressed. The results demonstrate a wide distribution of Wnt5A expression in cells of the IPF lung and reveal that it is significantly increased by Wnt7B and TGF-β1, which, in combination, could represent key signaling pathways that modulate the pathogenesis of IPF. © 2016 The Histochemical Society.

  4. Identification of different subsets of lung cells using Raman microspectroscopy and whole cell nucleus isolation.

    Science.gov (United States)

    Pijanka, Jacek K; Stone, Nicholas; Rutter, Abigail V; Forsyth, Nicholas; Sockalingum, Ganesh D; Yang, Ying; Sulé-Suso, Josep

    2013-09-07

    Raman spectroscopy has been widely used to study its possible clinical application in cancer diagnosis. However, in order to make it into clinical practice, it is important that this technique is able not only to identify cancer cells from their normal counterparts, but also from the array of cells present in human tissues. To this purpose, we used Raman spectroscopy to assess whether this technique was able to differentiate not only between lung cancer cells and lung epithelial cells but also from lung fibroblasts. Furthermore, we studied whether the differences were due to cell lineage (epithelial versus fibroblast) or to different proliferative characteristics of cells, and where in the cell compartment these differences might reside. To answer these questions we studied cell cytoplasm, cell nucleus and isolated whole cell nuclei. Our data suggests that Raman spectroscopy can differentiate between lung cancer, lung epithelial cells and lung fibroblasts. More important, it can also differentiate between 2 cells from the same lineage (fibroblast) but with one of them rendered immortal and with an increased proliferative activity. Finally, it seems that the main spectral differences reside in the cell nucleus and that the study of isolated nuclei strengthens the differences between cells.

  5. KL-6, a human MUC1 mucin, promotes proliferation and survival of lung fibroblasts

    International Nuclear Information System (INIS)

    Ohshimo, Shinichiro; Yokoyama, Akihito; Hattori, Noboru; Ishikawa, Nobuhisa; Hirasawa, Yutaka; Kohno, Nobuoki

    2005-01-01

    The serum level of KL-6, a MUC1 mucin, is a clinically useful marker for various interstitial lung diseases. Previous studies demonstrated that KL-6 promotes chemotaxis of human fibroblasts. However, the pathophysiological role of KL-6 remains poorly understood. Here, we further investigate the functional aspects of KL-6 in proliferation and apoptosis of lung fibroblasts. KL-6 accelerated the proliferation and inhibited the apoptosis of all human lung fibroblasts examined. An anti-KL-6 monoclonal antibody counteracted both of these effects induced by KL-6 on human lung fibroblasts. The pro-fibroproliferative and anti-apoptotic effects of KL-6 are greater than and additive to those of the maximum effective concentrations of platelet-derived growth factor, basic fibroblast growth factor, and transforming growth factor-β. These findings indicate that increased levels of KL-6 in the epithelial lining fluid may stimulate fibrotic processes in interstitial lung diseases and raise the possibility of applying an anti-KL-6 antibody to treat interstitial lung diseases

  6. Sirtuin 7 is decreased in pulmonary fibrosis and regulates the fibrotic phenotype of lung fibroblasts.

    Science.gov (United States)

    Wyman, Anne E; Noor, Zahid; Fishelevich, Rita; Lockatell, Virginia; Shah, Nirav G; Todd, Nevins W; Atamas, Sergei P

    2017-06-01

    Pulmonary fibrosis is a severe condition with no cure and limited therapeutic options. A better understanding of its pathophysiology is needed. Recent studies have suggested that pulmonary fibrosis may be driven by accelerated aging-related mechanisms. Sirtuins (SIRTs), particularly SIRT1, SIRT3, and SIRT6, are well-known mediators of aging; however, limited data exist on the contribution of sirtuins to lung fibrosis. We assessed the mRNA and protein levels of all seven known sirtuins in primary lung fibroblasts from patients with idiopathic pulmonary fibrosis (IPF) and systemic sclerosis-associated interstitial lung disease (SSc-ILD) in comparison with lung fibroblasts from healthy controls. These unbiased tests revealed a tendency for all sirtuins to be expressed at lower levels in fibroblasts from patients compared with controls, but the greatest decrease was observed with SIRT7. Similarly, SIRT7 was decreased in lung tissues of bleomycin-challenged mice. Inhibition of SIRT7 with siRNA in cultured lung fibroblasts resulted in an increase in collagen and α-smooth muscle actin (α-SMA). Reciprocally, overexpression of SIRT7 resulted in lower basal and TGF-β-induced levels of COL1A1, COL1A2, COL3A1, and α-SMA mRNAs, as well as collagen and α-SMA proteins. Induced changes in SIRT7 had no effect on endogenous TGF-β mRNA levels or latent TGF-β activation, but overexpression of SIRT7 reduced the levels of Smad3 mRNA and protein. In conclusion, the decline in SIRT7 in lung fibroblasts has a profibrotic effect, which is mediated by changes in Smad3 levels.

  7. Lung Beractant Increases Free Cytosolic Levels of Ca2+ in Human Lung Fibroblasts.

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    Alejandro Guzmán-Silva

    Full Text Available Beractant, a natural surfactant, induces an antifibrogenic phenotype and apoptosis in normal human lung fibroblasts (NHLF. As intracellular Ca2+ signalling has been related to programmed cell death, we aimed to assess the effect of beractant on intracellular Ca2+ concentration ([Ca2+]i in NHLF in vitro. Cultured NHLF were loaded with Fura-2 AM (3 μM and Ca2+ signals were recorded by microfluorimetric techniques. Beractant causes a concentration-dependent increase in [Ca2+]i with a EC50 of 0.82 μg/ml. The application of beractant, at a concentration of 500 μg/ml, which has been shown to exert an apoptotic effect in human fibroblasts, elicited different patterns of Ca2+ signals in NHLF: a a single Ca2+ spike which could be followed by b Ca2+ oscillations, c a sustained Ca2+ plateau or d a sustained plateau overlapped by Ca2+ oscillations. The amplitude and pattern of Ca2+ transients evoked by beractant were dependent on the resting [Ca2+]i. Pharmacological manipulation revealed that beractant activates a Ca2+ signal through Ca2+ release from intracellular stores mediated by phospholipase Cβ (PLCβ, Ca2+ release from inositol 1,4,5-trisphosphate receptors (IP3Rs and Ca2+ influx via a store-operated pathway. Moreover, beractant-induced Ca2+ release was abolished by preventing membrane depolarization upon removal of extracellular Na+ and Ca2+. Finally, the inhibition of store-operated channels prevented beractant-induced NHLF apoptosis and downregulation of α1(I procollagen expression. Therefore, beractant utilizes SOCE to exert its pro-apoptotic and antifibrinogenic effect on NHLF.

  8. Advanced Research of Fibroblast Growth Factor Receptor 
in Non-small Cell Lung Cancer

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    Dan PU

    2013-11-01

    Full Text Available Lung cancer is severely threatening human health. In recent years, the treatment for lung adenocarcinoma has made a great progress, targeted therapy has been widely applied in clinic, and benefits amount of patients. However, in squamous cell lung cancer, the incidence of epidermal growth factor receptor (EGFR gene mutant and ALK fusion gene are low,and targeted therapy like Tarceva and crizotinib, can hardly work. Since the fibroblast growth factors (fibroblast growth factor, FGF pathway is considered to be related to tumor cell proliferation, metastasis and angiogenesis, more and more researches proved the amplification of fibroblast growth factor receptor (FGFR in squamous cell lung cancer. Experiments in vivo and in vitro found that blocking FGF pathway could reduce the proliferation of tumor cells and inhibit metastasis. The FGF pathway might be a new target for treatment of squamous cell lung cancer. This article reviews the effect of FGFR in tumorigenesis,as well as the prospect as a therapeutic target in non-small cell lung cancer.

  9. Teaching basic lung isolation skills on human anatomy simulator: attainment and retention of lung isolation skills.

    Science.gov (United States)

    Latif, Rana K; VanHorne, Edgar M; Kandadai, Sunitha Kanchi; Bautista, Alexander F; Neamtu, Aurel; Wadhwa, Anupama; Carter, Mary B; Ziegler, Craig H; Memon, Mohammed Faisal; Akça, Ozan

    2016-01-20

    Lung isolation skills, such as correct insertion of double lumen endobronchial tube and bronchial blocker, are essential in anesthesia training; however, how to teach novices these skills is underexplored. Our aims were to determine (1) if novices can be trained to a basic proficiency level of lung isolation skills, (2) whether video-didactic and simulation-based trainings are comparable in teaching lung isolation basic skills, and (3) whether novice learners' lung isolation skills decay over time without practice. First, five board certified anesthesiologist with experience of more than 100 successful lung isolations were tested on Human Airway Anatomy Simulator (HAAS) to establish Expert proficiency skill level. Thirty senior medical students, who were naive to bronchoscopy and lung isolation techniques (Novice) were randomized to video-didactic and simulation-based trainings to learn lung isolation skills. Before and after training, Novices' performances were scored for correct placement using pass/fail scoring and a 5-point Global Rating Scale (GRS); and time of insertion was recorded. Fourteen novices were retested 2 months later to assess skill decay. Experts' and novices' double lumen endobronchial tube and bronchial blocker passing rates showed similar success rates after training (P >0.99). There were no differences between the video-didactic and simulation-based methods. Novices' time of insertion decayed within 2 months without practice. Novices could be trained to basic skill proficiency level of lung isolation. Video-didactic and simulation-based methods we utilized were found equally successful in training novices for lung isolation skills. Acquired skills partially decayed without practice.

  10. Adiponectin attenuates lung fibroblasts activation and pulmonary fibrosis induced by paraquat.

    Directory of Open Access Journals (Sweden)

    Rong Yao

    Full Text Available Pulmonary fibrosis is one of the most common complications of paraquat (PQ poisoning, which demands for more effective therapies. Accumulating evidence suggests adiponectin (APN may be a promising therapy against fibrotic diseases. In the current study, we determine whether the exogenous globular APN isoform protects against pulmonary fibrosis in PQ-treated mice and human lung fibroblasts, and dissect the responsible underlying mechanisms. BALB/C mice were divided into control group, PQ group, PQ + low-dose APN group, and PQ + high-dose APN group. Mice were sacrificed 3, 7, 14, and 21 days after PQ treatment. We compared pulmonary histopathological changes among different groups on the basis of fibrosis scores, TGF-β1, CTGF and α-SMA pulmonary content via Western blot and real-time quantitative fluorescence-PCR (RT-PCR. Blood levels of MMP-9 and TIMP-1 were determined by ELISA. Human lung fibroblasts WI-38 were divided into control group, PQ group, APN group, and APN receptor (AdipoR 1 small-interfering RNA (siRNA group. Fibroblasts were collected 24, 48, and 72 hours after PQ exposure for assay. Cell viability and apoptosis were determined via Kit-8 (CCK-8 and fluorescein Annexin V-FITC/PI double labeling. The protein and mRNA expression level of collagen type III, AdipoR1, and AdipoR2 were measured by Western blot and RT-PCR. APN treatment significantly decreased the lung fibrosis scores, protein and mRNA expression of pulmonary TGF-β1, CTGF and α-SMA content, and blood MMP-9 and TIMP-1 in a dose-dependent manner (p<0.05. Pretreatment with APN significantly attenuated the reduced cell viability and up-regulated collagen type III expression induced by PQ in lung fibroblasts, (p<0.05. APN pretreatment up-regulated AdipoR1, but not AdipoR2, expression in WI-38 fibroblasts. AdipoR1 siRNA abrogated APN-mediated protective effects in PQ-exposed fibroblasts. Taken together, our data suggests APN protects against PQ-induced pulmonary fibrosis in a

  11. Ca2+ influx and ATP release mediated by mechanical stretch in human lung fibroblasts

    International Nuclear Information System (INIS)

    Murata, Naohiko; Ito, Satoru; Furuya, Kishio; Takahara, Norihiro; Naruse, Keiji; Aso, Hiromichi; Kondo, Masashi; Sokabe, Masahiro; Hasegawa, Yoshinori

    2014-01-01

    Highlights: • Uniaxial stretching activates Ca 2+ signaling in human lung fibroblasts. • Stretch-induced intracellular Ca 2+ elevation is mainly via Ca 2+ influx. • Mechanical strain enhances ATP release from fibroblasts. • Stretch-induced Ca 2+ influx is not mediated by released ATP or actin cytoskeleton. - Abstract: One cause of progressive pulmonary fibrosis is dysregulated wound healing after lung inflammation or damage in patients with idiopathic pulmonary fibrosis and severe acute respiratory distress syndrome. The mechanical forces are considered to regulate pulmonary fibrosis via activation of lung fibroblasts. In this study, the effects of mechanical stretch on the intracellular Ca 2+ concentration ([Ca 2+ ] i ) and ATP release were investigated in primary human lung fibroblasts. Uniaxial stretch (10–30% in strain) was applied to fibroblasts cultured in a silicone chamber coated with type I collagen using a stretching apparatus. Following stretching and subsequent unloading, [Ca 2+ ] i transiently increased in a strain-dependent manner. Hypotonic stress, which causes plasma membrane stretching, also transiently increased the [Ca 2+ ] i . The stretch-induced [Ca 2+ ] i elevation was attenuated in Ca 2+ -free solution. In contrast, the increase of [Ca 2+ ] i by a 20% stretch was not inhibited by the inhibitor of stretch-activated channels GsMTx-4, Gd 3+ , ruthenium red, or cytochalasin D. Cyclic stretching induced significant ATP releases from fibroblasts. However, the stretch-induced [Ca 2+ ] i elevation was not inhibited by ATP diphosphohydrolase apyrase or a purinergic receptor antagonist suramin. Taken together, mechanical stretch induces Ca 2+ influx independently of conventional stretch-sensitive ion channels, the actin cytoskeleton, and released ATP

  12. Ca{sup 2+} influx and ATP release mediated by mechanical stretch in human lung fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Murata, Naohiko [Department of Respiratory Medicine, Nagoya University Graduate School of Medicine, Nagoya 466-8550 (Japan); Ito, Satoru, E-mail: itori@med.nagoya-u.ac.jp [Department of Respiratory Medicine, Nagoya University Graduate School of Medicine, Nagoya 466-8550 (Japan); Furuya, Kishio [Mechanobiology Laboratory, Nagoya University Graduate School of Medicine, Nagoya 466-8550 (Japan); Takahara, Norihiro [Department of Respiratory Medicine, Nagoya University Graduate School of Medicine, Nagoya 466-8550 (Japan); Naruse, Keiji [Department of Cardiovascular Physiology, Okayama University Graduate School of Medicine, Okayama 700-8558 (Japan); Aso, Hiromichi; Kondo, Masashi [Department of Respiratory Medicine, Nagoya University Graduate School of Medicine, Nagoya 466-8550 (Japan); Sokabe, Masahiro [Mechanobiology Laboratory, Nagoya University Graduate School of Medicine, Nagoya 466-8550 (Japan); Hasegawa, Yoshinori [Department of Respiratory Medicine, Nagoya University Graduate School of Medicine, Nagoya 466-8550 (Japan)

    2014-10-10

    Highlights: • Uniaxial stretching activates Ca{sup 2+} signaling in human lung fibroblasts. • Stretch-induced intracellular Ca{sup 2+} elevation is mainly via Ca{sup 2+} influx. • Mechanical strain enhances ATP release from fibroblasts. • Stretch-induced Ca{sup 2+} influx is not mediated by released ATP or actin cytoskeleton. - Abstract: One cause of progressive pulmonary fibrosis is dysregulated wound healing after lung inflammation or damage in patients with idiopathic pulmonary fibrosis and severe acute respiratory distress syndrome. The mechanical forces are considered to regulate pulmonary fibrosis via activation of lung fibroblasts. In this study, the effects of mechanical stretch on the intracellular Ca{sup 2+} concentration ([Ca{sup 2+}]{sub i}) and ATP release were investigated in primary human lung fibroblasts. Uniaxial stretch (10–30% in strain) was applied to fibroblasts cultured in a silicone chamber coated with type I collagen using a stretching apparatus. Following stretching and subsequent unloading, [Ca{sup 2+}]{sub i} transiently increased in a strain-dependent manner. Hypotonic stress, which causes plasma membrane stretching, also transiently increased the [Ca{sup 2+}]{sub i}. The stretch-induced [Ca{sup 2+}]{sub i} elevation was attenuated in Ca{sup 2+}-free solution. In contrast, the increase of [Ca{sup 2+}]{sub i} by a 20% stretch was not inhibited by the inhibitor of stretch-activated channels GsMTx-4, Gd{sup 3+}, ruthenium red, or cytochalasin D. Cyclic stretching induced significant ATP releases from fibroblasts. However, the stretch-induced [Ca{sup 2+}]{sub i} elevation was not inhibited by ATP diphosphohydrolase apyrase or a purinergic receptor antagonist suramin. Taken together, mechanical stretch induces Ca{sup 2+} influx independently of conventional stretch-sensitive ion channels, the actin cytoskeleton, and released ATP.

  13. Adiponectin attenuates lung fibroblasts activation and pulmonary fibrosis induced by paraquat.

    Science.gov (United States)

    Yao, Rong; Cao, Yu; He, Ya-rong; Lau, Wayne Bond; Zeng, Zhi; Liang, Zong-an

    2015-01-01

    Pulmonary fibrosis is one of the most common complications of paraquat (PQ) poisoning, which demands for more effective therapies. Accumulating evidence suggests adiponectin (APN) may be a promising therapy against fibrotic diseases. In the current study, we determine whether the exogenous globular APN isoform protects against pulmonary fibrosis in PQ-treated mice and human lung fibroblasts, and dissect the responsible underlying mechanisms. BALB/C mice were divided into control group, PQ group, PQ + low-dose APN group, and PQ + high-dose APN group. Mice were sacrificed 3, 7, 14, and 21 days after PQ treatment. We compared pulmonary histopathological changes among different groups on the basis of fibrosis scores, TGF-β1, CTGF and α-SMA pulmonary content via Western blot and real-time quantitative fluorescence-PCR (RT-PCR). Blood levels of MMP-9 and TIMP-1 were determined by ELISA. Human lung fibroblasts WI-38 were divided into control group, PQ group, APN group, and APN receptor (AdipoR) 1 small-interfering RNA (siRNA) group. Fibroblasts were collected 24, 48, and 72 hours after PQ exposure for assay. Cell viability and apoptosis were determined via Kit-8 (CCK-8) and fluorescein Annexin V-FITC/PI double labeling. The protein and mRNA expression level of collagen type III, AdipoR1, and AdipoR2 were measured by Western blot and RT-PCR. APN treatment significantly decreased the lung fibrosis scores, protein and mRNA expression of pulmonary TGF-β1, CTGF and α-SMA content, and blood MMP-9 and TIMP-1 in a dose-dependent manner (ppulmonary fibrosis in a dose-dependent manner, via suppression of lung fibroblast activation. Functional AdipoR1 are expressed by human WI-38 lung fibroblasts, suggesting potential future clinical applicability of APN against pulmonary fibrosis.

  14. The effects of collagen-rich extracellular matrix on the intracellular delivery of glycol chitosan nanoparticles in human lung fibroblasts.

    Science.gov (United States)

    Yhee, Ji Young; Yoon, Hong Yeol; Kim, Hyunjoon; Jeon, Sangmin; Hergert, Polla; Im, Jintaek; Panyam, Jayanth; Kim, Kwangmeyung; Nho, Richard Seonghun

    2017-01-01

    Recent progress in nanomedicine has shown a strong possibility of targeted therapy for obstinate chronic lung diseases including idiopathic pulmonary fibrosis (IPF). IPF is a fatal lung disease characterized by persistent fibrotic fibroblasts in response to type I collagen-rich extracellular matrix. As a pathological microenvironment is important in understanding the biological behavior of nanoparticles, in vitro cellular uptake of glycol chitosan nanoparticles (CNPs) in human lung fibroblasts was comparatively studied in the presence or absence of type I collagen matrix. Primary human lung fibroblasts from non-IPF and IPF patients (n=6/group) showed significantly increased cellular uptake of CNPs (>33.6-78.1 times) when they were cultured on collagen matrix. To elucidate the underlying mechanism of enhanced cellular delivery of CNPs in lung fibroblasts on collagen, cells were pretreated with chlorpromazine, genistein, and amiloride to inhibit clathrin-mediated endocytosis, caveolae-mediated endocytosis, and macropinocytosis, respectively. Amiloride pretreatment remarkably reduced the cellular uptake of CNPs, suggesting that lung fibroblasts mainly utilize the macropinocytosis-dependent mechanism when interacted with collagen. In addition, the internalization of CNPs was predominantly suppressed by a phosphoinositide 3-kinase (PI3K) inhibitor in IPF fibroblasts, indicating that enhanced PI3K activity associated with late-stage macropinocytosis can be particularly important for the enhanced cellular delivery of CNPs in IPF fibroblasts. Our study strongly supports the concept that a pathological microenvironment which surrounds lung fibroblasts has a significant impact on the intracellular delivery of nanoparticles. Based on the property of enhanced intracellular delivery of CNPs when fibroblasts are made to interact with a collagen-rich matrix, we suggest that CNPs may have great potential as a drug-carrier system for targeting fibrotic lung fibroblasts.

  15. Fibroblasts are in a position to provide directional information to migrating neutrophils during pneumonia in rabbit lungs.

    Science.gov (United States)

    Behzad, A R; Chu, F; Walker, D C

    1996-05-01

    Previous findings have shown that pulmonary fibroblasts are associated with preexisting holes in the endothelial and epithelial basal laminae through which neutrophils appear to enter and leave the interstitium as they migrate from capillaries to alveoli. To determine their role in neutrophil migration, fibroblast organization within the interstitium was assessed by transmission electron microscope observations of serial-sectioned rabbit lung tissue. Interstitial fibroblasts were found to physically interconnect the endothelial basal lamina holes to epithelial basal lamina holes. Morphometric assessment of rabbit lung tissue instilled with Streptococcus pneumoniae revealed that approximately 70% of the surface area density of migrating neutrophils is in close contact (15 nm or less) with interstitial fibroblasts and extracellular matrix elements (30 and 40%, respectively). Although migrating neutrophils were close enough to adhere to both fibroblasts and extracellular elements, the interstitial fibroblasts are organized in a manner that would allow them to provide directional information to the neutrophils. A model illustrating this process is proposed.

  16. Hydroxychloroquine susceptibility determination of Coxiella burnetii in human embryonic lung (HEL) fibroblast cells.

    Science.gov (United States)

    Angelakis, Emmanouil; Khalil, Jacques Bou; Le Bideau, Marion; Perreal, Celine; La Scola, Bernard; Raoult, Didier

    2017-07-01

    Coxiella burnetii, the causative agent of Q fever, survives and replicates in the acidic environment of monocytes/macrophages; hydroxychloroquine, through alkalinisation of the acidic vacuoles, is critical for the management of Q fever. In this study, a collection of C. burnetii strains isolated from human samples was tested to evaluate the in vitro minimum inhibitory concentrations (MICs) of doxycycline and hydroxychloroquine. Serial two-fold dilutions of doxycycline (0.25-8 mg/L) and hydroxychloroquine (0.25-4 mg/L) were added to C. burnetii-infected human embryonic lung (HEL) fibroblast cells after 48 h of incubation, in duplicate. DNA was detected by C. burnetii-specific semi-quantitative PCR with primers and probes designed for amplification of the IS1111 and IS30A spacers. A total of 29 C. burnetii isolates obtained from 29 patients were tested. Doxycycline MICs ranged from 0.25 mg/L to 0.5 mg/L and hydroxychloroquine MICs from 0.25 mg/L to >4 mg/L. Four C. burnetii stains had hydroxychloroquine MICs ≤ 1 mg/L. The concentration of hydroxychloroquine was associated with a significant decrease in C. burnetii DNA copies in HEL cells based on linear regression analysis (P= 0.01). Recommended serum concentrations of hydroxychloroquine significantly reduced the growth of C. burnetii. Moreover, some C. burnetii strains presented hydroxychloroquine MICs below the recommended serum concentrations, indicating that, for these cases, hydroxychloroquine treatment alone may even be effective. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

  17. PTEN gene and phosphorylation of Akt protein expression in the LPS-induced lung fibroblast

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    Mao-lin HUANG

    2014-09-01

    Full Text Available Objective: To investigate PTEN gene expression and the Akt phosphorylation of protein expression in the LPS-induced lung fibroblast, to initially reveal the relation between PTEN gene and the Akt phosphorylated proteins to LPS-induced lung fibroblast proliferation mechanism. Methods: BrdU experiments was performed to evaluate the LPS-induced lung fibroblast proliferation,  RT-PCR and Western Blot analysis were used to analyze the PTEN gene expression and Western blot was performed to analyze Akt phosphorylated protein expression. Results: PTEN mRNA level of the experimental group were significantly lower than the control group (P<0.05 with LPS simulation for 24h and 72h , and there were no significant difference between the experimental group and control group the experimental group and control group (P>0.05 . PTEN protein expression levels of the experimental group were significantly lower than the control group (P<0.05 , at 72h, and PTEN mRNA levels had no significant differences between these of the experimental and control group at 6h,12h and 24h(p>0.05. Phosphorylation Akt protein level (relative to total Akt protein was significantly higer than the control group (P<0.05 at 24h and 72h, and phosphorylation Akt protein levels had no significant differences between these of the experimental and control group at 6h and 12h (P>0.05 .Conclusion: PTEN gene and phosphorylation Akt protein involve in LPS-induced lung fibroblast proliferation signal transduction pathway.

  18. Innate Immune Cytokines, Fibroblast Phenotypes, and Regulation of Extracellular Matrix in Lung.

    Science.gov (United States)

    Richards, Carl D

    2017-02-01

    Chronic inflammation can be caused by adaptive immune responses in autoimmune and allergic conditions, driven by a T lymphocyte subset balance (TH1, TH2, Th17, Th22, and/or Treg) and skewed cellular profiles in an antigen-specific manner. However, several chronic inflammatory diseases have no clearly defined adaptive immune mechanisms that drive chronicity. These conditions include those that affect the lung such as nonatopic asthma or idiopathic pulmonary fibrosis comprising significant health problems. The remodeling of extracellular matrix (ECM) causes organ dysfunction, and it is largely generated by fibroblasts as the major cell controlling net ECM. As such, these are potential targets of treatment approaches in the context of ECM pathology. Fibroblast phenotypes contribute to ECM and inflammatory cell accumulation, and they are integrated into chronic disease mechanisms including cancer. Evidence suggests that innate cytokine responses may be critical in nonallergic/nonautoimmune disease, and they enable environmental agent exposure mechanisms that are independent of adaptive immunity. Innate immune cytokines derived from macrophage subsets (M1/M2) and innate lymphoid cell (ILC) subsets can directly regulate fibroblast function. We also suggest that STAT3-activating gp130 cytokines can sensitize fibroblasts to the innate cytokine milieu to drive phenotypes and exacerbate existing adaptive responses. Here, we review evidence exploring innate cytokine regulation of fibroblast behavior.

  19. Cigarette smoke exposure inhibits extracellular MMP-2 (gelatinase A activity in human lung fibroblasts

    Directory of Open Access Journals (Sweden)

    Cappello Francesco

    2007-03-01

    Full Text Available Abstract Background Exposure to cigarette smoke is considered a major risk factor for the development of lung diseases, since its causative role has been assessed in the induction and maintenance of an inflamed state in the airways. Lung fibroblasts can contribute to these processes, due to their ability to produce proinflammatory chemotactic molecules and extracellular matrix remodelling proteinases. Among proteolytic enzymes, gelatinases A and B have been studied for their role in tissue breakdown and mobilisation of matrix-derived signalling molecules. Multiple reports linked gelatinase deregulation and overexpression to the development of inflammatory chronic lung diseases such as COPD. Methods In this study we aimed to determine variations in the gelatinolytic pattern of human lung fibroblasts (HFL-1 cell line exposed to cigarette smoke extract (CSE. Gelatinolytic activity levels were determined by using gelatin zymography for the in-gel detection of the enzymes (proenzyme and activated forms, and the subsequent semi-quantitative densitometric evaluation of lytic bands. Expression of gelatinases was evaluated also by RT-PCR, zymography of the cell lysates and by western blotting. Results CSE exposure at the doses used (1–10% did not exert any significant cytotoxic effects on fibroblasts. Zymographic analysis showed that CSE exposure resulted in a linear decrease of the activity of gelatinase A. Control experiments allowed excluding a direct inhibitory effect of CSE on gelatinases. Zymography of cell lysates confirmed the expression of MMP-2 in all conditions. Semi-quantitative evaluation of mRNA expression allowed assessing a reduced transcription of the enzyme, as well as an increase in the expression of TIMP-2. Statistical analyses showed that the decrease of MMP-2 activity in conditioned media reached the statistical significance (p = 0.0031 for 24 h and p = 0.0012 for 48 h, while correlation analysis showed that this result was

  20. Cellular Glycolysis and The Differential Survival of Lung Fibroblast and Lung Carcinoma Cell Lines.

    Science.gov (United States)

    Farah, Ibrahim O

    2016-04-01

    Tumor growth and abnormal cell survival were shown to be associated with a number of cellular metabolic abnormalities revealed by impaired oral glucose tolerance, depressed lipoprotein lipase activity leading to hypertriglyceridemia, and changes in amino acid profile as evidenced by increased plasma free tryptophan levels in patients with breast, lung, colon, stomach, and other cancers from various origins. The above findings seem to relate to or indicate a shift to non-oxidative metabolic pathways in cancer. In contrast to normal cells, cancer cells may lose the ability to utilize aerobic respiration due to either defective mitochondria or hypoxia within the tumor microenvironments. Glucose was shown to be the major energy source in cancer cells where it utilizes aerobic /anaerobic glycolysis with the resultant lactic acid formation. The role of energetic modulations and use of glycolytic inhibitors on cancer/normal cell survival is not clearly established in the literature. We hypothesize that natural intermediates of glycolysis and the citric acid cycle will differentially and negatively impact the cancer phenotype in contrast to their no effects on the normal cell phenotype. Therefore, the purpose of this study was to evaluate six potential glycolytic modulators namely, Pyruvic acid, oxalic acid, Zn acetate, sodium citrate, fructose diphosphate (FDP) and sodium bicarbonate at μM concentrations on growing A549 (lung cancer) and MRC-5 (normal; human lung fibroblast) cell lines with the objective of determining their influence on visual impact, cell metabolic activity, cell viability and end-point cell survival. Exposed and non-exposed cells were tested with phase-contrast micro-scanning, survival/death and metabolic activity trends through MTT-assays, as well as death end-point determinations by testing re-growth on complete media and T4 cellometer counts. Results showed that oxalic acid and Zn acetate both influenced the pH of the medium and resulted in

  1. Fibroblast Activation Protein (FAP) Accelerates Collagen Degradation and Clearance from Lungs in Mice*

    Science.gov (United States)

    Fan, Ming-Hui; Zhu, Qiang; Li, Hui-Hua; Ra, Hyun-Jeong; Majumdar, Sonali; Gulick, Dexter L.; Jerome, Jacob A.; Madsen, Daniel H.; Christofidou-Solomidou, Melpo; Speicher, David W.; Bachovchin, William W.; Feghali-Bostwick, Carol; Puré, Ellen

    2016-01-01

    Idiopathic pulmonary fibrosis is a disease characterized by progressive, unrelenting lung scarring, with death from respiratory failure within 2–4 years unless lung transplantation is performed. New effective therapies are clearly needed. Fibroblast activation protein (FAP) is a cell surface-associated serine protease up-regulated in the lungs of patients with idiopathic pulmonary fibrosis as well as in wound healing and cancer. We postulate that FAP is not only a marker of disease but influences the development of pulmonary fibrosis after lung injury. In two different models of pulmonary fibrosis, intratracheal bleomycin instillation and thoracic irradiation, we find increased mortality and increased lung fibrosis in FAP-deficient mice compared with wild-type mice. Lung extracellular matrix analysis reveals accumulation of intermediate-sized collagen fragments in FAP-deficient mouse lungs, consistent with in vitro studies showing that FAP mediates ordered proteolytic processing of matrix metalloproteinase (MMP)-derived collagen cleavage products. FAP-mediated collagen processing leads to increased collagen internalization without altering expression of the endocytic collagen receptor, Endo180. Pharmacologic FAP inhibition decreases collagen internalization as expected. Conversely, restoration of FAP expression in the lungs of FAP-deficient mice decreases lung hydroxyproline content after intratracheal bleomycin to levels comparable with that of wild-type controls. Our findings indicate that FAP participates directly, in concert with MMPs, in collagen catabolism and clearance and is an important factor in resolving scar after injury and restoring lung homeostasis. Our study identifies FAP as a novel endogenous regulator of fibrosis and is the first to show FAP's protective effects in the lung. PMID:26663085

  2. Inhibition of fibroblast growth factor receptor 3-dependent lung adenocarcinoma with a human monoclonal antibody

    Directory of Open Access Journals (Sweden)

    Yongjun Yin

    2016-05-01

    Full Text Available Activating mutations in fibroblast growth factor receptor 3 (FGFR3 have been identified in multiple types of human cancer and in congenital birth defects. In human lung cancer, fibroblast growth factor 9 (FGF9, a high-affinity ligand for FGFR3, is overexpressed in 10% of primary resected non-small cell lung cancer (NSCLC specimens. Furthermore, in a mouse model where FGF9 can be induced in lung epithelial cells, epithelial proliferation and ensuing tumorigenesis is dependent on FGFR3. To develop new customized therapies for cancers that are dependent on FGFR3 activation, we have used this mouse model to evaluate a human monoclonal antibody (D11 with specificity for the extracellular ligand-binding domain of FGFR3, that recognizes both human and mouse forms of the receptor. Here, we show that D11 effectively inhibits signaling through FGFR3 in vitro, inhibits the growth of FGFR3-dependent FGF9-induced lung adenocarcinoma in mice, and reduces tumor-associated morbidity. Given the potency of FGF9 in this mouse model and the absolute requirement for signaling through FGFR3, this study validates the D11 antibody as a potentially useful and effective reagent for treating human cancers or other pathologies that are dependent on activation of FGFR3.

  3. RhoA signaling modulates cyclin D1 expression in human lung fibroblasts; implications for idiopathic pulmonary fibrosis

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    Hoban PR

    2006-06-01

    Full Text Available Abstract Background Idiopathic Pulmonary Fibrosis (IPF is a debilitating disease characterized by exaggerated extracellular matrix deposition and aggressive lung structural remodeling. Disease pathogenesis is driven by fibroblastic foci formation, consequent on growth factor overexpression and myofibroblast proliferation. We have previously shown that both CTGF overexpression and myofibroblast formation in IPF cell lines are dependent on RhoA signaling. As RhoA-mediated regulation is also involved in cell cycle progression, we hypothesise that this pathway is key to lung fibroblast turnover through modulation of cyclin D1 kinetic expression. Methods Cyclin D1 expression was compared in primary IPF patient-derived fibroblasts and equivalent normal control cells. Quantitative real time PCR was employed to examine relative expression levels of cyclin D1 mRNA; protein expression was confirmed by western blotting. Effects of Rho signaling were investigated using transient transfection of constitutively active and dominant negative RhoA constructs as well as pharmacological inhibitors. Cellular proliferation of lung fibroblasts was determined by BrdU incorporation ELISA. To further explore RhoA regulation of cyclin D1 in lung fibroblasts and associated cell cycle progression, an established Rho inhibitor, Simvastatin, was incorporated in our studies. Results Cyclin D1 expression was upregulated in IPF compared to normal lung fibroblasts under exponential growth conditions (p Conclusion These findings report for the first time that cyclin D1 expression is deregulated in IPF through a RhoA dependent mechanism that influences lung fibroblast proliferation. This potentially unravels new molecular targets for future anti-IPF strategies; accordingly, Simvastatin inhibition of Rho-mediated cyclin D1 expression in IPF fibroblasts merits further exploitation.

  4. Aluminum is More Cytotoxic than Lunar Dust in Human Skin and Lung Fibroblasts

    Science.gov (United States)

    Hammond, D.; Shehata, T.; Hammond, D.; Shehata, T.; Wise, J.P.; Martino, J; Wise, J.P.; Wise, J.P.

    2009-01-01

    NASA plans to build a permanent space station on the moon to explore its surface. The surface of the moon is covered in lunar dust, which consists of fine particles that contain silicon, aluminum and titanium, among others. Because this will be a manned base, the potential toxicity of this dust has to be studied. Also, toxicity standards for potential exposure have to be set. To properly address the potential toxicity of lunar dust we need to understand the toxicity of its individual components, as well as their combined effects. In order to study this we compared NASA simulant JSC-1AVF (volcanic ash particles), that simulates the dust found on the moon, to aluminum, the 3rd most abundant component in lunar dust. We tested the cytotoxicity of both compounds on human lung and skin fibroblasts (WTHBF-6 and BJhTERT cell lines, respectively). Aluminum oxide was more cytotoxic than lunar dust to both cell lines. In human lung fibroblasts 5, 10 and 50 g/sq cm of aluminum oxide induced 85%, 61% and 30% relative survival, respectively. For human skin fibroblasts the same concentrations induced 58%, 41% and 58% relative survival. Lunar dust was also cytotoxic to both cell lines, but its effects were seen at higher concentrations: 50, 100, 200 and 400 g/sq cm of lunar dust induced a 69%, 46%, 35% and 30% relative survival in the skin cells and 53%, 16%, 8% and 2% on the lung cells. Overall, for both compounds, lung cells were more sensitive than skin cells. This work was supported by a NASA EPSCoR grant through the Maine Space Grant Consortium (JPW), the Maine Center for Toxicology and Environmental Health., a Fulbright Grant (JM) and a Delta Kappa Gamma Society International World Fellowship (JM).

  5. Knockdown of versican 1 blocks cigarette-induced loss of insoluble elastin in human lung fibroblasts.

    Science.gov (United States)

    Xu, Lu-lu; Lu, Yun-tao; Zhang, Jing; Wu, Lian; Merrilees, Mervyn J; Qu, Jie-ming

    2015-08-15

    COPD lung is characterized by loss of alveolar elastic fibers and an increase in the chondroitin sulfate (CS) matrix proteoglycan versican V1 (V1). V1 is a known inhibitor of elastic fiber deposition and this study investigates the effects of knockdown of V1, and add-back of CS, on CCL-210 lung fibroblasts treated with cigarette smoke extract (CSE) as a model for COPD. CSE inhibited fibroblast proliferation, viability, tropoelastin synthesis, and elastin deposition, and increased V1 synthesis and secretion. V1 siRNA decreased V1 and constituent CS, did not affect tropoelastin production, but blocked the CSE-induced loss in insoluble elastin. Exogenous CS reduced insoluble elastin, even in the presence of V1 siRNA. These findings confirm that V1 and CS impair the assembly of tropoelastin monomers into insoluble fibers, and further demonstrate that specific knockdown of V1 alleviates the impaired assembly of elastin seen in cultures of pulmonary fibroblasts exposed to CSE, indicating a regulatory role for this protein in the pathophysiology of COPD. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Lewis lung carcinoma progression is facilitated by TIG-3 fibroblast cells.

    Science.gov (United States)

    Yamauchi, Yoshikane; Izumi, Yotaro; Asakura, Keisuke; Kawai, Kenji; Wakui, Masatoshi; Ohmura, Mitsuyo; Suematsu, Makoto; Nomori, Hiroaki

    2013-09-01

    The interactions of tumor cells with stromal fibroblasts influence tumor biology, but the exact mechanisms involved are still unclear. In the present study, we evaluated the effects of a human lung fibroblast cell line, TIG-3, on Lewis lung carcinoma (LLC) cells both in vitro and in vivo. LLC and TIG-3 cells were co-cultured/co-implanted in vitro and in vivo. Cell invasion was assayed. Local tumor growth, as well as lung metastasis, were evaluated after subcutaneous cell co-implantation into NOD/SCID/γ-null (NOG) mice. LLC, and TIG-3 cells were pre-treated with either SB431542, a small molecule TGF-β receptor antagonist, or siRNA for transforming growth factor (TGF)-β before co-culture or co-implantation, and the effects of pre-treatments were compared both in cell culture and in mice. Subcutaneous LLC tumor growth (L group) in NOG mice was significantly increased by co-implantation of TIG-3 cells (L+T group) at four weeks. The number of macroscopic lung metastases was also significantly increased in the L+T group in comparison to the L group. In vitro cell invasion was significantly increased in the L+T group in comparison to the L group. In vitro expression of phosphorylated-SMAD3 was significantly increased in the L+T group in comparison to the L group. Furthermore, pre-treatment with either SB431542 or siRNA for TGF-β reduced the invasiveness both in culture and in mice. This study suggested that in vitro as well as in vivo progression of LLC was facilitated by co-culture/co-implantation with TIG-3 cells, and that this process was at least in part dependent on TGF-β-mediated interactions.

  7. The cytotoxicity and genotoxicity of soluble and particulate cobalt in human lung fibroblast cells

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Leah J.; Holmes, Amie L. [Wise Laboratory of Environmental and Genetic Toxicology, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Maine Center for Environmental Toxicology and Health, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Department of Applied Medical Science, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Kandpal, Sanjeev Kumar; Mason, Michael D. [Department of Chemical and Biological Engineering, University of Maine, Orono, ME (United States); Zheng, Tongzhang [Department of Environmental Health Sciences, Yale School of Public Health, New Haven, CT (United States); Wise, John Pierce, E-mail: John.Wise@usm.maine.edu [Wise Laboratory of Environmental and Genetic Toxicology, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Maine Center for Environmental Toxicology and Health, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States); Department of Applied Medical Science, University of Southern Maine, 96 Falmouth St., P.O. Box 9300, Portland, ME 04101-9300 (United States)

    2014-08-01

    Cobalt exposure is increasing as cobalt demand rises worldwide due to its use in enhancing rechargeable battery efficiency, super-alloys, and magnetic products. Cobalt is considered a possible human carcinogen with the lung being a primary target. However, few studies have considered cobalt-induced toxicity in human lung cells. Therefore, in this study, we sought to determine the cytotoxicity and genotoxicity of particulate and soluble cobalt in human lung cells. Cobalt oxide and cobalt chloride were used as representative particulate and soluble cobalt compounds, respectively. Exposure to both particulate and soluble cobalt induced a concentration-dependent increase in cytotoxicity, genotoxicity, and intracellular cobalt ion levels. Based on intracellular cobalt ion levels, we found that soluble cobalt was more cytotoxic than particulate cobalt while particulate and soluble cobalt induced similar levels of genotoxicity. However, soluble cobalt induced cell cycle arrest indicated by the lack of metaphases at much lower intracellular cobalt concentrations compared to cobalt oxide. Accordingly, we investigated the role of particle internalization in cobalt oxide-induced toxicity and found that particle-cell contact was necessary to induce cytotoxicity and genotoxicity after cobalt exposure. These data indicate that cobalt compounds are cytotoxic and genotoxic to human lung fibroblasts, and solubility plays a key role in cobalt-induced lung toxicity. - Highlights: • Particulate and soluble cobalt are cytotoxic and genotoxic to human lung cells. • Soluble cobalt induces more cytotoxicity compared to particulate cobalt. • Soluble and particulate cobalt induce similar levels of genotoxicity. • Particle-cell contact is required for particulate cobalt-induced toxicity.

  8. Connective tissue growth factor stimulates the proliferation, migration and differentiation of lung fibroblasts during paraquat-induced pulmonary fibrosis.

    Science.gov (United States)

    Yang, Zhizhou; Sun, Zhaorui; Liu, Hongmei; Ren, Yi; Shao, Danbing; Zhang, Wei; Lin, Jinfeng; Wolfram, Joy; Wang, Feng; Nie, Shinan

    2015-07-01

    It is well established that paraquat (PQ) poisoning can cause severe lung injury during the early stages of exposure, finally leading to irreversible pulmonary fibrosis. Connective tissue growth factor (CTGF) is an essential growth factor that is involved in tissue repair and pulmonary fibrogenesis. In the present study, the role of CTGF was examined in a rat model of pulmonary fibrosis induced by PQ poisoning. Histological examination revealed interstitial edema and extensive cellular thickening of interalveolar septa at the early stages of poisoning. At 2 weeks after PQ administration, lung tissue sections exhibited a marked thickening of the alveolar walls with an accumulation of interstitial cells with a fibroblastic appearance. Masson's trichrome staining revealed a patchy distribution of collagen deposition, indicating pulmonary fibrogenesis. Western blot analysis and immunohistochemical staining of tissue samples demonstrated that CTGF expression was significantly upregulated in the PQ-treated group. Similarly, PQ treatment of MRC-5 human lung fibroblast cells caused an increase in CTGF in a dose-dependent manner. Furthermore, the addition of CTGF to MRC-5 cells triggered cellular proliferation and migration. In addition, CTGF induced the differentiation of fibroblasts to myofibroblasts, as was evident from increased expression of α-smooth muscle actin (α-SMA) and collagen. These findings demonstrate that PQ causes increased CTGF expression, which triggers proliferation, migration and differentiation of lung fibroblasts. Therefore, CTGF may be important in PQ-induced pulmonary fibrogenesis, rendering this growth factor a potential pharmacological target for reducing lung injury.

  9. Growth characteristics of fibroblasts isolated from the trunk and distal aspect of the limb of horses and ponies.

    Science.gov (United States)

    Miller, C B; Wilson, D A; Keegan, K G; Kreeger, J M; Adelstein, E H; Ganjam, V K

    2000-01-01

    To determine if there is a difference in in vitro growth of fibroblasts isolated from the trunk and distal aspect of the limb of horses and ponies. To determine the effects of a corticosteroid and monokine on in vitro growth of fibroblasts isolated from the trunk and distal aspect of the limb of horses and ponies. Growth of fibroblasts from tissues harvested from the trunk and limb were compared from horse and pony samples grown in control media and control media with triamcinolone or monokine added. Dermal and subcutaneous tissue from 22 horses and 17 ponies of various ages and breeds. Fibroblast growth was assessed by tritiated thymidine uptake using standard cell culture techniques. The effect of a monokine or triamcinolone plus control media were compared with control media for fibroblast growth. Fibroblast growth from tissues isolated from the horse limb was significantly less than growth from the horse trunk and the limb and trunk of ponies. Monokine was more effective than triamcinolone in suppressing fibroblast growth from tissues isolated from the trunk and limb in both horses and ponies. There are growth differences in fibroblasts isolated from the limb of horses compared with those isolated from the trunk and from the limb and trunk of ponies. The difference in fibroblast growth from tissues isolated from the trunk and limb of horses and ponies may provide evidence for the difference reported in the healing characteristics of limb wounds in horses and ponies. Influencing fibroblast growth may provide a key to controlling the development of exuberant granulation tissue in horses and ponies.

  10. Hypoxia-Induced Collagen Synthesis of Human Lung Fibroblasts by Activating the Angiotensin System

    Directory of Open Access Journals (Sweden)

    Shan-Shan Liu

    2013-12-01

    Full Text Available The exact molecular mechanism that mediates hypoxia-induced pulmonary fibrosis needs to be further clarified. The aim of this study was to explore the effect and underlying mechanism of angiotensin II (Ang II on collagen synthesis in hypoxic human lung fibroblast (HLF cells. The HLF-1 cell line was used for in vitro studies. Angiotensinogen (AGT, angiotensin converting enzyme (ACE, angiotensin II type 1 receptor (AT1R and angiotensin II type 2 receptor (AT2R expression levels in human lung fibroblasts were analysed using real-time polymerase chain reaction (RT-PCR after hypoxic treatment. Additionally, the collagen type I (Col-I, AT1R and nuclear factor κappaB (NF-κB protein expression levels were detected using Western blot analysis, and NF-κB nuclear translocation was measured using immunofluorescence localization analysis. Ang II levels in HLF-1 cells were measured with an enzyme-linked immunosorbent assay (ELISA. We found that hypoxia increased Col-I mRNA and protein expression in HLF-1 cells, and this effect could be inhibited by an AT1R or AT2R inhibitor. The levels of NF-κB, RAS components and Ang II production in HLF-1 cells were significantly increased after the hypoxia exposure. Hypoxia or Ang II increased NF-κB-p50 protein expression in HLF-1 cells, and the special effect could be inhibited by telmisartan (TST, an AT1R inhibitor, and partially inhibited by PD123319, an AT2R inhibitor. Importantly, hypoxia-induced NF-κB nuclear translocation could be nearly completely inhibited by an AT1R or AT2R inhibitor. Furthermore pyrrolidine dithiocarbamate (PDTC, a NF-κB blocker, abolished the expression of hypoxia-induced AT1R and Col-I in HLF-1 cells. Our results indicate that Ang II-mediated NF-κB signalling via ATR is involved in hypoxia-induced collagen synthesis in human lung fibroblasts.

  11. Bronchodilatory effect of deep inspiration in freshly isolated sheep lungs.

    Science.gov (United States)

    Wong, William D; Wang, Lu; Paré, Peter D; Seow, Chun Y

    2017-02-01

    Taking a big breath is known to reverse bronchoconstriction induced by bronchochallenge in healthy subjects; this bronchodilatory effect of deep inspiration (DI) is diminished in asthmatics. The mechanism underlying the DI effect is not clear. Observations from experiments using isolated airway smooth muscle (ASM) preparations and airway segments suggest that straining of ASM due to DI could lead to bronchodilation, possibly due to strain-induced reduction in ASM contractility. However, factors external to the lung cannot be excluded as potential causes for the DI effect. Neural reflex initiated by stretch receptors in the lung are known to inhibit the broncho-motor tone and enhance vasodilatation; the former directly reduces airway resistance, and the latter facilitates removal of contractile agonists through the bronchial circulation. If the DI effect is solely mediated by factors extrinsic to the lung, the DI effect would be absent in isolated, nonperfused lungs. Here we examined the DI effect in freshly isolated, nonperfused sheep lungs. We found that imposition of DI on isolated lungs resulted in significant bronchodilation, that this DI effect was present only after the lungs were challenged with a contractile agonist (acetylcholine or histamine), and that the effect was independent of the difference in lung volume observed pre- and post-DI. We conclude that a significant portion of the bronchodilatory DI effect stems from factors internal to the lung related to the activation of ASM. Copyright © 2017 the American Physiological Society.

  12. Isolation of Moellerella wisconsensis from the lung of a goat.

    Science.gov (United States)

    Casalinuovo, F; Musarella, R

    2009-09-18

    This report describes the first isolation of Moellerella wisconsensis from the lung of a goat in Italy and represents the fourth isolation of this rare enterobacterial species from non-human sources reported in the literature. In fact, since its first isolation, M. wisconsensis has only been described in a few other occasions and often in clinical samples associated with cases of human enteritis.

  13. Changes in CD11b and L-selectin expression on eosinophils are mediated by human lung fibroblasts in vitro

    NARCIS (Netherlands)

    Spoelstra, FM; Hovenga, H; Noordhoek, JA; Postma, DS; Kauffman, HF

    Eosinophilic airway infiltration is a central feature in asthma. Eosinophils recovered from bronchoalveolar fluid show an activated phenotype, e.g., increased CD11b and decreased L-selectin expression. We investigated whether lung fibroblasts are able to activate eosinophils in vitro, and if so,

  14. Isolation and culture of primary adult skin fibroblasts from the Asian elephant (Elephas maximus

    Directory of Open Access Journals (Sweden)

    Puntita Siengdee

    2018-01-01

    Full Text Available Background Primary cultures from Asian elephants (Elephas maximus allow scientists to obtain representative cells that have conserved most of their original characteristics, function, physiology and biochemistry. This technique has thus gained significant importance as a foundation for further cellular, cell biology and molecular research. Therefore, the aim of this study was to describe conditions for the successful establishment of primary adult fibroblasts from Asian elephant carcasses. Methods Ear tissue sample collection from Asian elephant carcasses and our recommendations are given. We describe here a simple modified protocol for successful isolation and maintenance of primary adult fibroblasts from elephant ear skin. Ear samples from each individual (five 3 × 3 cm2 pieces were brought to the laboratory within 3 h after collection, kept in transportation medium at 0–4 °C. The ear tissues were prepared by a combination of 10% collagenase type II digestion procedure together with a simple explant procedure. Primary fibroblasts were cultured at 37 °C in Dulbecco’s modified Eagle’s medium (DMEM with 20% fetal calf serum (FCS in a humidified atmosphere containing 5% CO2. After the third passage, fibroblasts were routinely trypsinized with 0.25% trypsin/EDTA and cultured in DMEM with 10% FCS at 37 °C and 5% CO2. Traditional cell counting method was used to measure cell viability and growth curve. Long-term storage of cells used freezing medium consisting of 40% FCS (v/v. Results We explored the most suitable conditions during sample collection (post-mortem storage time and sample storage temperature, which is the most important step in determining primary outgrowth. Our study successfully established and cultured primary adult skin fibroblasts obtained from post-mortem E. maximus ear skin tissues from six carcasses, with a success rate of around 83.3%. Outgrowth could be seen 4–12 days after explantation, and epithelial

  15. Interactions of ozone and antineoplastic drugs on rat lung fibroblasts and Walker rat carcinoma cells

    International Nuclear Information System (INIS)

    Wenzel, D.G.; Morgan, D.L.

    1983-01-01

    Cultured rat lung fibroblasts (F-cells) and Walker rat carcinoma cells (WRC-cells) labeled with 51 Cr were exposed to the following antitumor drugs alone or with O 3 : carmustine (BCNU), doxorubicin (Dox), cisplatin (CPt), mitomycin C (Mit C) or vitamin K 3 (Vit K). Release of 51 Cr (cell injury) was greater for F-cells than WRC-cells with any single treatment. Pretreatment with any drug (400 microM), except for Vit K with WRC-cells, did not significantly increase O 3 -induced loss of 51 Cr. Co-exposure of F-cells to drugs and O 3 resulted in a marked potentiation of O 3 -induced injury with Vit K, and an inhibition with Dox

  16. Histone Deacetylase Inhibition Downregulates Collagen 3A1 in Fibrotic Lung Fibroblasts

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    Victor J. Thannickal

    2013-09-01

    Full Text Available Idiopathic pulmonary fibrosis (IPF is a deadly disease characterized by chronic inflammation and excessive collagen accumulation in the lung. Myofibroblasts are the primary collagen-producing cells in pulmonary fibrosis. Histone deacetylase inhibitor (HDACi can affect gene expression, and some, such as suberoylanilide hydroxamic acid (SAHA, are US FDA approved for cancer treatment. In this study, we investigated SAHA’s effects on the expression of collagen III alpha 1 (COL3A1 in primary human IPF fibroblasts and in a murine model of pulmonary fibrosis. We observed that increased COL3A1 expression in IPF fibroblasts can be substantially reduced by SAHA treatment at the level of transcription as detected by RT-PCR; collagen III protein level was also reduced, as detected by Western blots and immunofluorescence. The deacetylation inhibitor effect of SAHA was verified by observing higher acetylation levels of both histone H3 and H4 in treated IPF cells. Chromatin immunoprecipitation (ChIP experiments demonstrated that the reduced expression of COL3A1 by SAHA is with increased association of the repressive chromatin marker, H3K27Me3, and decreased association of the active chromatin marker, H3K9Ac. In our murine model of bleomycin-induced pulmonary fibrosis, the SAHA treated group demonstrated significantly less collagen III, as detected by immunohistochemistry. Our data indicate that the HDACi SAHA alters the chromatin associated with COL3A1, resulting in its decreased expression.

  17. Electronic cigarette aerosols and copper nanoparticles induce mitochondrial stress and promote DNA fragmentation in lung fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Lerner, Chad A.; Rutagarama, Pierrot; Ahmad, Tanveer; Sundar, Isaac K.; Elder, Alison; Rahman, Irfan, E-mail: irfan_rahman@urmc.rochester.edu

    2016-09-02

    Oxidants or nanoparticles have recently been identified as constituents of aerosols released from various styles of electronic cigarettes (E-cigs). Cells in the lung may be directly exposed to these constituents and harbor reactive properties capable of incurring acute cell injury. Our results show mitochondria are sensitive to both E-cig aerosols and aerosol containing copper nanoparticles when exposed to human lung fibroblasts (HFL-1) using an Air-Liquid Interface culture system, evident by elevated levels of mitochondrial ROS (mtROS). Increased mtROS after aerosol exposure is associated with reduced stability of OxPhos electron transport chain (ETC) complex IV subunit and nuclear DNA fragmentation. Increased levels of IL-8 and IL-6 in HFL-1 conditioned media were also observed. These findings reveal both mitochondrial, genotoxic, and inflammatory stresses are features of direct cell exposure to E-cig aerosols which are ensued by inflammatory duress, raising a concern on deleterious effect of vaping. - Graphical abstract: Oxidants and possibly reactive properties of metal particles in E-cig aerosols impart mitochondrial oxidative stress and DNA damage. These biological effects accompany inflammatory response which may raise concern regarding long term E-cig use. Mitochondria may be particularly sensitive to reactive properties of E-cig aerosols in addition to the potential for them to induce genotoxic stress by generating increased ROS. - Highlights: • Mitochondria are sensitive to both E-cig aerosols and metal nanoparticles. • Increased mtROS by E-cig aerosol is associated with disrupted mitochondrial energy. • E-cig causes nuclear DNA fragmentation. • E-cig aerosols induce pro-inflammatory response in human fibroblasts.

  18. Electronic cigarette aerosols and copper nanoparticles induce mitochondrial stress and promote DNA fragmentation in lung fibroblasts

    International Nuclear Information System (INIS)

    Lerner, Chad A.; Rutagarama, Pierrot; Ahmad, Tanveer; Sundar, Isaac K.; Elder, Alison; Rahman, Irfan

    2016-01-01

    Oxidants or nanoparticles have recently been identified as constituents of aerosols released from various styles of electronic cigarettes (E-cigs). Cells in the lung may be directly exposed to these constituents and harbor reactive properties capable of incurring acute cell injury. Our results show mitochondria are sensitive to both E-cig aerosols and aerosol containing copper nanoparticles when exposed to human lung fibroblasts (HFL-1) using an Air-Liquid Interface culture system, evident by elevated levels of mitochondrial ROS (mtROS). Increased mtROS after aerosol exposure is associated with reduced stability of OxPhos electron transport chain (ETC) complex IV subunit and nuclear DNA fragmentation. Increased levels of IL-8 and IL-6 in HFL-1 conditioned media were also observed. These findings reveal both mitochondrial, genotoxic, and inflammatory stresses are features of direct cell exposure to E-cig aerosols which are ensued by inflammatory duress, raising a concern on deleterious effect of vaping. - Graphical abstract: Oxidants and possibly reactive properties of metal particles in E-cig aerosols impart mitochondrial oxidative stress and DNA damage. These biological effects accompany inflammatory response which may raise concern regarding long term E-cig use. Mitochondria may be particularly sensitive to reactive properties of E-cig aerosols in addition to the potential for them to induce genotoxic stress by generating increased ROS. - Highlights: • Mitochondria are sensitive to both E-cig aerosols and metal nanoparticles. • Increased mtROS by E-cig aerosol is associated with disrupted mitochondrial energy. • E-cig causes nuclear DNA fragmentation. • E-cig aerosols induce pro-inflammatory response in human fibroblasts.

  19. Clarithromycin attenuates IL-13–induced periostin production in human lung fibroblasts

    Directory of Open Access Journals (Sweden)

    Kosaku Komiya

    2017-02-01

    Full Text Available Abstract Background Periostin is a biomarker indicating the presence of type 2 inflammation and submucosal fibrosis; serum periostin levels have been associated with asthma severity. Macrolides have immunomodulatory effects and are considered a potential therapy for patients with severe asthma. Therefore, we investigated whether macrolides can also modulate pulmonary periostin production. Methods Using quantitative PCR and ELISA, we measured periostin production in human lung fibroblasts stimulated by interleukin-13 (IL-13 in the presence of two 14-member–ring macrolides—clarithromycin or erythromycin—or a 16-member–ring macrolide, josamycin. Phosphorylation of signal transducers and activators of transcription 6 (STAT6, downstream of IL-13 signaling, was evaluated by Western blotting. Changes in global gene expression profile induced by IL-13 and/or clarithromycin were assessed by DNA microarray analysis. Results Clarithromycin and erythromycin, but not josamycin, inhibited IL-13–stimulated periostin production. The inhibitory effects of clarithromycin were stronger than those of erythromycin. Clarithromycin significantly attenuated STAT6 phosphorylation induced by IL-13. Global gene expression analyses demonstrated that IL-13 increased mRNA expression of 454 genes more than 4-fold, while decreasing its expression in 390 of these genes (85.9%, mainly “extracellular,” “plasma membrane,” or “defense response” genes. On the other hand, clarithromycin suppressed 9.8% of the genes in the absence of IL-13. Clarithromycin primarily attenuated the gene expression of extracellular matrix protein, including periostin, especially after IL-13. Conclusions Clarithromycin suppressed IL-13–induced periostin production in human lung fibroblasts, in part by inhibiting STAT6 phosphorylation. This suggests a novel mechanism of the immunomodulatory effect of clarithromycin in asthmatic airway inflammation and fibrosis.

  20. Cellular metabolic rates from primary dermal fibroblast cells isolated from birds of different body masses.

    Science.gov (United States)

    Jimenez, Ana Gabriela; Williams, Joseph B

    2014-10-01

    The rate of metabolism is the speed at which organisms use energy, an integration of energy transformations within the body; it governs biological processes that influence rates of growth and reproduction. Progress at understanding functional linkages between whole organism metabolic rate and underlying mechanisms that influence its magnitude has been slow despite the central role this issue plays in evolutionary and physiological ecology. Previous studies that have attempted to relate how cellular processes translate into whole-organism physiology have done so over a range of body masses of subjects. However, the data still remains controversial when observing metabolic rates at the cellular level. To bridge the gap between these ideas, we examined cellular metabolic rate of primary dermal fibroblasts isolated from 49 species of birds representing a 32,000-fold range in body masses to test the hypothesis that metabolic rate of cultured cells scales with body size. We used a Seahorse XF-96 Extracellular flux analyzer to measure cellular respiration in fibroblasts. Additionally, we measured fibroblast size and mitochondrial content. We found no significant correlation between cellular metabolic rate, cell size, or mitochondrial content and body mass. Additionally, there was a significant relationship between cellular basal metabolic rate and proton leak in these cells. We conclude that metabolic rate of cells isolated in culture does not scale with body mass, but cellular metabolic rate is correlated to growth rate in birds. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. TRX is up-regulated by fibroblast growth factor-2 in lung carcinoma.

    Science.gov (United States)

    Deng, Zheng-Hao; Cao, Hui-Qiu; Hu, Yong-Bin; Wen, Ji-Fang; Zhou, Jian-Hua

    2011-01-01

    We have previously shown that exogenous fibroblast growth factor-2 (FGF-2) inhibits apoptosis of the small-cell lung cancer (SCLC) cell line NCI-H446, but the underlying mechanism remains unknown. In this study, the protein profiles of FGF-2-treated and untreated NCI-H446 cells were determined by 2-D gel electrophoresis combined with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and bioinformatics. Differential expression analysis of the protein profiles after FGF-2 treatment identified a total of 24 protein spots, of which nine were up-regulated and 15 were down-regulated. Four proteins were identified by MALDI-TOF-MS: thioredoxin (TRX), visfatin, ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) and Cu/Zn superoxide dismutase (CuZn-SOD). Western blotting revealed that TRX was up-regulated in NCI-H446 and A549 cells treated with FGF-2. Furthermore, immunohistochemical staining confirmed that both FGF-2 and TRX were overexpressed in lung cancer tissues and could be correlated with both lymph node metastasis and clinical stage. These data indicate that TRX may be involved in the FGF-2 signaling pathway. © 2010 The Authors. APMIS © 2010 APMIS.

  2. The alpha2-adrenoreceptor agonist dexmedetomidine protects against lipopolysaccharide-induced apoptosis via inhibition of gap junctions in lung fibroblasts.

    Science.gov (United States)

    Zhang, Yuan; Tan, Xiaoming; Xue, Lianfang

    2018-01-01

    The α2-adrenoceptor inducer dexmedetomidine protects against acute lung injury (ALI), but the mechanism of this effect is largely unknown. The present study investigated the effect of dexmedetomidine on apoptosis induced by lipopolysaccharide (LPS) and the relationship between this effect and gap junction intercellular communication in human lung fibroblast cell line. Flow cytometry was used to detect apoptosis induced by LPS. Parachute dye coupling assay was used to measure gap junction function, and western blot analysis was used to determine the expression levels of connexin43 (Cx43). The results revealed that exposure of human lung fibroblast cell line to LPS for 24 h increased the apoptosis, and pretreatment of dexmedetomidine and 18α-GA significantly reduced LPS-induced apoptosis. Dexmedetomidine exposure for 1 h inhibited gap junction function mainly via a decrease in Cx43 protein levels in human lung fibroblast cell line. These results demonstrated that the inhibition of gap junction intercellular communication by dexmedetomidine affected the LPS-induced apoptosis through inhibition of gap junction function by reducing Cx43 protein levels. The present study provides evidence of a novel mechanism underlying the effects of analgesics in counteracting ALI. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. FK506-binding protein 10 (FKBP10) regulates lung fibroblast migration via collagen VI synthesis.

    Science.gov (United States)

    Knüppel, Larissa; Heinzelmann, Katharina; Lindner, Michael; Hatz, Rudolf; Behr, Jürgen; Eickelberg, Oliver; Staab-Weijnitz, Claudia A

    2018-04-19

    In idiopathic pulmonary fibrosis (IPF), fibroblasts gain a more migratory phenotype and excessively secrete extracellular matrix (ECM), ultimately leading to alveolar scarring and progressive dyspnea. Here, we analyzed the effects of deficiency of FK506-binding protein 10 (FKBP10), a potential IPF drug target, on primary human lung fibroblast (phLF) adhesion and migration. Using siRNA, FKBP10 expression was inhibited in phLF in absence or presence of 2ng/ml transforming growth factor-β1 (TGF-β1) and 0.1mM 2-phosphoascorbate. Effects on cell adhesion and migration were monitored by an immunofluorescence (IF)-based attachment assay, a conventional scratch assay, and single cell tracking by time-lapse microscopy. Effects on expression of key players in adhesion dynamics and migration were analyzed by qPCR and Western Blot. Colocalization was evaluated by IF microscopy and by proximity ligation assays. FKBP10 knockdown significantly attenuated adhesion and migration of phLF. Expression of collagen VI was decreased, while expression of key components of the focal adhesion complex was mostly upregulated. The effects on migration were 2-phosphoascorbate-dependent, suggesting collagen synthesis as the underlying mechanism. FKBP10 colocalized with collagen VI and coating culture dishes with collagen VI, and to a lesser extent with collagen I, abolished the effect of FKBP10 deficiency on migration. These findings show, to our knowledge for the first time, that FKBP10 interacts with collagen VI and that deficiency of FKBP10 reduces phLF migration mainly by downregulation of collagen VI synthesis. The results strengthen FKBP10 as an important intracellular regulator of ECM remodeling and support the concept of FKBP10 as drug target in IPF.

  4. Quiescence does not affect p53 and stress response by irradiation in human lung fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Dai, Jiawen [Molecular Radiobiology Laboratory, Division of Cellular and Molecular Research (Singapore); Itahana, Koji, E-mail: koji.itahana@duke-nus.edu.sg [Cancer and Stem Cell Biology Program, Duke-NUS Graduate Medical School (Singapore); Baskar, Rajamanickam, E-mail: r.baskar@nccs.com.sg [Molecular Radiobiology Laboratory, Division of Cellular and Molecular Research (Singapore); Department of Radiation Oncology, National Cancer Centre (Singapore)

    2015-02-27

    Cells in many organs exist in both proliferating and quiescent states. Proliferating cells are more radio-sensitive, DNA damage pathways including p53 pathway are activated to undergo either G{sub 1}/S or G{sub 2}/M arrest to avoid entering S and M phase with DNA damage. On the other hand, quiescent cells are already arrested in G{sub 0}, therefore there may be fundamental difference of irradiation response between proliferating and quiescent cells, and this difference may affect their radiosensitivity. To understand these differences, proliferating and quiescent human normal lung fibroblasts were exposed to 0.10–1 Gy of γ-radiation. The response of key proteins involved in the cell cycle, cell death, and metabolism as well as histone H2AX phosphorylation were examined. Interestingly, p53 and p53 phosphorylation (Ser-15), as well as the cyclin-dependent kinase inhibitors p21 and p27, were induced similarly in both proliferating and quiescent cells after irradiation. Furthermore, the p53 protein half-life, and expression of cyclin A, cyclin E, proliferating cell nuclear antigen (PCNA), Bax, or cytochrome c expression as well as histone H2AX phosphorylation were comparable after irradiation in both phases of cells. The effect of radioprotection by a glycogen synthase kinase 3β inhibitor on p53 pathway was also similar between proliferating and quiescent cells. Our results showed that quiescence does not affect irradiation response of key proteins involved in stress and DNA damage at least in normal fibroblasts, providing a better understanding of the radiation response in quiescent cells, which is crucial for tissue repair and regeneration. - Highlights: • p53 response by irradiation was similar between proliferating and quiescent cells. • Quiescent cells showed similar profiles of cell cycle proteins after irradiation. • Radioprotection of GSK-3β inhibitor caused similar effects between these cells. • Quiescence did not affect p53 response despite its

  5. Quiescence does not affect p53 and stress response by irradiation in human lung fibroblasts

    International Nuclear Information System (INIS)

    Dai, Jiawen; Itahana, Koji; Baskar, Rajamanickam

    2015-01-01

    Cells in many organs exist in both proliferating and quiescent states. Proliferating cells are more radio-sensitive, DNA damage pathways including p53 pathway are activated to undergo either G 1 /S or G 2 /M arrest to avoid entering S and M phase with DNA damage. On the other hand, quiescent cells are already arrested in G 0 , therefore there may be fundamental difference of irradiation response between proliferating and quiescent cells, and this difference may affect their radiosensitivity. To understand these differences, proliferating and quiescent human normal lung fibroblasts were exposed to 0.10–1 Gy of γ-radiation. The response of key proteins involved in the cell cycle, cell death, and metabolism as well as histone H2AX phosphorylation were examined. Interestingly, p53 and p53 phosphorylation (Ser-15), as well as the cyclin-dependent kinase inhibitors p21 and p27, were induced similarly in both proliferating and quiescent cells after irradiation. Furthermore, the p53 protein half-life, and expression of cyclin A, cyclin E, proliferating cell nuclear antigen (PCNA), Bax, or cytochrome c expression as well as histone H2AX phosphorylation were comparable after irradiation in both phases of cells. The effect of radioprotection by a glycogen synthase kinase 3β inhibitor on p53 pathway was also similar between proliferating and quiescent cells. Our results showed that quiescence does not affect irradiation response of key proteins involved in stress and DNA damage at least in normal fibroblasts, providing a better understanding of the radiation response in quiescent cells, which is crucial for tissue repair and regeneration. - Highlights: • p53 response by irradiation was similar between proliferating and quiescent cells. • Quiescent cells showed similar profiles of cell cycle proteins after irradiation. • Radioprotection of GSK-3β inhibitor caused similar effects between these cells. • Quiescence did not affect p53 response despite its known role in

  6. Identification of pulmonary PDGFRalpha-positive fibroblast specific miRNA and mRNA expression profiles during postnatal lung development

    OpenAIRE

    Dontireddy, Daria Agnieszka

    2015-01-01

    BACKGROUND AND AIM The process of alveolarization is tightly regulated and requires the contribution of different subpopulations of fibroblasts such as myofibroblasts, lipofibroblasts and platelet-derived growth factor receptor alpha (PDGFRalpha)-positive fibroblasts. Each of this fibroblasts subset fulfills certain functions during lung development in a time-dependent manner. In particular PDGFRalpha-positive cells are crucial for alveolar septation and myofibroblasts differentiation. PDG...

  7. MicroRNA-26a modulates transforming growth factor beta-1-induced proliferation in human fetal lung fibroblasts

    International Nuclear Information System (INIS)

    Li, Xiaoou; Liu, Lian; Shen, Yongchun; Wang, Tao; Chen, Lei; Xu, Dan; Wen, Fuqiang

    2014-01-01

    Highlights: • Endogenous miR-26a inhibits TGF-beta 1 induced proliferation of lung fibroblasts. • miR-26a induces G1 arrest through directly targeting 3′-UTR of CCND2. • TGF indispensable receptor, TGF-beta R I, is regulated by miR-26a. • miR-26a acts through inhibiting TGF-beta 2 feedback loop to reduce TGF-beta 1. • Collagen type I and connective tissue growth factor are suppressed by miR-26a. - Abstract: MicroRNA-26a is a newly discovered microRNA that has a strong anti-tumorigenic capacity and is capable of suppressing cell proliferation and activating tumor-specific apoptosis. However, whether miR-26a can inhibit the over-growth of lung fibroblasts remains unclear. The relationship between miR-26a and lung fibrosis was explored in the current study. We first investigated the effect of miR-26a on the proliferative activity of human lung fibroblasts with or without TGF-beta1 treatment. We found that the inhibition of endogenous miR-26a promoted proliferation and restoration of mature miR-26a inhibited the proliferation of human lung fibroblasts. We also examined that miR-26a can block the G1/S phase transition via directly targeting 3′-UTR of CCND2, degrading mRNA and decreasing protein expression of Cyclin D2. Furthermore, we showed that miR-26a mediated a TGF-beta 2-TGF-beta 1 feedback loop and inhibited TGF-beta R I activation. In addition, the overexpression of miR-26a also significantly suppressed the TGF-beta 1-interacting-CTGF–collagen fibrotic pathway. In summary, our studies indicated an essential role of miR-26a in the anti-fibrotic mechanism in TGF-beta1-induced proliferation in human lung fibroblasts, by directly targeting Cyclin D2, regulating TGF-beta R I as well as TGF-beta 2, and suggested the therapeutic potential of miR-26a in ameliorating lung fibrosis

  8. MicroRNA-26a modulates transforming growth factor beta-1-induced proliferation in human fetal lung fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xiaoou [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Department of Respiratory Medicine, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Liu, Lian [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Shen, Yongchun; Wang, Tao; Chen, Lei; Xu, Dan [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Department of Respiratory Medicine, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Wen, Fuqiang, E-mail: wenfuqiang.scu@gmail.com [Division of Pulmonary Diseases, State Key Laboratory of Biotherapy of China, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China); Department of Respiratory Medicine, West China Hospital, West China School of Medicine, Sichuan University, Chengdu, Sichuan (China)

    2014-11-28

    Highlights: • Endogenous miR-26a inhibits TGF-beta 1 induced proliferation of lung fibroblasts. • miR-26a induces G1 arrest through directly targeting 3′-UTR of CCND2. • TGF indispensable receptor, TGF-beta R I, is regulated by miR-26a. • miR-26a acts through inhibiting TGF-beta 2 feedback loop to reduce TGF-beta 1. • Collagen type I and connective tissue growth factor are suppressed by miR-26a. - Abstract: MicroRNA-26a is a newly discovered microRNA that has a strong anti-tumorigenic capacity and is capable of suppressing cell proliferation and activating tumor-specific apoptosis. However, whether miR-26a can inhibit the over-growth of lung fibroblasts remains unclear. The relationship between miR-26a and lung fibrosis was explored in the current study. We first investigated the effect of miR-26a on the proliferative activity of human lung fibroblasts with or without TGF-beta1 treatment. We found that the inhibition of endogenous miR-26a promoted proliferation and restoration of mature miR-26a inhibited the proliferation of human lung fibroblasts. We also examined that miR-26a can block the G1/S phase transition via directly targeting 3′-UTR of CCND2, degrading mRNA and decreasing protein expression of Cyclin D2. Furthermore, we showed that miR-26a mediated a TGF-beta 2-TGF-beta 1 feedback loop and inhibited TGF-beta R I activation. In addition, the overexpression of miR-26a also significantly suppressed the TGF-beta 1-interacting-CTGF–collagen fibrotic pathway. In summary, our studies indicated an essential role of miR-26a in the anti-fibrotic mechanism in TGF-beta1-induced proliferation in human lung fibroblasts, by directly targeting Cyclin D2, regulating TGF-beta R I as well as TGF-beta 2, and suggested the therapeutic potential of miR-26a in ameliorating lung fibrosis.

  9. Induction of Mitochondrial DNA Deletion by Ionizing Radiation in Human Lung Fibroblast IMR-90 Cells

    International Nuclear Information System (INIS)

    Eom, Hyeon Soo; Jung, U Hee; Park, Hae Ran; Jo, Sung Kee

    2009-01-01

    Mitochondrial DNA (mtDNA) deletion is a well-known marker for oxidative stress and aging and also contributes to their unfavorable effects in cultured cells and animal tissues. This study was conducted to investigate the effect of ionizing radiation (IR) on mtDNA deletion and the involvement of reactive oxygen species (ROS) in this process in human lung fibroblast (IMR-90) cells. Young IMR-90 cells at population doubling (PD) 39 were irradiated with 137 Cs -rays and the intracellular ROS level was determined by 2',7'-dichlorofluorescein diacetate (DCFH-DA) and mtDNA common deletion (4977bp) was detected by nested PCR. Old cells at PD 55 and H 2 O 2 -treated young cells were compared as the positive control. IR increased the intracellular ROS level and mtDNA 4977 bp deletion in IMR-90 cells dose-dependently. The increases of ROS level and mtDNA deletion were also observed in old cells and H 2 O 2 -treated young cells. To confirm the increased ROS level is essential for mtDNA deletion in irradiated cells, the effects of N-acetylcysteine (NAC) on IRinduced ROS and mtDNA deletion were examined. 5 mM NAC significantly attenuated the IR-induced ROS increase and mtDNA deletion. These results suggest that IR induces the mtDNA deletion and this process is mediated by ROS in IMR-90 cells

  10. Induction of Mitochondrial DNA Deletion by Ionizing Radiation in Human Lung Fibroblast IMR-90 Cells

    Energy Technology Data Exchange (ETDEWEB)

    Eom, Hyeon Soo; Jung, U Hee; Park, Hae Ran; Jo, Sung Kee [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2009-06-15

    Mitochondrial DNA (mtDNA) deletion is a well-known marker for oxidative stress and aging and also contributes to their unfavorable effects in cultured cells and animal tissues. This study was conducted to investigate the effect of ionizing radiation (IR) on mtDNA deletion and the involvement of reactive oxygen species (ROS) in this process in human lung fibroblast (IMR-90) cells. Young IMR-90 cells at population doubling (PD) 39 were irradiated with {sup 137}Cs -rays and the intracellular ROS level was determined by 2',7'-dichlorofluorescein diacetate (DCFH-DA) and mtDNA common deletion (4977bp) was detected by nested PCR. Old cells at PD 55 and H{sub 2}O{sub 2}-treated young cells were compared as the positive control. IR increased the intracellular ROS level and mtDNA 4977 bp deletion in IMR-90 cells dose-dependently. The increases of ROS level and mtDNA deletion were also observed in old cells and H{sub 2}O{sub 2}-treated young cells. To confirm the increased ROS level is essential for mtDNA deletion in irradiated cells, the effects of N-acetylcysteine (NAC) on IRinduced ROS and mtDNA deletion were examined. 5 mM NAC significantly attenuated the IR-induced ROS increase and mtDNA deletion. These results suggest that IR induces the mtDNA deletion and this process is mediated by ROS in IMR-90 cells.

  11. Fusobacterium necrophorum presenting as isolated lung nodules

    Directory of Open Access Journals (Sweden)

    Rajiv Sonti

    2015-01-01

    Full Text Available Fusobacterium necrophorum causes Lemierre's syndrome - a dramatic and distinct condition beginning with pharyngitis before proceeding to internal jugular vein septic thrombophlebitis and respiratory tract infection in otherwise healthy individuals. It is rare, but by far the most common pathway to parenchymal lung disease with this organism. Here we describe we a 34 year old healthy lady who was nontoxic without any antecedent illness who presented with lung nodules due to fusobacterium necrophorum as the sole manifestation of disease. Leading diagnostic consideration prior to culture data was pulmonary vasculitis. Identifying her disease process was a somewhat chance occurrence, and it began to resolve prior to antibiotic therapy. Though it would be difficult to recommend keen awareness of this organism given its rarity, it is important to consider that its scope may be broader than traditionally considered.

  12. Low-Dose Radiation Induces Cell Proliferation in Human Embryonic Lung Fibroblasts but not in Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Xinyue Liang

    2016-01-01

    Full Text Available Hormesis and adaptive responses are 2 important biological effects of low-dose ionizing radiation (LDR. In normal tissue, LDR induces hormesis as evinced by increased cell proliferation; however, whether LDR also increases tumor cell proliferation needs to be investigated. In this study, cell proliferation was assayed by total cell numbers and the Cell Counting Kit 8 assay. Mitogen-activated protein kinases (MAPK/extracellular signal-regulated kinase (ERK and phosphatidylinositol 3′ -kinase(PI3K-Akt (PI3K/AKT phosphorylation were determined by Western blot analysis. Human embryonic lung fibroblast 2BS and lung cancer NCI-H446 cell lines were irradiated with LDR at different doses (20-100 mGy. In response to 20 to 75 mGy X-rays, cell proliferation was significantly increased in 2BS but not in NCI-H446 cells. In 2BS cells, LDR at 20 to 75 mGy also stimulated phosphorylation of MAPK/ERK pathway proteins including ERK, MEK, and Raf and of the PI3K/AKT pathway protein AKT. To test whether ERK1/2 and AKT pathway activation was involved in the stimulation of cell proliferation in 2BS cells, the MAPK/ERK and PI3K/AKT pathways were inhibited using their specific inhibitors, U0126 and LY294002. U0126 decreased the phosphorylation of ERK1/2, and LY294002 decreased the phosphorylation of AKT; each could significantly inhibit LDR-induced 2BS cell proliferation. However, LDR did not stimulate these kinases, and kinase inhibitors also did not affect cell proliferation in the NCI-H446 cells. These results suggest that LDR stimulates cell proliferation via the activation of both MAPK/ERK and PI3K/AKT signaling pathways in 2BS but not in NCI-H446 cells. This finding implies the potential for applying LDR to protect normal tissues from radiotherapy without diminishing the efficacy of tumor therapy.

  13. Proteomic Analysis Shows Constitutive Secretion of MIF and p53-associated Activity of COX-2−/− Lung Fibroblasts

    Directory of Open Access Journals (Sweden)

    Mandar Dave

    2017-12-01

    Full Text Available The differential expression of two closelyassociated cyclooxygenase isozymes, COX-1 and COX-2, exhibited functions beyond eicosanoid metabolism. We hypothesized that COX-1 or COX-2 knockout lung fibroblasts may display altered protein profiles which may allow us to further differentiate the functional roles of these isozymes at the molecular level. Proteomic analysis shows constitutive production of macrophage migration inhibitory factor (MIF in lung fibroblasts derived from COX-2−/− but not wild-type (WT or COX-1−/− mice. MIF was spontaneously released in high levels into the extracellular milieu of COX2−/− fibroblasts seemingly from the preformed intracellular stores, with no change in the basal gene expression of MIF. The secretion and regulation of MIF in COX-2−/− was “prostaglandin-independent.” GO analysis showed that concurrent with upregulation of MIF, there is a significant surge in expression of genes related to fibroblast growth, FK506 binding proteins, and isomerase activity in COX-2−/− cells. Furthermore, COX-2−/− fibroblasts also exhibit a significant increase in transcriptional activity of various regulators, antagonists, and co-modulators of p53, as well as in the expression of oncogenes and related transcripts. Integrative Oncogenomics Cancer Browser (IntroGen analysis shows downregulation of COX-2 and amplification of MIF and/or p53 activity during development of glioblastomas, ependymoma, and colon adenomas. These data indicate the functional role of the MIF-COX-p53 axis in inflammation and cancer at the genomic and proteomic levels in COX-2-ablated cells. This systematic analysis not only shows the proinflammatory state but also unveils a molecular signature of a pro-oncogenic state of COX-1 in COX-2 ablated cells.

  14. Prevention of reperfusion lung injury by lidocaine in isolated rat lung ventilated with higher oxygen levels.

    Directory of Open Access Journals (Sweden)

    Das K

    2003-01-01

    Full Text Available BACKGROUND: Lidocaine, an antiarrhythmic drug has been shown to be effective against post-ischaemic reperfusion injury in heart. However, its effect on pulmonary reperfusion injury has not been investigated. AIMS: We investigated the effects of lidocaine on a postischaemic reperfused rat lung model. MATERIALS AND METHODS: Lungs were isolated and perfused at constant flow with Krebs-Henseilet buffer containing 4% bovine serum albumin, and ventilated with 95% oxygen mixed with 5% CO2. Lungs were subjected to ischaemia by stopping perfusion for 60 minutes followed by reperfusion for 10 minutes. Ischaemia was induced in normothermic conditions. RESULTS: Postischaemic reperfusion caused significant (p < 0.0001 higher wet-to-dry lung weight ratio, pulmonary arterial pressure and peak airway pressure compared to control lungs. Lidocaine, at a dose of 5mg/Kg b.w. was found to significantly (p < 0.0001 attenuate the increase in the wet-to-dry lung weight ratio, pulmonary arterial pressure and peak airway pressure observed in post-ischaemic lungs. CONCLUSION: Lidocaine is effective in preventing post-ischaemic reperfusion injury in isolated, perfused rat lung.

  15. Niacinamide mitigated the acute lung injury induced by phorbol myristate acetate in isolated rat's lungs

    Directory of Open Access Journals (Sweden)

    Lin Chia-Chih

    2012-03-01

    Full Text Available Abstract Background Phorbol myristate acetate (PMA is a strong neutrophil activator and has been used to induce acute lung injury (ALI. Niacinamide (NAC is a compound of B complex. It exerts protective effects on the ALI caused by various challenges. The purpose was to evaluate the protective effects of niacinamide (NAC on the PMA-induced ALI and associated changes. Methods The rat's lungs were isolated in situ and perfused with constant flow. A total of 60 isolated lungs were randomized into 6 groups to received Vehicle (DMSO 100 μg/g, PMA 4 μg/g (lung weight, cotreated with NAC 0, 100, 200 and 400 mg/g (lung weight. There were 10 isolated lungs in each group. We measured the lung weight and parameters related to ALI. The pulmonary arterial pressure and capillary filtration coefficient (Kfc were determined in isolated lungs. ATP (adenotriphosphate and PARP [poly(adenosine diphophate-ribose polymerase] contents in lung tissues were detected. Real-time PCR was employed to display the expression of inducible and endothelial NO synthases (iNOS and eNOS. The neutrophil-derived mediators in lung perfusate were determined. Results PMA caused increases in lung weight parameters. This agent produced pulmonary hypertension and increased microvascular permeability. It resulted in decrease in ATP and increase in PARP. The expression of iNOS and eNOS was upregulated following PMA. PMA increased the neutrophil-derived mediators. Pathological examination revealed lung edema and hemorrhage with inflammatory cell infiltration. Immunohistochemical stain disclosed the presence of iNOS-positive cells in macrophages and endothelial cells. These pathophysiological and biochemical changes were diminished by NAC treatment. The NAC effects were dose-dependent. Conclusions Our results suggest that neutrophil activation and release of neutrophil-derived mediators by PMA cause ALI and associated changes. NO production through the iNOS-producing cells plays a detrimental

  16. Niacinamide mitigated the acute lung injury induced by phorbol myristate acetate in isolated rat's lungs.

    Science.gov (United States)

    Lin, Chia-Chih; Hsieh, Nan-Kuang; Liou, Huey Ling; Chen, Hsing I

    2012-03-01

    Phorbol myristate acetate (PMA) is a strong neutrophil activator and has been used to induce acute lung injury (ALI). Niacinamide (NAC) is a compound of B complex. It exerts protective effects on the ALI caused by various challenges. The purpose was to evaluate the protective effects of niacinamide (NAC) on the PMA-induced ALI and associated changes. The rat's lungs were isolated in situ and perfused with constant flow. A total of 60 isolated lungs were randomized into 6 groups to received Vehicle (DMSO 100 μg/g), PMA 4 μg/g (lung weight), cotreated with NAC 0, 100, 200 and 400 mg/g (lung weight). There were 10 isolated lungs in each group. We measured the lung weight and parameters related to ALI. The pulmonary arterial pressure and capillary filtration coefficient (Kfc) were determined in isolated lungs. ATP (adenotriphosphate) and PARP [poly(adenosine diphophate-ribose) polymerase] contents in lung tissues were detected. Real-time PCR was employed to display the expression of inducible and endothelial NO synthases (iNOS and eNOS). The neutrophil-derived mediators in lung perfusate were determined. PMA caused increases in lung weight parameters. This agent produced pulmonary hypertension and increased microvascular permeability. It resulted in decrease in ATP and increase in PARP. The expression of iNOS and eNOS was upregulated following PMA. PMA increased the neutrophil-derived mediators. Pathological examination revealed lung edema and hemorrhage with inflammatory cell infiltration. Immunohistochemical stain disclosed the presence of iNOS-positive cells in macrophages and endothelial cells. These pathophysiological and biochemical changes were diminished by NAC treatment. The NAC effects were dose-dependent. Our results suggest that neutrophil activation and release of neutrophil-derived mediators by PMA cause ALI and associated changes. NO production through the iNOS-producing cells plays a detrimental role in the PMA-induced lung injury. ATP is beneficial

  17. The Apoptotic Effects of the P300 Activator on Breast Cancer and Lung Fibroblast Cell Lines

    Directory of Open Access Journals (Sweden)

    Mohammad Reza Salahshoor

    2013-10-01

    Full Text Available Background: P300 is an enzyme that acetylates histones during stress. It alsoacetylates several non-histone proteins, including P53 which is the most important tumorsuppressor gene. P53 plays an important role in the apoptosis of tumor cells. Hereby,this study describes the potency of cholera toxin B subunit as a P300 activator to induceapoptosis in a breast cancer cell line (MCF-7 and a lung fibroblast cell line (MRC-5as a non-tumorigenic control sample. Methods: MCF-7 and MRC-5 were cultured in RPMI-1640 and treated with orwithout cholera toxin B subunit at the concentration of 85.43 μmol/L, based on the half-maximal inhibitory concentration index at different times (24, 48 and 72 h. Thepercentage of apoptotic cells was measured by flow cytometry. Real-time quantitativeRT-PCR was performed to estimate the mRNA expression of P300 in MCF-7 and MRC-5 with cholera toxin B subunit at different times. We used the ELISA and Bradford proteintechniques to detect levels of total and acetylated P53 protein generated in MCF-7 andMRC-5. Results: Our findings indicated that the cholera toxin B subunit effectively andsignificantly induced more apoptosis in MCF-7 compared to MRC-5. We showed thatexpression of P300 up-regulated by increasing the time of the cholera toxin B subunittreatment in MCF-7 but not in MRC-5. In addition, the acetylated and total P53protein levels increased more in MCF-7 cells than in MRC-5 cells.Conclusion: Cholera toxin B subunit induced significant cell death in MCF-7, butit could be well tolerated in MRC-5. Therefore, cholera toxin B subunit can besuggested as an anti-cancer agent.

  18. Hypercapnic acidosis modulates inflammation, lung mechanics, and edema in the isolated perfused lung.

    Science.gov (United States)

    De Smet, Hilde R; Bersten, Andrew D; Barr, Heather A; Doyle, Ian R

    2007-12-01

    Low tidal volume (V(T)) ventilation strategies may be associated with permissive hypercapnia, which has been shown by ex vivo and in vivo studies to have protective effects. We hypothesized that hypercapnic acidosis may be synergistic with low V(T) ventilation; therefore, we studied the effects of hypercapnia and V(T) on unstimulated and lipopolysaccharide-stimulated isolated perfused lungs. Isolated perfused rat lungs were ventilated for 2 hours with low (7 mL/kg) or moderately high (20 mL/kg) V(T) and 5% or 20% CO(2), with lipopolysaccharide or saline added to the perfusate. Hypercapnia resulted in reduced pulmonary edema, lung stiffness, tumor necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6) in the lavage and perfusate. The moderately high V(T) did not cause lung injury but increased lavage IL-6 and perfusate IL-6 as well as TNF-alpha. Pulmonary edema and respiratory mechanics improved, possibly as a result of a stretch-induced increase in surfactant turnover. Lipopolysaccharide did not induce significant lung injury. We conclude that hypercapnia exerts a protective effect by modulating inflammation, lung mechanics, and edema. The moderately high V(T) used in this study stimulated inflammation but paradoxically improved edema and lung mechanics with an associated increase in surfactant release.

  19. Co-expression of podoplanin and fibroblast growth factor 1 predicts poor prognosis in patients with lung squamous cell carcinoma.

    Science.gov (United States)

    Li, Juan; Chen, Han; Li, Xiaoqing; Wang, Linlin; Gao, Aiqin; Zhang, Pei; Lin, Wenli; Gao, Wei; Yang, Dong; Guo, Xiaosun; Liu, Jie; Dang, Qi; Sun, Yuping

    2017-08-01

    Podoplanin and fibroblast growth factor (FGF) 1 have been detected more frequently in lung squamous cell carcinoma (SQCC) compared with lung adenocarcinoma. Furthermore, it has been previous demonstrated that FGF1 is located on the edge of tumor nests in certain lung SQCC sections, which resembles the characteristic expression pattern of podoplanin. Podoplanin and FGF1 have roles in lymphangiogenesis and angiogenesis. Based on their consistently specific expression in lung SQCC and similar localization patterns, the present study aimed to investigate whether the expression of podoplanin in tumor cells is correlated with FGF1 expression in lung SQCC and whether their co‑expression has clinicopathological significance, particularly for lymphangiogenesis/angiogenesis. The correlation between podoplanin and FGF1 expression in tumor cells of 82 lung SQCC cases was investigated by immunohistochemical staining and the association between the co‑expression of podoplanin and FGF1, and clinicopathological factors such as microvessel density (MVD), was examined in these samples. In addition, the prognostic value of co‑expression of podoplanin and FGF1 in tumor cells was determined, and the regulation of FGF1 expression and angiogenesis by podoplanin was examined in vitro in a human lung SQCC cell line. Immunohistochemical analysis demonstrated that there was a significant correlation between podoplanin and FGF1 expression in lung SQCC tumor cells (R=0.591; P<0.0001). Co‑expression of podoplanin and FGF1 was significantly associated with larger primary tumor size, advanced TNM stage and higher intratumoral MVD. Survival analysis demonstrated that cases with podoplanin and FGF1 double‑positive staining had a significantly lower survival rate compared with cases with double‑negative staining. In vitro experiments revealed that podoplanin regulated FGF1 expression and affected tube formation of human umbilical vein endothelial cells. Combined, the results

  20. Reproducible isolation of lymph node stromal cells reveals site-dependent differences in fibroblastic reticular cells

    Directory of Open Access Journals (Sweden)

    Anne L Fletcher

    2011-09-01

    Full Text Available Within lymph nodes, non-hematopoietic stromal cells organize and interact with leukocytes in an immunologically important manner. In addition to organizing T and B cell segregation and expressing lymphocyte survival factors, several recent studies have shown that lymph node stromal cells shape the naïve T cell repertoire, expressing self-antigens which delete self-reactive T cells in a unique and non-redundant fashion. A fundamental role in peripheral tolerance, in addition to an otherwise extensive functional portfolio, necessitates closer study of lymph node stromal cell subsets using modern immunological techniques; however this has not routinely been possible in the field, due to difficulties reproducibly isolating these rare subsets. Techniques were therefore developed for successful ex vivo and in vitro manipulation and characterization of lymph node stroma. Here we discuss and validate these techniques in mice and humans, and apply them to address several unanswered questions regarding lymph node composition. We explored the steady-state stromal composition of lymph nodes isolated from mice and humans, and found that marginal reticular cells and lymphatic endothelial cells required lymphocytes for their normal maturation in mice. We also report alterations in the proportion and number of fibroblastic reticular cells (FRCs between skin-draining and mesenteric lymph nodes. Similarly, transcriptional profiling of FRCs revealed changes in cytokine production from these sites. Together, these methods permit highly reproducible stromal cell isolation, sorting, and culture.

  1. Reproducible isolation of lymph node stromal cells reveals site-dependent differences in fibroblastic reticular cells.

    Science.gov (United States)

    Fletcher, Anne L; Malhotra, Deepali; Acton, Sophie E; Lukacs-Kornek, Veronika; Bellemare-Pelletier, Angelique; Curry, Mark; Armant, Myriam; Turley, Shannon J

    2011-01-01

    Within lymph nodes, non-hematopoietic stromal cells organize and interact with leukocytes in an immunologically important manner. In addition to organizing T and B cell segregation and expressing lymphocyte survival factors, several recent studies have shown that lymph node stromal cells shape the naïve T cell repertoire, expressing self-antigens which delete self-reactive T cells in a unique and non-redundant fashion. A fundamental role in peripheral tolerance, in addition to an otherwise extensive functional portfolio, necessitates closer study of lymph node stromal cell subsets using modern immunological techniques; however this has not routinely been possible in the field, due to difficulties reproducibly isolating these rare subsets. Techniques were therefore developed for successful ex vivo and in vitro manipulation and characterization of lymph node stroma. Here we discuss and validate these techniques in mice and humans, and apply them to address several unanswered questions regarding lymph node composition. We explored the steady-state stromal composition of lymph nodes isolated from mice and humans, and found that marginal reticular cells and lymphatic endothelial cells required lymphocytes for their normal maturation in mice. We also report alterations in the proportion and number of fibroblastic reticular cells (FRCs) between skin-draining and mesenteric lymph nodes. Similarly, transcriptional profiling of FRCs revealed changes in cytokine production from these sites. Together, these methods permit highly reproducible stromal cell isolation, sorting, and culture.

  2. 99Tcm-N(NOEt2 Uptake Kinetics Difference among KMB17 Human Embryonic Lung Diploid Fibroblast and Different Human Lung Cancer Cells

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    Wei JIA

    2010-04-01

    Full Text Available Background and objective PET/CT imaging is expensive, so searching the tumor imaging agent for SPECT/CT is necessary. 99Tcm-N(NOEt2 [bis (N-ethoxy-N-ethyl dithiocarbamato nitrido99Tcm (V] can be uptaken by lung cancer cells and other cells alike. The aim of this study is to evaluate the distinctive value in lung tumor with 99Tcm-N(NOEt2, the difference in its uptake kinetics in human embryonic lung diploid fibroblasts KMB17 and several kinds of lung cancer cells lines. Methods Firstly, six different cell culture medium which contained YTMLC Gejiu human lung squamous carcinoma cell, SPC-A1 human lung adenocarcinoma cell, AGZY low metastatic human lung adenocarcinoma, 973 high metastatic human lung adenocarcinoma cell, GLC-82 Gejiu human lung adenocarcinoma cell, and KMB17 human embryonic lung diploid fibroblast, respectively with equal cell density of 1×106/mL and the same volume were prepared; secondly, the same radioactive dose of 99Tcm-N(NOEt2 was added into each sample and then 300 μL mixed sample was taken out respectively and cultured in 37 oC culture box; Finally, 5 min, 15 min, 30 min, 45 min, 60 min, 75 min, 90 min after cultivation, centrifuged each cultured sample and determined the intracellular radiocounts of each sample, calculated each cell sample’s uptake rate of 99Tcm-N(NOEt2 at different time. Results Statistical difference was found among six cell samples, and the uptake rate sequence from high to low is 973 and SPC-A1>YTMLC>GLC-82>AGZY>KMB17 respectively; furthermore, 30 min-45 min after culture, the uptake rate reached stability, and the 45 min uptake rate of each sample was higher than its 96.7% uptake peak. Conclusion Based on the results above mentioned, it is supposed that there are discriminative clinical value when using 99Tcm-N(NOEt2 as a tumor targeting imaging agent, and 30 min or so after injection may be the best imaging time in the early imaging stage.

  3. Gallic Acid Induces a Reactive Oxygen Species-Provoked c-Jun NH2-Terminal Kinase-Dependent Apoptosis in Lung Fibroblasts

    Science.gov (United States)

    Chen, Chiu-Yuan; Chen, Kun-Chieh; Yang, Tsung-Ying; Liu, Hsiang-Chun; Hsu, Shih-Lan

    2013-01-01

    Idiopathic pulmonary fibrosis is a chronic lung disorder characterized by fibroblasts proliferation and extracellular matrix accumulation. Induction of fibroblast apoptosis therefore plays a crucial role in the resolution of this disease. Gallic acid (3,4,5-trihydroxybenzoic acid), a common botanic phenolic compound, has been reported to induce apoptosis in tumor cell lines and renal fibroblasts. The present study was undertaken to examine the role of mitogen-activated protein kinases (MAPKs) in lung fibroblasts apoptosis induced by gallic acid. We found that treatment with gallic acid resulted in activation of c-Jun NH2-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and protein kinase B (PKB, Akt), but not p38MAPK, in mouse lung fibroblasts. Inhibition of JNK using pharmacologic inhibitor (SP600125) and genetic knockdown (JNK specific siRNA) significantly inhibited p53 accumulation, reduced PUMA and Fas expression, and abolished apoptosis induced by gallic acid. Moreover, treatment with antioxidants (vitamin C, N-acetyl cysteine, and catalase) effectively diminished gallic acid-induced hydrogen peroxide production, JNK and p53 activation, and cell death. These observations imply that gallic acid-mediated hydrogen peroxide formation acts as an initiator of JNK signaling pathways, leading to p53 activation and apoptosis in mouse lung fibroblasts. PMID:23533505

  4. Combination of roflumilast with a beta-2 adrenergic receptor agonist inhibits proinflammatory and profibrotic mediator release from human lung fibroblasts

    Directory of Open Access Journals (Sweden)

    Tannheimer Stacey L

    2012-03-01

    Full Text Available Abstract Background Small airway narrowing is an important pathology which impacts lung function in chronic obstructive pulmonary disease (COPD. The accumulation of fibroblasts and myofibroblasts contribute to inflammation, remodeling and fibrosis by production and release of mediators such as cytokines, profibrotic factors and extracellular matrix proteins. This study investigated the effects of the phosphodiesterase 4 inhibitor roflumilast, combined with the long acting β2 adrenergic agonist indacaterol, both approved therapeutics for COPD, on fibroblast functions that contribute to inflammation and airway fibrosis. Methods The effects of roflumilast and indacaterol treatment were characterized on transforming growth factor β1 (TGFβ1-treated normal human lung fibroblasts (NHLF. NHLF were evaluated for expression of the profibrotic mediators endothelin-1 (ET-1 and connective tissue growth factor (CTGF, expression of the myofibroblast marker alpha smooth muscle actin, and fibronectin (FN secretion. Tumor necrosis factor-α (TNF-α was used to induce secretion of chemokine C-X-C motif ligand 10 (CXCL10, chemokine C-C motif ligand 5 (CCL5 and granulocyte macrophage colony-stimulating factor (GM-CSF from NHLF and drug inhibition was assessed. Results Evaluation of roflumilast (1-10 μM showed no significant inhibition alone on TGFβ1-induced ET-1 and CTGF mRNA transcripts, ET-1 and FN protein production, alpha smooth muscle expression, or TNF-α-induced secretion of CXCL10, CCL5 and GM-CSF. A concentration-dependent inhibition of ET-1 and CTGF was shown with indacaterol treatment, and a submaximal concentration was chosen for combination studies. When indacaterol (0.1 nM was added to roflumilast, significant inhibition was seen on all inflammatory and fibrotic mediators evaluated, which was superior to the inhibition seen with either drug alone. Roflumilast plus indacaterol combination treatment resulted in significantly elevated phosphorylation

  5. TNFSF14 (LIGHT Exhibits Inflammatory Activities in Lung Fibroblasts Complementary to IL-13 and TGF-β

    Directory of Open Access Journals (Sweden)

    Ricardo da Silva Antunes

    2018-03-01

    Full Text Available The cytokine TNFSF14 [homologous to Lymphotoxin, exhibits Inducible expression and competes with HSV Glycoprotein D for binding to HVEM, a receptor expressed on T lymphocytes (LIGHT] has been shown in mouse models to be important for development of lung tissue remodeling that is characteristic of asthma, idiopathic pulmonary fibrosis (IPF, and systemic sclerosis (SSc. However, its cellular targets are not fully delineated. In the present report, we show that LTβR and HVEM, the receptors for LIGHT, are constitutively expressed in primary human lung fibroblasts (HLFs. We asked whether LIGHT could promote inflammatory and remodeling-relevant activity in HLFs and how this was similar to, or distinct from, IL-13 or TGF-β, two cytokines strongly implicated in the pathogenesis of asthma, IPF, and SSc. Accumulation of myofibroblasts expressing alpha smooth muscle actin is a feature of lung inflammatory diseases. LIGHT promoted cell cycle progression and proliferation of HLFs, but not alpha smooth muscle actin expression. In contrast, TGF-β upregulated alpha smooth muscle actin but did not drive their proliferation. LIGHT also increased the gene or protein expression of a number of proinflammatory mediators, including ICAM-1 and VCAM-1, IL-6 and GM-CSF, the chemokines CCL5 and 20, and CXCL5, 11, and 12, and lung remodeling-associated proteinases MMP-9 and ADAM8. These were dependent on LTβR but not HVEM. LIGHT displayed overlapping and synergistic activities with IL-13 for a number of the activities, but LIGHT additionally enhanced the gene expression of several molecules, including the innate cytokines IL-33 and TSLP, which were not upregulated by IL-13. Our results highlight the varied and pleiotropic effects of LIGHT in HLFs. LIGHT might then be a therapeutic target for modulation of inflammation and remodeling associated with asthma and other similar diseases of the lung that involve fibroblasts.

  6. Bambuterol: uptake and metabolism in guinea pig isolated lungs

    International Nuclear Information System (INIS)

    Ryrfeldt, A.; Nilsson, E.; Tunek, A.; Svensson, L.A.

    1988-01-01

    The lung uptake and biotransformation of 3 H-bambuterol, a prodrug to terbutaline, were studied using isolated perfused and ventilated guinea pig lungs. 14 C-Sucrose was used as an extracellular marker. The lung uptake of bambuterol was significantly (0.05 greater than or equal to P greater than or equal to 0.001) higher than that found for sucrose in single-pass perfusion experiments. High-performance liquid chromatographic (HPLC) analysis showed that 95.6 +/- 3.6% of the effluent 3 H radioactivity was attributable to bambuterol. In recirculating experiments (120 min) the lung biotransformation of 3 H-bambuterol (8.5 pmol/ml) was studied. Both oxidative and hydrolytic metabolism took place. The dominating metabolites were hydroxylated bambuterol and the monocarbamate derivative which is a product of hydrolysis of bambuterol. Traces of terbutaline were also formed. The results show that bambuterol has a certain affinity to lung tissue and that the drug is, to some extent, biotransformed in the guinea pig lung

  7. Isolation and Characterization of Human Lung Lymphatic Endothelial Cells

    Science.gov (United States)

    Lorusso, Bruno; Falco, Angela; Madeddu, Denise; Frati, Caterina; Cavalli, Stefano; Graiani, Gallia; Gervasi, Andrea; Rinaldi, Laura; Lagrasta, Costanza; Maselli, Davide; Gnetti, Letizia; Silini, Enrico M.; Quaini, Eugenio; Ampollini, Luca; Carbognani, Paolo; Quaini, Federico

    2015-01-01

    Characterization of lymphatic endothelial cells from the respiratory system may be crucial to investigate the role of the lymphatic system in the normal and diseased lung. We describe a simple and inexpensive method to harvest, isolate, and expand lymphatic endothelial cells from the human lung (HL-LECs). Fifty-five samples of healthy lung selected from patients undergoing lobectomy were studied. A two-step purification tool, based on paramagnetic sorting with monoclonal antibodies to CD31 and Podoplanin, was employed to select a pure population of HL-LECs. The purity of HL-LECs was assessed by morphologic criteria, immunocytochemistry, flow cytometry, and functional assays. Interestingly, these cells retain in vitro several receptor tyrosine kinases (RTKs) implicated in cell survival and proliferation. HL-LECs represent a clinically relevant cellular substrate to study lymphatic biology, lymphoangiogenesis, interaction with microbial agents, wound healing, and anticancer therapy. PMID:26137493

  8. Sterols of Pneumocystis carinii hominis organisms isolated from human lungs

    DEFF Research Database (Denmark)

    Kaneshiro, E S; Amit, Z; Chandra, Jan Suresh

    1999-01-01

    in conjunction with analyses of chemically synthesized authentic standards. The sterol composition of isolated P. carinii hominis organisms has yet to be reported. If P. carinii from animal models is to be used for identifying potential drug targets and for developing chemotherapeutic approaches to clear human...... infections, it is important to determine whether the 24-alkylsterols of organisms found in rats are also present in organisms in humans. In the present study, sterol analyses of P. carinii hominis organisms isolated from cryopreserved human P. carinii-infected lungs and from bronchoalveolar lavage fluid were...

  9. Lung fibroblasts accelerate wound closure in human alveolar epithelial cells through hepatocyte growth factor/c-Met signaling.

    Science.gov (United States)

    Ito, Yoko; Correll, Kelly; Schiel, John A; Finigan, Jay H; Prekeris, Rytis; Mason, Robert J

    2014-07-01

    There are 190,600 cases of acute lung injury/acute respiratory distress syndrome (ALI/ARDS) each year in the United States, and the incidence and mortality of ALI/ARDS increase dramatically with age. Patients with ALI/ARDS have alveolar epithelial injury, which may be worsened by high-pressure mechanical ventilation. Alveolar type II (ATII) cells are the progenitor cells for the alveolar epithelium and are required to reestablish the alveolar epithelium during the recovery process from ALI/ARDS. Lung fibroblasts (FBs) migrate and proliferate early after lung injury and likely are an important source of growth factors for epithelial repair. However, how lung FBs affect epithelial wound healing in the human adult lung has not been investigated in detail. Hepatocyte growth factor (HGF) is known to be released mainly from FBs and to stimulate both migration and proliferation of primary rat ATII cells. HGF is also increased in lung tissue, bronchoalveolar lavage fluid, and serum in patients with ALI/ARDS. Therefore, we hypothesized that HGF secreted by FBs would enhance wound closure in alveolar epithelial cells (AECs). Wound closure was measured using a scratch wound-healing assay in primary human AEC monolayers and in a coculture system with FBs. We found that wound closure was accelerated by FBs mainly through HGF/c-Met signaling. HGF also restored impaired wound healing in AECs from the elderly subjects and after exposure to cyclic stretch. We conclude that HGF is the critical factor released from FBs to close wounds in human AEC monolayers and suggest that HGF is a potential strategy for hastening alveolar repair in patients with ALI/ARDS. Copyright © 2014 the American Physiological Society.

  10. Podoplanin expression in cancer-associated fibroblasts predicts unfavourable prognosis in patients with pathological stage IA lung adenocarcinoma.

    Science.gov (United States)

    Kubouchi, Yasuaki; Yurugi, Yohei; Wakahara, Makoto; Sakabe, Tomohiko; Haruki, Tomohiro; Nosaka, Kanae; Miwa, Ken; Araki, Kunio; Taniguchi, Yuji; Shiomi, Tatsushi; Nakamura, Hiroshige; Umekita, Yoshihisa

    2018-02-01

    Podoplanin expression in cancer-associated fibroblasts (CAFs) has been proposed as an unfavourable indicator in squamous cell carcinoma of the lung, but little is known about its clinical significance in early-stage lung adenocarcinoma. We evaluated the prognostic impact of podoplanin expression in patients with pathological stage (p-stage) IA lung adenocarcinoma as categorised by the 8th edition of the tumour-node-metastasis classification for lung cancer. Immunohistochemical analyses using anti-podoplanin antibody were performed on resected specimens from 158 patients with p-stage IA lung adenocarcinoma. When more than 10% of cancer cells or CAFs showed immunoreactivity with podoplanin, the specimens were classified as podoplanin-positive. Podoplanin-positive status in cancer cells (n = 8) was not correlated with clinicopathological factors or with patient prognosis. Podoplanin-positive status in CAFs (n = 41) was correlated significantly with poorer tumour differentiation (P < 0.001), the presence of lymphatic invasion (P < 0.001) and high-grade (solid and/or micropapillary) components constituting ≥1% of the entire tumour (P < 0.001). The log-rank test showed that podoplanin-positive status in CAFs was associated significantly with shorter disease-free survival (DFS) (P < 0.001) and disease-specific survival (P = 0.015). In Cox's multivariate analysis, podoplanin-positive status in CAFs had the most significant effect on shorter DFS [hazard ratio (HR) = 4.411, P = 0.004], followed by the presence of high-grade components (HR = 3.581, P = 0.013). Podoplanin expression in CAFs could be an independent predictor of increased risk of recurrence in patients with p-stage IA lung adenocarcinoma. © 2017 John Wiley & Sons Ltd.

  11. Senescence-associated microRNAs target cell cycle regulatory genes in normal human lung fibroblasts.

    Science.gov (United States)

    Markopoulos, Georgios S; Roupakia, Eugenia; Tokamani, Maria; Vartholomatos, George; Tzavaras, Theodore; Hatziapostolou, Maria; Fackelmayer, Frank O; Sandaltzopoulos, Raphael; Polytarchou, Christos; Kolettas, Evangelos

    2017-10-01

    Senescence recapitulates the ageing process at the cell level. A senescent cell stops dividing and exits the cell cycle. MicroRNAs (miRNAs) acting as master regulators of transcription, have been implicated in senescence. In the current study we investigated and compared the expression of miRNAs in young versus senescent human fibroblasts (HDFs), and analysed the role of mRNAs expressed in replicative senescent HFL-1 HDFs. Cell cycle analysis confirmed that HDFs accumulated in G 1 /S cell cycle phase. Nanostring analysis of isolated miRNAs from young and senescent HFL-1 showed that a distinct set of 15 miRNAs were significantly up-regulated in senescent cells including hsa-let-7d-5p, hsa-let-7e-5p, hsa-miR-23a-3p, hsa-miR-34a-5p, hsa-miR-122-5p, hsa-miR-125a-3p, hsa-miR-125a-5p, hsa-miR-125b-5p, hsa-miR-181a-5p, hsa-miR-221-3p, hsa-miR-222-3p, hsa-miR-503-5p, hsa-miR-574-3p, hsa-miR-574-5p and hsa-miR-4454. Importantly, pathway analysis of miRNA target genes down-regulated during replicative senescence in a public RNA-seq data set revealed a significant high number of genes regulating cell cycle progression, both G 1 /S and G 2 /M cell cycle phase transitions and telomere maintenance. The reduced expression of selected miRNA targets, upon replicative and oxidative-stress induced senescence, such as the cell cycle effectors E2F1, CcnE, Cdc6, CcnB1 and Cdc25C was verified at the protein and/or RNA levels. Induction of G1/S cell cycle phase arrest and down-regulation of cell cycle effectors correlated with the up-regulation of miR-221 upon both replicative and oxidative stress-induced senescence. Transient expression of miR-221/222 in HDFs promoted the accumulation of HDFs in G1/S cell cycle phase. We propose that miRNAs up-regulated during replicative senescence may act in concert to induce cell cycle phase arrest and telomere erosion, establishing a senescent phenotype. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. P. aeruginosa in the paranasal sinuses and transplanted lungs have similar adaptive mutations as isolates from chronically infected CF lungs

    DEFF Research Database (Denmark)

    Ciofu, Oana; Johansen, Helle Krogh; Aanaes, Kasper

    2013-01-01

    BACKGROUND: Pseudomonas aeruginosa cells are present as biofilms in the paranasal sinuses and the lungs of chronically infected cystic fibrosis (CF) patients. Since different inflammatory responses and selective antibiotic pressures are acting in the sinuses compared with the lungs, we compared...... the adaptive profiles of mucoid and non-mucoid isolates from the two locations. METHODS: We studied the genetic basis of phenotypic diversification and gene expression profiles in sequential lung and sinus P. aeruginosa isolates from four chronically infected CF patients, including pre- and post-lung...... transplantation isolates. RESULTS: The same phenotypes caused by similar mutations and similar gene expression profiles were found in mucoid and non-mucoid isolates from the paranasal sinuses and from the lungs before and after transplantation. CONCLUSION: Bilateral exchange of P. aeruginosa isolates between...

  13. Interleukin-1β attenuates myofibroblast formation and extracellular matrix production in dermal and lung fibroblasts exposed to transforming growth factor-β1.

    Directory of Open Access Journals (Sweden)

    Masum M Mia

    Full Text Available One of the most potent pro-fibrotic cytokines is transforming growth factor (TGFβ. TGFβ is involved in the activation of fibroblasts into myofibroblasts, resulting in the hallmark of fibrosis: the pathological accumulation of collagen. Interleukin-1β (IL1β can influence the severity of fibrosis, however much less is known about the direct effects on fibroblasts. Using lung and dermal fibroblasts, we have investigated the effects of IL1β, TGFβ1, and IL1β in combination with TGFβ1 on myofibroblast formation, collagen synthesis and collagen modification (including prolyl hydroxylase, lysyl hydroxylase and lysyl oxidase, and matrix metalloproteinases (MMPs. We found that IL1β alone has no obvious pro-fibrotic effect on fibroblasts. However, IL1β is able to inhibit the TGFβ1-induced myofibroblast formation as well as collagen synthesis. Glioma-associated oncogene homolog 1 (GLI1, the Hedgehog transcription factor that is involved in the transformation of fibroblasts into myofibroblasts is upregulated by TGFβ1. The addition of IL1β reduced the expression of GLI1 and thereby also indirectly inhibits myofibroblast formation. Other potentially anti-fibrotic effects of IL1β that were observed are the increased levels of MMP1, -2, -9 and -14 produced by fibroblasts exposed to TGFβ1/IL1β in comparison with fibroblasts exposed to TGFβ1 alone. In addition, IL1β decreased the TGFβ1-induced upregulation of lysyl oxidase, an enzyme involved in collagen cross-linking. Furthermore, we found that lung and dermal fibroblasts do not always behave identically towards IL1β. Suppression of COL1A1 by IL1β in the presence of TGFβ1 is more pronounced in lung fibroblasts compared to dermal fibroblasts, whereas a higher upregulation of MMP1 is seen in dermal fibroblasts. The role of IL1β in fibrosis should be reconsidered, and the differences in phenotypical properties of fibroblasts derived from different organs should be taken into account in future

  14. Correlation of Hypoxia and Pro-senescence Protein Expression in Green Sea Turtle (Chelonia mydas Lung Epithelial and Dermal Fibroblast Cell Culture

    Directory of Open Access Journals (Sweden)

    Anggraini Barlian

    2018-03-01

    Full Text Available Recent studies have shown hypoxia-induced gene expression correlated with cellular senescence. HIF-1α (hypoxia-inducible factor 1-alpha, p53, and pRB were induced under hypoxia and correlated with cellular senescence. The localization and expression of HIF-1α, p53, and pRB in Chelonia mydas lung epithelial and dermal fibroblast cell cultures were analyzed under normoxic and hypoxic conditions (at 4 and 24 hours. Human dermal fibroblast was used for comparison purposes. Protein localization was analyzed with immunocytochemistry, while protein expression was analyzed with the Western blot and enhanced chemiluminescence (ECL method. HIF-1α, p53, and pRB were localized in the nuclei of the C. mydas cell cultures treated with hypoxia. The C. mydas lung epithelial cell cultures had a higher increase of HIF-1α expression than the human dermal fibroblast cell culture. The hypoxic conditions did not affect p53 expression significantly in C. mydas lung epithelial and dermal fibroblast cell cultures. Meanwhile, pRB expression changed significantly under hypoxia in the C. mydas dermal fibroblast cells. Expression of p53 and pRB in the human cell cultures was higher than in the C. mydas cell cultures. This research suggests that C. mydas and human cell cultures have different pro-senescence protein expression responses under hypoxic conditions.

  15. Human lung fibroblast-derived matrix facilitates vascular morphogenesis in 3D environment and enhances skin wound healing.

    Science.gov (United States)

    Du, Ping; Suhaeri, Muhammad; Ha, Sang Su; Oh, Seung Ja; Kim, Sang-Heon; Park, Kwideok

    2017-05-01

    Extracellular matrix (ECM) is crucial to many aspects of vascular morphogenesis and maintenance of vasculature function. Currently the recapitulation of angiogenic ECM microenvironment is still challenging, due mainly to its diverse components and complex organization. Here we investigate the angiogenic potential of human lung fibroblast-derived matrix (hFDM) in creating a three-dimensional (3D) vascular construct. hFDM was obtained via decellularization of in vitro cultured human lung fibroblasts and analyzed via immunofluorescence staining and ELISA, which detect multiple ECM macromolecules and angiogenic growth factors (GFs). Human umbilical vein endothelial cells (HUVECs) morphology was more elongated and better proliferative on hFDM than on gelatin-coated substrate. To prepare 3D construct, hFDM is collected, quantitatively analyzed, and incorporated in collagen hydrogel (Col) with HUVECs. Capillary-like structure (CLS) formation at 7day was significantly better with the groups containing higher doses of hFDM compared to the Col group (control). Moreover, the group (Col/hFDM/GFs) with both hFDM and angiogenic GFs (VEGF, bFGF, SDF-1) showed the synergistic activity on CLS formation and found much larger capillary lumen diameters with time. Further analysis of hFDM via angiogenesis antibody array kit reveals abundant biochemical cues, such as angiogenesis-related cytokines, GFs, and proteolytic enzymes. Significantly up-regulated expression of VE-cadherin and ECM-specific integrin subunits was also noticed in Col/hFDM/GFs. In addition, transplantation of Col/hFMD/GFs with HUVECs in skin wound model presents more effective re-epithelialization, many regenerated hair follicles, better transplanted cells viability, and advanced neovascularization. We believe that current system is a very promising platform for 3D vasculature construction in vitro and for cell delivery toward therapeutic applications in vivo. Functional 3D vasculature construction in vitro is still

  16. Fibroblast Activation Protein (FAP) Accelerates Collagen Degradation and Clearance from Lungs in Mice

    DEFF Research Database (Denmark)

    Fan, Ming-Hui; Zhu, Qiang; Li, Hui-Hua

    2016-01-01

    , intratracheal bleomycin instillation and thoracic irradiation, we find increased mortality and increased lung fibrosis in FAP-deficient mice compared with wild-type mice. Lung extracellular matrix analysis reveals accumulation of intermediate-sized collagen fragments in FAP-deficient mouse lungs, consistent...... within vitrostudies showing that FAP mediates ordered proteolytic processing of matrix metalloproteinase (MMP)-derived collagen cleavage products. FAP-mediated collagen processing leads to increased collagen internalization without altering expression of the endocytic collagen receptor, Endo180....... Pharmacologic FAP inhibition decreases collagen internalization as expected. Conversely, restoration of FAP expression in the lungs of FAP-deficient mice decreases lung hydroxyproline content after intratracheal bleomycin to levels comparable with that of wild-type controls. Our findings indicate that FAP...

  17. Nonmuscle myosin IIA and IIB differentially contribute to intrinsic and directed migration of human embryonic lung fibroblasts.

    Science.gov (United States)

    Kuragano, Masahiro; Murakami, Yota; Takahashi, Masayuki

    2018-03-25

    Nonmuscle myosin II (NMII) plays an essential role in directional cell migration. In this study, we investigated the roles of NMII isoforms (NMIIA and NMIIB) in the migration of human embryonic lung fibroblasts, which exhibit directionally persistent migration in an intrinsic manner. NMIIA-knockdown (KD) cells migrated unsteadily, but their direction of migration was approximately maintained. By contrast, NMIIB-KD cells occasionally reversed their direction of migration. Lamellipodium-like protrusions formed in the posterior region of NMIIB-KD cells prior to reversal of the migration direction. Moreover, NMIIB KD led to elongation of the posterior region in migrating cells, probably due to the lack of load-bearing stress fibers in this area. These results suggest that NMIIA plays a role in steering migration by maintaining stable protrusions in the anterior region, whereas NMIIB plays a role in maintenance of front-rear polarity by preventing aberrant protrusion formation in the posterior region. These distinct functions of NMIIA and NMIIB might promote intrinsic and directed migration of normal human fibroblasts. Copyright © 2018 Elsevier Inc. All rights reserved.

  18. Concentration of (+/-)-propranolol in isolated, perfused lungs of rat.

    Science.gov (United States)

    Dollery, C T; Junod, A F

    1976-01-01

    1 The metabolism and the accumulation of (+/-)-propranolol have been studied in isolated lungs of the rat, perfused with an artificial medium. 2 Little or no metabolism took place during the perfusion periods (up to 10 minutes). 3 Accumulation was observed with high tissue/medium ratios for substrate concentrations of 0.2 muM to 1 mM; there was evidence for saturability, but no real plateau could be seen. The presence of two binding sites with different affinities was established. 4 Cold greatly inhibited the accumulation process at low substrate concentrations, but had no effect at 1 mM propranolol. 5 Inhibition of accumulation was measured in the presence of imipramine, desmethylimipramine, nortryptiline, chlorpromazine and of Na+-free medium. Cocaine, 5-hydroxytryptamine and noradrenaline had no effect. Lidocaine enhanced the accumulation process. Release of previously bound propranolol was accelerated in the presence of propranolol and imipramine, unaffected by a Na+-free medium and decreased by cold and by lidocaine. 6 Experiments on lung tissue slices yielded qualitatively similar results to those obtained with perfused lungs. Ouabain and KCN had no or little effect on propranolol accumulation. PMID:1276542

  19. Expression of TGF-β3 in Isolated Fibroblasts from Foreskin

    Directory of Open Access Journals (Sweden)

    Mahnaz Mahmoudi Rad

    2015-05-01

    Full Text Available Background: The multifunctional transforming growth factor beta (TGF-β is a glycoprotein that exists in three isoforms. TGF-β3 expression increases in fetal wound healing and reduces fibronectin and collagen I and III deposition, and also improves the architecture of the neodermis which is a combination of blood vessels and connective tissue during wound healing. Fibroblasts are key cells in the wound healing process. TGF-β3 plays a critical role in scar-free wound healing and fibroblast actions in the wound healing process. The aim of this study was to express the TGF-β3 gene (tgf-b3 in human foreskin fibroblasts (HFF’s. Methods: We obtained HFF’s from a newborn and a primary fibroblast culture was prepared. The cells were transfected with TGF-β3-pCMV6-XL5 plasmid DNA by both lipofection and electroporation. Expression of TGF-β3 was measured by enzyme-linked immunosorbent assay (ELISA. Results: The highest TGF-β3 expression (8.3-fold greater than control was obtained by lipofection after 72 hours using 3 μl of transfection reagent. Expression was 1.4-fold greater than control by electroporation. Conclusions: In this study, we successfully increased TGF-β3 expression in primary fibroblast cells. In the future, grafting these transfected fibroblasts onto wounds can help the healing process without scarring.

  20. Static inflation attenuates ischemia/reperfusion injury in an isolated rat lung in situ.

    Science.gov (United States)

    Kao, Shang Jyh; Wang, David; Yeh, Diana Yu-Wung; Hsu, Kang; Hsu, Yung Hsiang; Chen, Hsing I

    2004-08-01

    Ischemia (I)/reperfusion (R) lung injury is an important clinical issue in lung transplantation. In the present study, we observed the effects of lung static inflation, different perfusates, and ventilatory gas with nitrogen or oxygen on the I/R-induced pulmonary damage. A total of 96 male Sprague-Dawley rats were used. The lung was isolated in situ. In an isolated lung, the capillary filtration coefficient (Kfc), lung weight gain (LWG), lung weight (LW)/body weight (BW) ratio, and protein concentration in BAL fluid (PCBAL) were measured or calculated to evaluate the degree of lung injury. Histologic examinations with hematoxylin-eosin staining were performed. I/R caused lung injury, as reflected by increases in Kfc, LWG, LW/BW, and PCBAL. The histopathologic picture revealed the presence of hyaline membrane formation and the infiltration of inflammatory cells. These values were significantly attenuated by static lung inflation. The I/R lung damage appeared to be less in the lung perfused with whole blood than in the lung perfused with an isotonic solution. Therapy with ventilatory air (ie, nitrogen or oxygen) did not alter the I/R lung damage. The data suggest that lung inflation is protective to I/R injury, irrespective of the type of ventilatory air used for treatment. The preservation of the lung for transplantation is better kept at a static inflation state and perfused with whole blood instead of an isotonic physiologic solution.

  1. Aryl hydrocarbon receptor-dependent regulation of miR-196a expression controls lung fibroblast apoptosis but not proliferation

    International Nuclear Information System (INIS)

    Hecht, Emelia; Zago, Michela; Sarill, Miles; Rico de Souza, Angela; Gomez, Alvin; Matthews, Jason; Hamid, Qutayba; Eidelman, David H.; Baglole, Carolyn J.

    2014-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor implicated in the regulation of apoptosis and proliferation. Although activation of the AhR by xenobiotics such as dioxin inhibits the cell cycle and control apoptosis, paradoxically, AhR expression also promotes cell proliferation and survival independent of exogenous ligands. The microRNA (miRNA) miR-196a has also emerged as a regulator of proliferation and apoptosis but a relationship between the AhR and miR-196a is not known. Therefore, we hypothesized that AhR-dependent regulation of endogenous miR-196a expression would promote cell survival and proliferation. Utilizing lung fibroblasts from AhR deficient (AhR −/− ) and wild-type (AhR +/+ ) mice, we show that there is ligand-independent regulation of miRNA, including low miR-196a in AhR −/− cells. Validation by qRT-PCR revealed a significant decrease in basal expression of miR-196a in AhR −/− compared to AhR +/+ cells. Exposure to AhR agonists benzo[a]pyrene (B[a]P) and FICZ as well as AhR antagonist CH-223191 decreased miR-196a expression in AhR +/+ fibroblasts concomitant with decreased AhR protein levels. There was increased proliferation only in AhR +/+ lung fibroblasts in response to serum, corresponding to a decrease in p27 KIP1 protein, a cyclin-dependent kinase inhibitor. Increasing the cellular levels of miR-196a had no effect on proliferation or expression of p27 KIP1 in AhR −/− fibroblasts but attenuated cigarette smoke-induced apoptosis. This study provides the first evidence that AhR expression is essential for the physiological regulation of cellular miRNA levels- including miR-196a. Future experiments designed to elucidate the functional relationship between the AhR and miR-196a may delineate additional novel ligand-independent roles for the AhR. - Highlights: • The AhR controls proliferation and apoptosis in lung cells. • The AhR regulates the expression of the microRNA miR-196a independent of

  2. Aryl hydrocarbon receptor-dependent regulation of miR-196a expression controls lung fibroblast apoptosis but not proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Hecht, Emelia [Department of Medicine, McGill University, Montreal, Quebec (Canada); Zago, Michela [Research Institute of the McGill University Health Centre, McGill University, Montreal, Quebec (Canada); Sarill, Miles [Department of Medicine, McGill University, Montreal, Quebec (Canada); Rico de Souza, Angela [Research Institute of the McGill University Health Centre, McGill University, Montreal, Quebec (Canada); Gomez, Alvin; Matthews, Jason [Department of Pharmacology and Toxicology, University of Toronto, Toronto, ON (Canada); Hamid, Qutayba; Eidelman, David H. [Department of Medicine, McGill University, Montreal, Quebec (Canada); Research Institute of the McGill University Health Centre, McGill University, Montreal, Quebec (Canada); Baglole, Carolyn J., E-mail: Carolyn.baglole@McGill.ca [Department of Medicine, McGill University, Montreal, Quebec (Canada); Research Institute of the McGill University Health Centre, McGill University, Montreal, Quebec (Canada)

    2014-11-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor implicated in the regulation of apoptosis and proliferation. Although activation of the AhR by xenobiotics such as dioxin inhibits the cell cycle and control apoptosis, paradoxically, AhR expression also promotes cell proliferation and survival independent of exogenous ligands. The microRNA (miRNA) miR-196a has also emerged as a regulator of proliferation and apoptosis but a relationship between the AhR and miR-196a is not known. Therefore, we hypothesized that AhR-dependent regulation of endogenous miR-196a expression would promote cell survival and proliferation. Utilizing lung fibroblasts from AhR deficient (AhR{sup −/−}) and wild-type (AhR{sup +/+}) mice, we show that there is ligand-independent regulation of miRNA, including low miR-196a in AhR{sup −/−} cells. Validation by qRT-PCR revealed a significant decrease in basal expression of miR-196a in AhR{sup −/−} compared to AhR{sup +/+} cells. Exposure to AhR agonists benzo[a]pyrene (B[a]P) and FICZ as well as AhR antagonist CH-223191 decreased miR-196a expression in AhR{sup +/+} fibroblasts concomitant with decreased AhR protein levels. There was increased proliferation only in AhR{sup +/+} lung fibroblasts in response to serum, corresponding to a decrease in p27{sup KIP1} protein, a cyclin-dependent kinase inhibitor. Increasing the cellular levels of miR-196a had no effect on proliferation or expression of p27{sup KIP1} in AhR{sup −/−} fibroblasts but attenuated cigarette smoke-induced apoptosis. This study provides the first evidence that AhR expression is essential for the physiological regulation of cellular miRNA levels- including miR-196a. Future experiments designed to elucidate the functional relationship between the AhR and miR-196a may delineate additional novel ligand-independent roles for the AhR. - Highlights: • The AhR controls proliferation and apoptosis in lung cells. • The AhR regulates the

  3. Propofol attenuates oxidant-induced acute lung injury in an isolated perfused rabbit-lung model.

    Science.gov (United States)

    Yumoto, Masato; Nishida, Osamu; Nakamura, Fujio; Katsuya, Hirotada

    2005-01-01

    Reactive oxygen species have been strongly implicated in the pathogenesis of acute lung injury (ALI). Some animal studies suggest that free radical scavengers inhibit the onset of oxidant-induced ALI. Propofol (2,6-diisopropylphenol) is chemically similar to phenol-based free radical scavengers such as the endogenous antioxidant vitamin E. Both in vivo and in vitro studies have suggested that propofol has antioxidant potential. We hypothesized that propofol may attenuate ALI by acting as a free-radical scavenger. We investigated the effects of propofol on oxidant-induced ALI induced by purine and xanthine oxidase (XO), in isolated perfused rabbit lung, in two series of experiments. In series 1, we examined the relationship between the severity of ALI and the presence of hydrogen peroxide (H2O2). In series 2, we evaluated the effects of propofol on attenuating ALI and the dose dependence of these effects. The lungs were perfused for 90 min, and we evaluated the effects on the severity of ALI by monitoring the pulmonary capillary filtration coefficient (Kfc), pulmonary arterial pressure (Ppa), and the pulmonary capillary hydrostatic pressure (Ppc). In series 1, treatment with catalase (an H2O2 scavenger) prior to the addition of purine and XO resulted in complete prevention of ALI, suggesting that H2O2 may be involved closely in the pathogenesis of ALI. In series 2, pretreatment with propofol at concentrations in excess of 0.5 mM significantly inhibited the increases in the Kfc values, and that in excess of 0.75 mM significantly inhibited the increase in the Ppa values. Propofol attenuates oxidant-induced ALI in an isolated perfused rabbit lung model, probably due to its antioxidant action.

  4. Isolated lung events following radiation for early stage breast cancer: incidence and predictors for primary lung vs metastatic breast cancer

    International Nuclear Information System (INIS)

    Van Buren, Teresa A; Harris, Jay R; Sugarbaker, David J; Schneider, Lindsey; Healey, Elizabeth A

    1995-01-01

    Purpose: 1) To define the incidence of isolated lung events in a cohort of women treated with conservative surgery (CS) and radiation therapy (RT) for early stage breast cancer. 2) Among such patients, to define the relative distribution of primary lung cancer, metastatic breast cancer, and indeterminate lesions; and to identify any predictors for a diagnosis of lung vs metastatic breast cancer. 3) To examine the cohort with respect to whether a higher than expected incidence of lung cancer is seen following breast irradiation. Materials and Methods: Between 1968 and 1986, 1865 patients with clinical stage I-II breast cancer were treated with CS and RT; the median follow-up for surviving patients is 129 months. The study population was limited to patients who developed a subsequent isolated lung event as the first site of distant disease. Isolated lung event was defined as disease limited to the thoracic cavity, without evidence of either uncontrolled local breast disease or metastatic disease elsewhere. Diagnosis of the lung event as a primary lung cancer, a metastatic breast lesion, or an indeterminate lesion was documented from the viewpoint of 1) the pathologic analysis and 2) the clinical impression at the time of the lung event. Results: Sixty six of the 1865 patients (3.5%) developed an isolated lung event. The relative distribution of the pathologic and clinical diagnoses is shown below: The 66 lung events were characterized either as a solitary pulmonary nodule (27), multiple nodules (23), pleural effusion alone (10), unknown (2), or miscellaneous other findings (4). Among the 47 patients for whom pathology was available, the diagnosis remained indeterminate for 24 (51%). For patients with a definitive pathologic diagnosis, 69% ((9(13))) of smokers had a new lung cancer compared to 20% ((2(10))) of non-smokers (p=0.036), and 67% ((10(15))) of patients with a solitary pulmonary nodule had lung cancer compared to 14% ((1(7))) for other lung presentations (p

  5. miR-199a-5p Is upregulated during fibrogenic response to tissue injury and mediates TGFbeta-induced lung fibroblast activation by targeting caveolin-1.

    Directory of Open Access Journals (Sweden)

    Christian Lacks Lino Cardenas

    Full Text Available As miRNAs are associated with normal cellular processes, deregulation of miRNAs is thought to play a causative role in many complex diseases. Nevertheless, the precise contribution of miRNAs in fibrotic lung diseases, especially the idiopathic form (IPF, remains poorly understood. Given the poor response rate of IPF patients to current therapy, new insights into the pathogenic mechanisms controlling lung fibroblasts activation, the key cell type driving the fibrogenic process, are essential to develop new therapeutic strategies for this devastating disease. To identify miRNAs with potential roles in lung fibrogenesis, we performed a genome-wide assessment of miRNA expression in lungs from two different mouse strains known for their distinct susceptibility to develop lung fibrosis after bleomycin exposure. This led to the identification of miR-199a-5p as the best miRNA candidate associated with bleomycin response. Importantly, miR-199a-5p pulmonary expression was also significantly increased in IPF patients (94 IPF versus 83 controls. In particular, levels of miR-199a-5p were selectively increased in myofibroblasts from injured mouse lungs and fibroblastic foci, a histologic feature associated with IPF. Therefore, miR-199a-5p profibrotic effects were further investigated in cultured lung fibroblasts: miR-199a-5p expression was induced upon TGFβ exposure, and ectopic expression of miR-199a-5p was sufficient to promote the pathogenic activation of pulmonary fibroblasts including proliferation, migration, invasion, and differentiation into myofibroblasts. In addition, we demonstrated that miR-199a-5p is a key effector of TGFβ signaling in lung fibroblasts by regulating CAV1, a critical mediator of pulmonary fibrosis. Remarkably, aberrant expression of miR-199a-5p was also found in unilateral ureteral obstruction mouse model of kidney fibrosis, as well as in both bile duct ligation and CCl4-induced mouse models of liver fibrosis, suggesting that

  6. Cellular uptake and cytotoxic potential of respirable bentonite particles with different quartz contents and chemical modifications in human lung fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Geh, Stefan; Rettenmeier, Albert W.; Dopp, Elke [University Hospital, Institute of Hygiene and Occupational Medicine, Essen (Germany); Yuecel, Raif [University Hospital, Institute of Cell Biology (Cancer Research), Essen (Germany); Duffin, Rodger [Institute of Environmental Health Research (IUF), Duesseldorf (Germany); University of Edinburgh, ELEGI COLT Lab, Scotland (United Kingdom); Albrecht, Catrin; Borm, Paul J.A. [Institute of Environmental Health Research (IUF), Duesseldorf (Germany); Armbruster, Lorenz [Verein fuer Technische Sicherheit und Umweltschutz e.V., Gotha (Germany); Raulf-Heimsoth, Monika; Bruening, Thomas [Research Institute for Occupational Medicine of the Institutions for Statutory Accident Insurance and Prevention (BGFA), Bochum (Germany); Hoffmann, Eik [University of Rostock, Institute of Biology, Department of Cell Biology and Biosystems Technology, Rostock (Germany)

    2006-02-01

    Considering the biological reactivity of pure quartz in lung cells, there is a strong interest to clarify the cellular effects of respirable siliceous dusts, like bentonites. In the present study, we investigated the cellular uptake and the cytotoxic potential of bentonite particles (Oe< 10 {mu}m) with an {alpha}-quartz content of up to 6% and different chemical modifications (activation: alkaline, acidic, organic) in human lung fibroblasts (IMR90). Additionally, the ability of the particles to induce apoptosis in IMR90-cells and the hemolytic activity was tested. All bentonite samples were tested for endotoxins with the in vitro-Pyrogen test and were found to be negative. Cellular uptake of particles by IMR90-cells was studied by transmission electron microscopy (TEM). Cytotoxicity was analyzed in IMR90-cells by determination of viable cells using flow cytometry and by measuring of the cell respiratory activity. Induced apoptotic cells were detected by AnnexinV/Propidiumiodide-staining and gel electrophoresis. Our results demonstrate that activated bentonite particles are better taken up by IMR90-cells than untreated (native) bentonite particles. Also, activated bentonite particles with a quartz content of 5-6% were more cytotoxic than untreated bentonites or bentonites with a quartz content lower than 4%. The bentonite samples induced necrotic as well as apoptotic cell death. In general, bentonites showed a high membrane-damaging potential shown as hemolytic activity in human erythrocytes. We conclude that cellular effects of bentonite particles in human lung cells are enhanced after chemical treatment of the particles. The cytotoxic potential of the different bentonites is primarily characterized by a strong lysis of the cell membrane. (orig.)

  7. Podoplanin Expression in Cancer-associated Fibroblasts Predicts Poor Prognosis in Patients with Squamous Cell Carcinoma of the Lung.

    Science.gov (United States)

    Yurugi, Yohei; Wakahara, Makoto; Matsuoka, Yuki; Sakabe, Tomohiko; Kubouchi, Yasuaki; Haruki, Tomohiro; Nosaka, Kanae; Miwa, Ken; Araki, Kunio; Taniguchi, Yuji; Shiomi, Tatsushi; Nakamura, Hiroshige; Umekita, Yoshihisa

    2017-01-01

    Podoplanin is a candidate cancer stem cell marker in squamous cell carcinoma (SCC). Several studies have reported the prognostic value of podoplanin expression in tumor cells in lung SCC but few have focused on its expression in cancer-associated fibroblasts (CAFs). The aim of this study was to analyze the prognostic significance of podoplanin expression, with special reference to the expression pattern in both tumor cells and CAFs. Immunohistochemical analyses using anti-podoplanin antibody were performed on 126 resected specimens of lung SCC. When more than 10% of tumor cells or CAFs showed immunoreactivity with podoplanin levels as strong as those of the positive controls, the specimens were classified as a podoplanin-positive. Podoplanin-positive status in tumor cells (n=54) was correlated with a lower incidence of lymphatic invasion (p=0.031) but there were no significant differences in disease-free survival (DFS) and disease-specific survival (DSS) by the log-rank test. Podoplanin-positive status in CAFs (n=41) was correlated with more advanced stage (p=0.008), higher frequency of pleural invasion (p=0.002) and both shorter DFS (p=0.006) and DSS (p=0.006). In Cox's multivariate analysis, podoplanin-positive status in CAFs was an independent negative prognostic factor for DFS (p=0.027) and DSS (p=0.027). Podoplanin expression in CAFs might be an independent unfavorable prognostic indicator in patients with lung SCC, irrespective of the expression status of tumor cells. Copyright© 2017 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  8. Budesonide and formoterol inhibit ICAM-1 and VCAM-1 expression of human lung fibroblasts

    NARCIS (Netherlands)

    Spoelstra, FM; Postma, DS; Hovenga, H; Noordhoek, JA; Kauffman, HF

    The glucocorticoid budesonide and the long-acting beta(2)-adrenoceptor agonist formoterol are used in asthma therapy for their anti-inflammatory and bronchodilating effects, respectively. Since expression of adhesion molecules on resident cells in the lung plays an important role in asthmatic

  9. Mast cell density in isolated monkey lungs on exposure to cigarette smoke.

    OpenAIRE

    Walter, A; Walter, S

    1982-01-01

    The density and percentage of degranulated cells of the mast cell population were studied in the isolated lungs of 25 monkeys (Macaca radiata radiata) before and after acute exposure to cigarette smoke. In each animal one lung was used as the test lung while the other lung was used as its control. In the control lungs the total mean mast cell count was 9.5/mm2 and the proportion of degranulated cells was 9.7%. In the lungs exposed to smoke the total counts were lower (7.3/mm2) and the percent...

  10. Transforming growth factor-β1 induces expression of human coagulation factor XII via Smad3 and JNK signaling pathways in human lung fibroblasts.

    Science.gov (United States)

    Jablonska, Ewa; Markart, Philipp; Zakrzewicz, Dariusz; Preissner, Klaus T; Wygrecka, Malgorzata

    2010-04-09

    Coagulation factor XII (FXII) is a liver-derived serine protease involved in fibrinolysis, coagulation, and inflammation. The regulation of FXII expression is largely unknown. Transforming growth factor-beta1 (TGF-beta1) is a multifunctional cytokine that has been linked to several pathological processes, including tissue fibrosis by modulating procoagulant and fibrinolytic activities. This study investigated whether TGF-beta1 may regulate FXII expression in human lung fibroblasts. Treatment of human lung fibroblasts with TGF-beta1 resulted in a time-dependent increase in FXII production, activation of p44/42, p38, JNK, and Akt, and phosphorylation and translocation into the nucleus of Smad3. However, TGF-beta1-induced FXII expression was repressed only by the JNK inhibitor and JNK and Smad3 antisense oligonucleotides but not by MEK, p38, or phosphoinositide 3-kinase blockers. JNK inhibition had no effect on TGF-beta1-induced Smad3 phosphorylation, association with Smad4, and its translocation into the nucleus but strongly suppressed Smad3-DNA complex formation. FXII promoter analysis revealed that the -299/+1 region was sufficient for TGF-beta1 to induce FXII expression. Sequence analysis of this region detected a potential Smad-binding element at position -272/-269 (SBE-(-272/-269)). Chromatin immunoprecipitation and streptavidin pulldown assays demonstrated TGF-beta1-dependent Smad3 binding to SBE-(-272/-269). Mutation or deletion of SBE-(-272/-269) substantially reduced TGF-beta1-mediated activation of the FXII promoter. Clinical relevance was demonstrated by elevated FXII levels and its co-localization with fibroblasts in the lungs of patients with acute respiratory distress syndrome. Our results show that JNK/Smad3 pathway plays a critical role in TGF-beta1-induced FXII expression in human lung fibroblasts and implicate its possible involvement in pathological conditions characterized by elevated TGF-beta1 levels.

  11. Cancer-Associated Fibroblasts from lung tumors maintain their immuno-suppressive abilities after high-dose irradiation

    Directory of Open Access Journals (Sweden)

    Laia eGorchs

    2015-05-01

    Full Text Available Accumulating evidence supports the notion that high-dose (>5 Gy radiotherapy (RT regimens are triggering stronger pro-immunogenic effects than standard low-dose (2 Gy regimens. However, the effects of RT on certain immunoregulatory elements in tumors remain unexplored. In this study we have investigated the effects of high-dose irradiation (HD-RT on the immunomodulating functions of cancer-associated fibroblasts (CAFs. Primary CAF cultures were established from lung cancer specimens derived from patients diagnosed for non-small cell lung cancer. Irradiated and non-irradiated CAFs were examined for immunomodulation in experiments with peripheral blood mononuclear cells from random, healthy donors. Regulation of lymphocytes behavior was checked by lymphocyte proliferation assays, lymphocyte migration assays and T-cell cytokine production. Additionally, CAF-secreted immuno-regulatory factors were studied by multiplex protein arrays, ELISAs and by LC-MS/MS proteomics. In all functional assays we observed a powerful immuno-suppressive effect exerted by CAF-conditioned medium on activated T-cells (p>0,001, and this effect was sustained after a single radiation dose of 18 Gy. Relevant immuno-suppressive molecules such as prostaglandin E2, interleukin-6 and -10, or transforming growth factor-β were found in CAF conditioned medium, but their secretion was unchanged after irradiation. Finally, immunogenic cell death responses in CAFs were studied by exploring the release of high motility group box-1 and ATP. Both alarmins remained undetectable before and after irradiation. In conclusion, CAFs play a powerful immuno-suppressive effect over activated T-cells, and this effect remains unchanged after HD-RT. Importantly, CAFs do not switch on immunogenic cell death responses after exposure to HD-RT.

  12. Repression of TSC1/TSC2 mediated by MeCP2 regulates human embryo lung fibroblast cell differentiation and proliferation.

    Science.gov (United States)

    Wang, Yuanyuan; Chen, Chen; Deng, Ziyu; Bian, Erbao; Huang, Cheng; Lei, Ting; Lv, Xiongwen; Liu, Liping; Li, Jun

    2017-03-01

    Pulmonary fibrosis (PF) is a severe inflammatory disease with limited effective treatments. It is known that the transdifferentiation of human embryo lung fibroblast (HELF) cells from pulmonary fibroblasts into myofibroblasts, contributes to the progression of pulmonary fibrogenesis. The tuberous sclerosis proteins TSC1 and TSC2 are two key signaling factors which can suppress cell growth and proliferation. However, the roles of TSC1 and TSC2 in lung fibroblast are unclear. Here, we developed a PF model with bleomycin (BLM) in mice and conducted several simulation experiments in HELF cells. Our study shows that the expression of TSC1 and TSC2 in fibrotic mice lung was reduced and stimulation of HELF cells with TGF-β1 resulted in a down-regulation of TSC1 and TSC2. In addition, overexpression of TSC1 or TSC2 decreased cell proliferation and differentiation. Furthermore, we found that reduced expression of TSC1 and TSC2 caused by TGF-β1 is associated with the promoter methylation status of TSC1 and TSC2. MeCP2, controls an epigenetic pathway that promotes myofibroblast transdifferentiation and fibrosis. We found that expression of TSC1 and TSC2 can be repressed by MeCP2, which regulates HELF cell differentiation and proliferation as myofibroblasts and lead to PF ultimately. Copyright © 2016. Published by Elsevier B.V.

  13. Two methods for isolating the lung area of a CT scan for density information

    International Nuclear Information System (INIS)

    Hedlund, L.W.; Anderson, R.F.; Goulding, P.L.; Beck, J.W.; Effmann, E.L.; Putman, C.E.

    1982-01-01

    Extracting density information from irregularly shaped tissue areas of CT scans requires automated methods when many scans are involved. We describe two computer methods that automatically isolate the lung area of a CT scan. Each starts from a single, operator specified point in the lung. The first method follows the steep density gradient boundary between lung and adjacent tissues; this tracking method is useful for estimating the overall density and total area of lung in a scan because all pixels within the lung area are available for statistical sampling. The second method finds all contiguous pixels of lung that are within the CT number range of air to water and are not a part of strong density gradient edges; this method is useful for estimating density and area of the lung parenchyma. Structures within the lung area that are surrounded by strong density gradient edges, such as large blood vessels, airways and nodules, are excluded from the lung sample while lung areas with diffuse borders, such as an area of mild or moderate edema, are retained. Both methods were tested on scans from an animal model of pulmonary edema and were found to be effective in isolating normal and diseased lungs. These methods are also suitable for isolating other organ areas of CT scans that are bounded by density gradient edges

  14. DNA-damage effect of polycyclic aromatic hydrocarbons from urban area, evaluated in lung fibroblast cultures

    International Nuclear Information System (INIS)

    Calesso Teixeira, Elba; Pra, Daniel; Idalgo, Daniele; Henriques, João Antonio Pêgas; Wiegand, Flavio

    2012-01-01

    This study was designed to biomonitor the effect of PAH extracts from urban areas on the DNA of lung cell cultures. The analyses of the polycyclic aromatic hydrocarbons (PAHs) were performed in atmospheric PM 2.5 and PM 10 collected at three sampling sites with heavy traffic located in the Metropolitan Area of Porto Alegre (MAPA) (Brazil). The concentrations of 16 major PAHs were determined according to EPA. Comet assay on V79 hamster lung cells was chosen for genotoxicity evaluation. Temperature, humidity, and wind speed were recorded. With regard to the damage index, higher levels were reported in the extract of particulate matter samples from the MAPA during the summer. High molecular weight compounds showed correlation with DNA damage frequency and their respective carcinogenicity. - Highlights: ► Cell line V79 was used to assess the effect of PAHs in PM 2.5 and PM 10 from urban area. ► Temperature showed a significant seasonal variation with the level of DNA damage. ► PAHs with higher molecular weight contributed to higher DNA damage levels. - DNA-damage effect of polycyclic aromatic hydrocarbons from urban area, showed difference according to season

  15. Antagonism of phenanthrene cytotoxicity for human embryo lung fibroblast cell line HFL-I by green tea polyphenols

    Energy Technology Data Exchange (ETDEWEB)

    Mei Xin [Department of Tea Science, Zhejiang University, Hangzhou 310029 (China); Key Laboratory of Horticultural Plant Growth Development and Biotechnology of Ministry of Agriculture, Zhejiang University, Hangzhou 310029 (China); Wu Yuanyuan; Mao Xiao [Department of Tea Science, Zhejiang University, Hangzhou 310029 (China); Tu Youying, E-mail: youytu@zju.edu.c [Department of Tea Science, Zhejiang University, Hangzhou 310029 (China)

    2011-01-15

    Polycyclic aromatic hydrocarbons (PAHs) have been detected in some commercial teas around the world and pose a threat to tea consumers. However, green tea polyphenols (GTP) possess remarkable antioxidant and anticancer effects. In this study, the potential of GTP to block the toxicity of the model PAH phenanthrene was examined in human embryo lung fibroblast cell line HFL-I. Both GTP and phenanthrene treatment individually caused dose-dependent inhibition of cell growth. A full factorial design experiment demonstrated that the interaction of phenanthrene and GTP significantly reduced growth inhibition. Using the median effect method showed that phenanthrene and GTP were antagonistic when the inhibitory levels were less than about 50%. Apoptosis and cell cycle detection suggested that only phenanthrene affected cell cycle significantly and caused cell death; GTP lowered the mortality of HFL-I cells exposed to phenanthrene; However, GTP did not affect modulation of the cell cycle by phenanthrene. - Green tea polyphenols antagonised cytotoxicity of a low-ring PAH phenanthrene.

  16. Characterization of Na+-linked and Na+-independent Cl-/HCO3- exchange systems in Chinese hamster lung fibroblasts

    International Nuclear Information System (INIS)

    Cassel, D.; Scharf, O.; Rotman, M.; Cragoe, E.J. Jr.; Katz, M.

    1988-01-01

    The PS120 variant of Chinese hamster lung fibroblasts which lacks Na + /H + exchange activity was used to investigate bicarbonate transport systems and their role in intracellular pH (pH/sub i/) regulation. When pH/sub i/ was decreased by acid load, bicarbonate caused pH/sub i/ increase and stimulated 36 Cl - efflux from the cells, both in a Na + -dependent manner. These results together with previous findings that bicarbonate stimulates 22 Na + uptake in PS120 cells demonstrate the presence of a Na + -linked Cl - /HCO 3 - exchange system. In cells with normal initial pH/sub i/, bicarbonate caused Na + -independent pH/sub i/ increase in Cl - -free solutions and stimulated Na + -independent 36 Cl - efflux, indicating that a Na + -independent Cl - /HCO 3 - exchanger is also present in the cell. Na + -linked and Na + -independent Cl - /HCO 3- exchange is apparently mediated by two distinct systems, since a [(tetrahydrofluorene-7-yl)oxy]acetic acid derivative selectively inhibits the Na + -independent exchanger. An additional distinctive features is a 10-fold lower affinity for chloride of the Na + -linked exchanger. The Na + -linked and Na + -independent Cl - /HCO 3 - exchange systems are likely to protect the cell from acid and alkaline load, respectively

  17. Antagonism of phenanthrene cytotoxicity for human embryo lung fibroblast cell line HFL-I by green tea polyphenols

    International Nuclear Information System (INIS)

    Mei Xin; Wu Yuanyuan; Mao Xiao; Tu Youying

    2011-01-01

    Polycyclic aromatic hydrocarbons (PAHs) have been detected in some commercial teas around the world and pose a threat to tea consumers. However, green tea polyphenols (GTP) possess remarkable antioxidant and anticancer effects. In this study, the potential of GTP to block the toxicity of the model PAH phenanthrene was examined in human embryo lung fibroblast cell line HFL-I. Both GTP and phenanthrene treatment individually caused dose-dependent inhibition of cell growth. A full factorial design experiment demonstrated that the interaction of phenanthrene and GTP significantly reduced growth inhibition. Using the median effect method showed that phenanthrene and GTP were antagonistic when the inhibitory levels were less than about 50%. Apoptosis and cell cycle detection suggested that only phenanthrene affected cell cycle significantly and caused cell death; GTP lowered the mortality of HFL-I cells exposed to phenanthrene; However, GTP did not affect modulation of the cell cycle by phenanthrene. - Green tea polyphenols antagonised cytotoxicity of a low-ring PAH phenanthrene.

  18. Multi-walled carbon nanotubes (NM401) induce ROS-mediated HPRT mutations in Chinese hamster lung fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Rubio, Laura [Grup de Mutagènesi, Departament de Genètica i de Microbiologia, Facultat de Biociències, Universitat Autònoma de Barcelona, Bellaterra (Spain); El Yamani, Naouale [Health Effects Laboratory-MILK, NILU-Norwegian Institute for Air Research, Kjeller (Norway); Kazimirova, Alena [Department of Biology, Slovak Medical University, Bratislava (Slovakia); Dusinska, Maria, E-mail: maria.dusinska@nilu.no [Health Effects Laboratory-MILK, NILU-Norwegian Institute for Air Research, Kjeller (Norway); Marcos, Ricard, E-mail: ricard.marcos@uab.es [Grup de Mutagènesi, Departament de Genètica i de Microbiologia, Facultat de Biociències, Universitat Autònoma de Barcelona, Bellaterra (Spain); CIBER Epidemiología y Salud Pública, Instituto de Salud Carlos III, Madrid (Spain)

    2016-04-15

    Although there is an important set of data showing potential genotoxic effects of nanomaterials (NMs) at the DNA (comet assay) and chromosome (micronucleus test) levels, few studies have been conducted to analyze their potential mutagenic effects at gene level. We have determined the ability of multi-walled carbon nanotubes (MWCNT, NM401), to induce mutations in the HPRT gene in Chinese hamster lung (V79) fibroblasts. NM401, characterized in the EU NanoGenotox project, were further studied within the EU Framework Programme Seven (FP7) project NANoREG. From the proliferation assay data we selected a dose-range of 0.12 to 12 µg/cm{sup 2} At these range we have been able to observe significant cellular uptake of MWCNT by using transmission electron microscopy (TEM), as well as a concentration-dependent induction of intracellular reactive oxygen species. In addition, a clear concentration-dependent increase in the induction of HPRT mutations was also observed. Data support a potential genotoxic/ carcinogenic risk associated with MWCNT exposure. - Highlights: • MWCNT were tested in V79 cells. • Cellular uptake of MWCNT was detected using TEM. • Intracellular ROS induction was observed after MWCNT exposure. • MWCNT induced a concentration-dependent increase of HPRT mutations.

  19. Acrolein-exposed normal human lung fibroblasts in vitro: cellular senescence, enhanced telomere erosion, and degradation of Werner's syndrome protein.

    Science.gov (United States)

    Jang, Jun-Ho; Bruse, Shannon; Huneidi, Salam; Schrader, Ronald M; Monick, Martha M; Lin, Yong; Carter, A Brent; Klingelhutz, Aloysius J; Nyunoya, Toru

    2014-09-01

    Acrolein is a ubiquitous environmental hazard to human health. Acrolein has been reported to activate the DNA damage response and induce apoptosis. However, little is known about the effects of acrolein on cellular senescence. We examined whether acrolein induces cellular senescence in cultured normal human lung fibroblasts (NHLF). We cultured NHLF in the presence or absence of acrolein and determined the effects of acrolein on cell proliferative capacity, senescence-associated β-galactosidase activity, the known senescence-inducing pathways (e.g., p53, p21), and telomere length. We found that acrolein induced cellular senescence by increasing both p53 and p21. The knockdown of p53 mediated by small interfering RNA (siRNA) attenuated acrolein-induced cellular senescence. Acrolein decreased Werner's syndrome protein (WRN), a member of the RecQ helicase family involved in DNA repair and telomere maintenance. Acrolein-induced down-regulation of WRN protein was rescued by p53 knockdown or proteasome inhibition. Finally, we found that acrolein accelerated p53-mediated telomere shortening. These results suggest that acrolein induces p53-mediated cellular senescence accompanied by enhanced telomere attrition and WRN protein down-regulation.

  20. Multi-walled carbon nanotubes (NM401) induce ROS-mediated HPRT mutations in Chinese hamster lung fibroblasts

    International Nuclear Information System (INIS)

    Rubio, Laura; El Yamani, Naouale; Kazimirova, Alena; Dusinska, Maria; Marcos, Ricard

    2016-01-01

    Although there is an important set of data showing potential genotoxic effects of nanomaterials (NMs) at the DNA (comet assay) and chromosome (micronucleus test) levels, few studies have been conducted to analyze their potential mutagenic effects at gene level. We have determined the ability of multi-walled carbon nanotubes (MWCNT, NM401), to induce mutations in the HPRT gene in Chinese hamster lung (V79) fibroblasts. NM401, characterized in the EU NanoGenotox project, were further studied within the EU Framework Programme Seven (FP7) project NANoREG. From the proliferation assay data we selected a dose-range of 0.12 to 12 µg/cm 2 At these range we have been able to observe significant cellular uptake of MWCNT by using transmission electron microscopy (TEM), as well as a concentration-dependent induction of intracellular reactive oxygen species. In addition, a clear concentration-dependent increase in the induction of HPRT mutations was also observed. Data support a potential genotoxic/ carcinogenic risk associated with MWCNT exposure. - Highlights: • MWCNT were tested in V79 cells. • Cellular uptake of MWCNT was detected using TEM. • Intracellular ROS induction was observed after MWCNT exposure. • MWCNT induced a concentration-dependent increase of HPRT mutations.

  1. Arterio-venous anastomoses in isolated, perfused rat lungs.

    Science.gov (United States)

    Conhaim, Robert L; Segal, Gilad S; Watson, Kal E

    2016-11-01

    Several studies have suggested that large-diameter (>25 μm) arterio-venous shunt pathways exist in the lungs of rats, dogs, and humans. We investigated the nature of these pathways by infusing specific-diameter fluorescent latex particles (4, 7, 15, 30, or 50 μm) into isolated, ventilated rat lungs perfused at constant pressure. All lungs received the same mass of latex (5 mg), which resulted in infused particle numbers that ranged from 1.7 × 10 7 4 μm particles to 7.5 × 10 4 50 μm particles. Particles were infused over 2 min. We used a flow cytometer to count particle appearances in venous effluent samples collected every 0.5 min for 12 min from the start of particle infusion. Cumulative percentages of infused particles that appeared in the samples averaged 3.17 ± 2.46% for 4 μm diameter particles, but ranged from 0.01% to 0.17% for larger particles. Appearances of 4 μm particles followed a rapid upslope beginning at 30 sec followed by a more gradual downslope that lasted for up to 12 min. All other particle diameters also began to appear at 30 sec, but followed highly irregular time courses. Infusion of 7 and 15 μm particles caused transient but significant perfusate flow reductions, while infusion of all other diameters caused insignificant reductions in flow. We conclude that small numbers of bypass vessels exist that can accommodate particle diameters of 7-to-50 μm. We further conclude that our 4 μm particle data are consistent with a well-developed network of serial and parallel perfusion pathways at the acinar level. Published 2016. This article is a U.S. Government work and is in the public domain in the USA. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

  2. Age-dependent oxidation of extracellular cysteine/cystine redox state (Eh(Cys/CySS)) in mouse lung fibroblasts is mediated by a decline in Slc7a11 expression.

    Science.gov (United States)

    Zheng, Yuxuan; Ritzenthaler, Jeffrey D; Burke, Tom J; Otero, Javier; Roman, Jesse; Watson, Walter H

    2018-04-01

    Aging is associated with progressive oxidation of the extracellular environment. The redox state of human plasma, defined by the concentrations of cysteine (Cys) and cystine (CySS), becomes more oxidized as we age. Recently, we showed that fibroblasts isolated from the lungs of young and old mice retain this differential phenotype; old cells produce and maintain a more oxidizing extracellular redox potential (E h (Cys/CySS)) than young cells. Microarray analysis identified down-regulation of Slc7a11, the light subunit of the CySS/glutamate transporter, as a potential mediator of age-related oxidation in these cells. The purpose of the present study was to investigate the mechanistic link between Slc7a11 expression and extracellular E h (Cys/CySS). Sulforaphane treatment or overexpression of Slc7a11 was used to increase Slc7a11 in lung fibroblasts from old mice, and sulfasalazine treatment or siRNA-mediated knock down was used to decrease Slc7a11 in young fibroblasts. Slc7a11 mRNA levels were measured by real-time PCR, Slc7a11 activity was determined by measuring the rate of glutamate release, Cys, CySS, glutathione (GSH) and its disulfide (GSSG) were measured by HPLC, and E h (Cys/CySS) was calculated from the Nernst equation. The results showed that both E h (Cys/CySS) and E h (GSH/GSSG) were more oxidized in the conditioned media of old cells than in young cells. Up-regulation of Slc7a11 via overexpression or sulforaphane treatment restored extracellular E h (Cys/CySS) in cultures of old cells, whereas down-regulation reproduced the oxidizing E h (Cys/CySS) in young cells. Only sulforaphane treatment was able to increase total GSH and restore E h (GSH/GSSG), whereas overexpression, knock down and sulfasalazine had no effect on these parameters. In addition, inhibition of GSH synthesis with buthionine sulfoximine had no effect on the ability of cells to restore their extracellular redox potential in response to an oxidative challenge. In conclusion, our study

  3. Presence of nanosilica (E551) in commercial food products: TNF-mediated oxidative stress and altered cell cycle progression in human lung fibroblast cells.

    Science.gov (United States)

    Athinarayanan, Jegan; Periasamy, Vaiyapuri Subbarayan; Alsaif, Mohammed A; Al-Warthan, Abdulrahman A; Alshatwi, Ali A

    2014-04-01

    Silica (E551) is commonly used as an anti-caking agent in food products. The morphology and the dimension of the added silica particles are not, however, usually stated on the food product label. The food industry has adapted nanotechnology using engineered nanoparticles to improve the quality of their products. However, there has been increased debate regarding the health and safety concerns related to the use of engineered nanoparticles in consumer products. In this study, we investigated the morphology and dimensions of silica (E551) particles in food. The silica content of commercial food products was determined using inductively coupled plasma optical emission spectrometry. The result indicates that 2.74-14. 45 μg/g silica was found in commercial food products; however, the daily dietary intake in increase causes adverse effects on human health. E551 was isolated from food products and the morphology, particle size, crystalline nature, and purity of the silica particles were analyzed using XRD, FTIR, TEM, EDX and DLS. The results of these analyses confirmed the presence of spherical silica nanoparticles (of amorphous nature) in food, approximately 10-50 nm in size. The effects of E551 on human lung fibroblast cell viability, intracellular ROS levels, cell cycle phase, and the expression levels of metabolic stress-responsive genes (CAT, GSTA4, TNF, CYP1A, POR, SOD1, GSTM3, GPX1, and GSR1) were studied. The results suggest that E551 induces a dose-dependent cytotoxicity and changes in ROS levels and alters the gene expression and cell cycle. Treatment with a high concentration of E551 caused significant cytotoxic effects on WI-38 cells. These findings have implications for the use of these nanoparticles in the food industry.

  4. Pock forming ability of fowl pox virus isolated from layer chicken and its adaptation in chicken embryo fibroblast cell culture.

    Science.gov (United States)

    Gilhare, Varsha Rani; Hirpurkar, S D; Kumar, Ashish; Naik, Surendra Kumar; Sahu, Tarini

    2015-03-01

    The objective of the present study was to examine pock forming ability of field strain and vaccine strain of fowl pox virus (FPV) in chorioallantoic membrane (CAM) of embryonated chicken eggs and its adaptation in chicken embryo fibroblast (CEF) cell culture. Dry scabs were collected from 25 affected birds in glycerin-saline and preserved at 4°C until processed. Virus was isolated in 10-day-old embryonated chicken eggs by dropped CAM method. The identity of the virus is confirmed by clinical findings of affected birds, pock morphology and histopathology of infected CAM. In addition one field isolate and vaccine strain of FPV was adapted to CEF cell culture. CEF cell culture was prepared from 9-day-old embryonated chicken eggs. Clinical symptoms observed in affected birds include pox lesion on comb, wattle, eyelids and legs, no internal lesions were observed. All field isolates produced similar findings in CAM. Pocks produced by field isolates ranged from 3 mm to 5 mm at the third passage while initial passages edematous thickening and necrosis of CAM was observed. Pocks formed by lyophilized strain were ranges from 0.5 mm to 2.5 mm in diameter scattered all over the membrane at the first passage. Intra-cytoplasmic inclusion bodies are found on histopathology of CAM. At third passage level, the CEF inoculated with FPV showed characteristic cytopathic effect (CPE) included aggregation of cells, syncytia and plaque formation. FPV field isolates and vaccine strain produced distinct pock lesions on CAMs. Infected CAM showed intracytoplasmic inclusion bodies. The CEF inoculated with FPV field isolate as well as a vaccine strain showed characteristic CPE at third passage level.

  5. Hormetic effects of noncoplanar PCB exposed to human lung fibroblast cells (HELF) and possible role of oxidative stress.

    Science.gov (United States)

    Hashmi, Muhammad Zaffar; Khan, Kiran Yasmin; Hu, Jinxing; Naveedullah; Su, Xiaomei; Abbas, Ghulam; Yu, Chunna; Shen, Chaofeng

    2015-12-01

    Hormesis, a biphasic dose-response phenomenon, which is characterized by stimulation of an end point at a low-dose and inhibition at a high-dose. In the present study we used human lungs fibroblast (HELF) cells as a test model to evaluate the role of oxidative stress (OS) in hormetic effects of non coplanar PCB 101. Results from 3-(4,5-dime-thylthiazol-2-yl)-2,5-diphenyltetrazo-lium bromide (MTT) assay indicated that PCB101 at lower concentrations (10(-5) to 10(-1) μg mL(-1) ) stimulated HELF cell proliferation and inhibited at high concentrations (1, 5, 10, and 20 μg mL(-1) ) in a dose- and time-dependent manner. Reactive oxygen species (ROS), malondialdehyde (MDA) and superoxide dismutase (SOD) (except 48 h) showed a significant increase at higher concentrations of PCB 101 than those at the lower concentrations with the passage of time. Antioxidant enzymes such as glutathione peroxidase (GSH-Px) exhibited decreasing trends in dose and time dependent manner. Lipid peroxidation assay resulted in a significant increase (P PCB 101-treated HELF cells compared with controls, suggesting that OS plays a key role in PCB 101-induced toxicity. Comet assay indicated a significant increase in genotoxicity at higher concentrations of PCB 101 exposure compared to lower concentrations. Overall, we found that HELF cell proliferation was higher at low ROS level and vice versa, which revealed activation of cell signaling-mediated hormetic mechanisms. The results suggested that PCB 101 has hormetic effects to HELF cells and these were associated with oxidative stress. © 2014 Wiley Periodicals, Inc.

  6. Dicholesteroyl diselenide: cytotoxicity, genotoxicity and mutagenicity in the yeast Saccharomyces cerevisiae and in Chinese hamster lung fibroblasts.

    Science.gov (United States)

    de Oliveira, Iuri Marques; Degrandi, Tiago Hoerbe; Jorge, Patrícia Mendes; Saffi, Jenifer; Rosa, Renato Moreira; Guecheva, Temenouga Nikolova; Henriques, João Antonio Pêgas

    2014-03-15

    The organoselenium compound, dicholesteroyl diselenide (DCDS) is a structural analogue of diphenyl diselenide (DPDS) and may be considered as a promising antioxidant drug in vivo. Nevertheless, little is known about the toxicological properties of DCDS. In the present study we evaluated the cytotoxic, genotoxic and mutagenic properties of DCDS in Chinese hamster lung fibroblasts (V79) and in strains of the yeast Saccharomyces cerevisiae, proficient and deficient in several DNA-repair pathways. The results with V79 cells show that DCDS induced cytotoxicity, GSH depletion and elevation of lipid peroxidation at lower concentrations than did DPDS. DCDS also generated single- and double-strand DNA breaks in V79 cells, both in the presence and in the absence of metabolic activation, as revealed by alkaline and neutral comet assays. Moreover, the induction of oxidative DNA base-damage was demonstrated by means of a modified comet assay with formamidopyrimidine-DNA glycosylase and endonuclease III. Treatment with DCDS also induced micronucleus formation in V79 cells as well as point and frame-shift mutations in a haploid wild-type strain of S. cerevisiae. Yeast mutants defective in base excision-repair proteins were the most sensitive to DCDS. Pre-incubation with N-acetylcysteine reduced DCDS's oxidative, genotoxic and mutagenic effects in yeast and in V79 cells. Our findings indicate that the presence of cholesteroyl substituents in DCDS results in elevation of its cytotoxic and genotoxic potential compared with that of DPDS in yeast and in V79 cells. However, due to dose-dependent contrasting behaviour of organoselenium compounds and differences in their toxicity in in vitro and in vivo systems, further studies are needed in order to establish the non-toxic concentration range for treatment in mammals. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. Design and validation of a clinical-scale bioreactor for long-term isolated lung culture.

    Science.gov (United States)

    Charest, Jonathan M; Okamoto, Tatsuya; Kitano, Kentaro; Yasuda, Atsushi; Gilpin, Sarah E; Mathisen, Douglas J; Ott, Harald C

    2015-06-01

    The primary treatment for end-stage lung disease is lung transplantation. However, donor organ shortage remains a major barrier for many patients. In recent years, techniques for maintaining lungs ex vivo for evaluation and short-term (advance to more complex interventions for lung repair and regeneration, the need for a long-term organ culture system becomes apparent. Herein we describe a novel clinical scale bioreactor capable of maintaining functional porcine and human lungs for at least 72 h in isolated lung culture (ILC). The fully automated, computer controlled, sterile, closed circuit system enables physiologic pulsatile perfusion and negative pressure ventilation, while gas exchange function, and metabolism can be evaluated. Creation of this stable, biomimetic long-term culture environment will enable advanced interventions in both donor lungs and engineered grafts of human scale. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Edaravone Decreases Paraquat Toxicity in A549 Cells and Lung Isolated Mitochondria

    OpenAIRE

    Shokrzadeh, Mohammad; Shaki, Fatemeh; Mohammadi, Ebrahim; Rezagholizadeh, Neda; Ebrahimi, Fatemeh

    2014-01-01

    Edaravone, an antioxidant and radical scavenger, showed protective effects against oxidative stress-like condition. Paraquat (PQ) is toxic herbicide considerable evidence suggests that oxidative stress and mitochondrial dysfunction contribute to PQ toxicity. In this study, protective effect of edaravone against PQ induced toxicity and reactive oxygen species (ROS) generation in A549 cells and lung isolated mitochondria were evaluated. A549 cells and lung isolated mitochondria were divided int...

  9. Kinetics of reversible-sequestration of leukocytes by the isolated perfused rat lung

    Energy Technology Data Exchange (ETDEWEB)

    Goliaei, B.

    1980-08-01

    The kinetics and morphology of sequestration and margination of rat leukocytes were studied using an isolated perfused and ventilated rat lung preparation. Whole rat blood, bone marrow suspension, or leukocyte suspensions, were used to perfuse the isolated rat lung. The lung was also perfused with latex particle suspensions and the passage of particles through the lung capillaries was studied. When a leukocyte suspension was perfused through the lung in the single-pass mode, the rate of sequestration decreased as more cells were perfused. In contrast, latex particles of a size comparable to that of leukocytes were totally stopped by the lung. When the leukocyte suspension was recirculated through the lung, cells were rapidly removed from circulation until a steady state was reached, after which no net removal of cells by the lung occurred. These results indicate that leukocytes are reversibly sequestered from circulation. The sequestered cells marginated and attached to the luminal surface of the endothelium of post-capillary venules and veins. A mathematical model was developed based on the assumption that the attachment and detachment of leukocytes to blood vessel walls follows first-order kinetics. The model correctly predicts the following characteristics of the system: (a) the kinetics of the sequestration of leukocytes by the lung; (b) the existence of a steady state when a suspension of leukocytes is recirculated through the lung; and (c) the independence of the fraction of cells remaining in circulation from the starting concentration for all values of starting concentration. (ERB)

  10. AhR-dependent secretion of PDGF-BB by human classically activated macrophages exposed to DEP extracts stimulates lung fibroblast proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Jaguin, Marie [UMR INSERM U1085, Institut de Recherche sur la Santé, l' Environnement et le Travail (IRSET), Université de Rennes 1, 2 Avenue du Pr Léon Bernard, 35043 Rennes Cedex (France); Fardel, Olivier [UMR INSERM U1085, Institut de Recherche sur la Santé, l' Environnement et le Travail (IRSET), Université de Rennes 1, 2 Avenue du Pr Léon Bernard, 35043 Rennes Cedex (France); Pôle Biologie, Centre Hospitalier Universitaire (CHU) Rennes, 2 rue Henri Le Guilloux, 35033 Rennes Cedex (France); Lecureur, Valérie, E-mail: valerie.lecureur@univ-rennes1.fr [UMR INSERM U1085, Institut de Recherche sur la Santé, l' Environnement et le Travail (IRSET), Université de Rennes 1, 2 Avenue du Pr Léon Bernard, 35043 Rennes Cedex (France)

    2015-06-15

    Lung diseases are aggravated by exposure to diesel exhaust particles (DEPs) found in air pollution. Macrophages are thought to play a crucial role in lung immune response to these pollutants, even if the mechanisms involved remain incompletely characterized. In the present study, we demonstrated that classically and alternative human macrophages (MΦ) exhibited increased secretion of PDGF-B in response to DEP extract (DEPe). This occurred via aryl hydrocarbon receptor (AhR)-activation because DEPe-induced PDGF-B overexpression was abrogated after AhR expression knock-down by RNA interference, in both M1 and M2 polarizing MΦ. In addition, TCDD and benzo(a)pyrene, two potent AhR ligands, also significantly increased mRNA expression of PDGF-B in M1 MΦ, whereas some weak ligands of AhR did not. We next evaluated the impact of conditioned media (CM) from MΦ culture exposed to DEPe or of recombinant PDGF-B onto lung fibroblast proliferation. The tyrosine kinase inhibitor, AG-1295, prevents phosphorylations of PDGF-Rβ, AKT and ERK1/2 and the proliferation of MRC-5 fibroblasts induced by recombinant PDGF-B and by CM from M1 polarizing MΦ, strongly suggesting that the PDGF-BB secreted by DEPe-exposed MΦ is sufficient to activate the PDGF-Rβ pathway of human lung fibroblasts. In conclusion, we demonstrated that human MΦ, whatever their polarization status, secrete PDGF-B in response to DEPe and that PDGF-B is a target gene of AhR. Therefore, induction of PDGF-B by DEP may participate in the deleterious effects towards human health triggered by such environmental urban contaminants. - Highlights: • PDGF-B expression and secretion are increased by DEPe exposure in human M1 and M2 MΦ. • DEPe-induced PDGF-B expression is aryl-hydrocarbon-dependent. • DEPe-exposed M1 MΦ secrete sufficient PDGF-B to increase lung fibroblast proliferation.

  11. Interleukin-1 or tumor necrosis factor-alpha augmented the cytotoxic effect of mycobacteria on human fibroblasts: application to evaluation of pathogenesis of clinical isolates of Mycobacterium tuberculosis and M. avium complex.

    Science.gov (United States)

    Takii, T; Abe, C; Tamura, A; Ramayah, S; Belisle, J T; Brennan, P J; Onozaki, K

    2001-03-01

    Mycobacteria-induced in vitro events reflecting human tuberculosis can contribute to the evaluation of the pathogenesis of Mycobacterium tuberculosis (MTB). In this study, we propose such an in vitro method based on live mycobacteria-induced cytotoxicity to human cell lines. When human lung-derived normal fibroblast cell line MRC-5 was infected with various strains of mycobacteria (M. tuberculosis H(37)Rv and H(37) Ra, Mycobacterium avium 427S and 2151SmO, and Mycobacterium bovis BCG Pasteur and Tokyo), the fibroblasts were killed by mycobacteria according to the degree of virulence. Other human originated macrophage (U-937, THP-1), myeloid (HL-60), and epithelial carcinoma (A549) cell lines exhibited a similar cytotoxic response to virulent mycobacteria. MRC-5 was most susceptible to virulent mycobacteria among various human cell lines examined. The cytotoxicity was enhanced by the proinflammatory cytokines, interleukin-1 (IL-1) and tumor necrosis factor-a (TNF-alpha), which in the absence of mycobacteria stimulate the growth of normal human fibroblasts. This in vitro evaluation system was applied to clinical isolates of drug-sensitive MTB (DS-MTB), drug-resistant MTB (DR-MTB) including multidrug-resistant (MDR-MTB), and M. avium complex (MAC). MTB strains (n = 24) exhibited strong cytotoxic activity, but MAC strains (n = 5) had only weak activity. Furthermore, there was no significant difference in cytotoxicity between DS-MTB (n = 11) and DR-MTB (n = 13). Collectively, these results suggest that this new in vitro system is useful for evaluating the pathogenesis of mycobacteria and that there was no difference in the pathogenesis between drug-susceptible and drug-resistant clinical isolates.

  12. The effect of cigarette smoke on the metabolism of arachidonic acid in isolated hamster lungs

    International Nuclear Information System (INIS)

    Maennistoe, J.; Toivonen, H.; Hartiala, J.; Bakhle, Y.S.; Uotila, P.

    1981-01-01

    The effects of cigarette smoke on the metabolism of exogenous arachidonic acid (AA) were investigated in isolated hamster lungs. Arachidonate was injected into the pulmonary circulation and the metabolites were analysed from the nonrecirculating perfusion effluent by thin layer chromatography. After the pulmonary injection of 66 nmol of 14C-AA about 20% of the injected radioactivity appeared in the perfusion effluent mostly as metabolites in six minutes. When isolated lungs were ventilated with cigarette smoke during the perfusion, the amounts of PGF2 alpha, PGE2 and two unidentified metabolite groups increased in the lung effluent. In two other experimental series hamsters were exposed to cigarette smoke before the lung perfusion either once for 30 min or during one hour daily for ten consecutive days. Neither pre-exposures caused any changes in the amounts of arachidonate metabolites in the lung effluent

  13. Global gene expression changes in human embryonic lung fibroblasts induced by organic extracts from respirable air particles

    Directory of Open Access Journals (Sweden)

    Líbalová Helena

    2012-01-01

    Full Text Available Abstract Background Recently, we used cell-free assays to demonstrate the toxic effects of complex mixtures of organic extracts from urban air particles (PM2.5 collected in four localities of the Czech Republic (Ostrava-Bartovice, Ostrava-Poruba, Karvina and Trebon which differed in the extent and sources of air pollution. To obtain further insight into the biological mechanisms of action of the extractable organic matter (EOM from ambient air particles, human embryonic lung fibroblasts (HEL12469 were treated with the same four EOMs to assess changes in the genome-wide expression profiles compared to DMSO treated controls. Method For this purpose, HEL cells were incubated with subtoxic EOM concentrations of 10, 30, and 60 μg EOM/ml for 24 hours and global gene expression changes were analyzed using human whole genome microarrays (Illumina. The expression of selected genes was verified by quantitative real-time PCR. Results Dose-dependent increases in the number of significantly deregulated transcripts as well as dose-response relationships in the levels of individual transcripts were observed. The transcriptomic data did not differ substantially between the localities, suggesting that the air pollution originating mainly from various sources may have similar biological effects. This was further confirmed by the analysis of deregulated pathways and by identification of the most contributing gene modulations. The number of significantly deregulated KEGG pathways, as identified by Goeman's global test, varied, depending on the locality, between 12 to 29. The Metabolism of xenobiotics by cytochrome P450 exhibited the strongest upregulation in all 4 localities and CYP1B1 had a major contribution to the upregulation of this pathway. Other important deregulated pathways in all 4 localities were ABC transporters (involved in the translocation of exogenous and endogenous metabolites across membranes and DNA repair, the Wnt and TGF-β signaling pathways

  14. Regulation of human lung fibroblast C1q-receptors by transforming growth factor-beta and tumor necrosis factor-alpha.

    Science.gov (United States)

    Lurton, J; Soto, H; Narayanan, A S; Raghu, G

    1999-03-01

    Transforming growth factor-beta (TGF-beta) and tumor necrosis factor-alpha (TNF-alpha) are two polypeptide mediators which are believed to play a role in the evolution of idiopathic pulmonary fibrosis (IPF). We have evaluated the effect of these two substances on the expression of receptors for collagen (cC1q-R) and globular (gC1q-R) domains of C1q and on type I collagen in human lung fibroblasts. Two fibroblast subpopulations differing in C1q receptor expression were obtained by culturing human lung explants in medium containing fresh human serum and heated plasma-derived serum and separating them based on C1q binding [Narayanan, Lurton and Raghu: Am J Resp Cell Mol Biol. 1998; 17:84]. The cells, referred to as HH and NL cells, respectively, were exposed to TGF-beta and TNF-alpha in serum-free conditions. The levels of mRNA were assessed by in situ hybridization and Northern analysis, and protein levels compared after SDS-polyacrylamide gel electrophoresis and Western blotting. NL cells exposed to TGF-beta and TNF-alpha contained 1.4 and 1.6 times as much cC1q-R mRNA, respectively, whereas in HH cells cC1q-R mRNA increased 2.0- and 2.4-fold. The gC1q-R mRNA levels increased to a lesser extent in both cells. These increases were not reflected in protein levels of CC1q-R and gC1q-R, which were similar to or less than controls. Both TGF-beta and TNF-alpha also increased procollagen [I] mRNA levels in both cells. Overall, TNF-alpha caused a greater increase and the degree of response by HH fibroblasts to both TGF-beta and TNF-alpha was higher than NL cells. These results indicated that TGF-beta and TNF-alpha upregulate the mRNA levels for cC1q-R and collagen and that they do not affect gC1q-R mRNA levels significantly. They also indicated different subsets of human lung fibroblasts respond differently to inflammatory mediators.

  15. Mechanistic study of DNA damage and radioprotection of small molecule treatment in the irradiated proliferating and quiescent human lung fibroblast cells

    International Nuclear Information System (INIS)

    Dai, Jiawen; Baskar, Rajamanickam

    2014-01-01

    Ionizing radiation is an invaluable diagnostic and treatment tool used in various clinical applications and also in cancer control. However, assessing normal tissue injury is of a great interest. Since radiation sensitivity varies with different phases of cell cycle, understanding how these cells differ in their sensitivity will help to prevent or reduce the radiation injury. We have used both proliferating and quiescent human normal lung fibroblast cells and investigated key proteins involved in the cell cycle, DNA damage and death, further the radioprotective role of small molecule after low doses (d ≤1Gy) of radiation exposure. Among the cell cycle/death proteins investigated, p53 and phosphorylation of p53 (Ser-15) were induced in both the proliferating and quiescent phases of cells when studied at different time intervals. In the proliferating cells after irradiation along with p53, cyclin dependent kinase (CDK) inhibitors p21, p27 were induced. However, similarly in the quiescent cells along with the p53, p21 and p27 were also induced. The DNA damage assessed by phosphorylation of histone H2AX expression showed an increase even in the non-dividing quiescent cells after 1 Gy of radiation exposure. Whereas cell cycle proteins Cyclin A and E and cell death proteins Bax and cytochrome-c did not show any increase in the quiescent cells. In conclusion, human normal lung fibroblast cells that are not actively dividing are also showed similar radiation response as of proliferating cells. Furthermore, proliferating and quiescent cells treated with small molecules attenuate p53 and its downstream target protein p21 indicating radioprotection of the cells. The specific activation of p53, phosphorylation of p53 (Ser-15), p21 and phosphorylation of histone H2AX following radiation doses of d ≤ 1 Gy in the quiescent cells demonstrated in this study may give us a better understanding about the radiation response of non-dividing fibroblast cells, which is present in many

  16. Effects of Perfluoroisobutene on Guinea Pig Isolated Lung Preparations.

    Science.gov (United States)

    guinea pigs showed increased sensitivity to 5-hydroxytryptamine, bradykinin, isoprenaline and a thromboxane A2 analogue, but not to carbachol, histamine, prostaglandin D2, adenosine or potassium. These changes were not detected in the TS. If PFIB-induced changes in lung pharmacology contribute to oedema, antagonists of identified mediators may be useful as pretreatments/therapies to pulmonary

  17. Lung Cancer Chemopreventive Activity of Patulin Isolated from Penicillium vulpinum

    Directory of Open Access Journals (Sweden)

    Aymeric Monteillier

    2018-03-01

    Full Text Available Lung cancer is the most lethal form of cancer in the world. Its development often involves an overactivation of the nuclear factor kappa B (NF-κB pathway, leading to increased cell proliferation, survival, mobility, and a decrease in apoptosis. Therefore, NF-κB inhibitors are actively sought after for both cancer chemoprevention and therapy, and fungi represent an interesting unexplored reservoir for such molecules. The aim of the present work was to find naturally occurring lung cancer chemopreventive compounds by investigating the metabolites of Penicillium vulpinum, a fungus that grows naturally on dung. Penicillium vulpinum was cultivated in Potato Dextrose Broth and extracted with ethyl acetate. Bioassay-guided fractionation of this extract was performed by measuring NF-κB activity using a HEK293 cell line transfected with an NF-κB-driven luciferase reporter gene. The mycotoxin patulin was identified as a nanomolar inhibitor of TNF-α-induced NF-κB activity. Immunocytochemistry and Western blot analyses revealed that its mechanism of action involved an inhibition of p65 nuclear translocation and was independent from the NF-κB inhibitor α (IκBα degradation process. Enhancing its interest in lung cancer chemoprevention, patulin also exhibited antiproliferative, proapoptotic, and antimigration effects on human lung adenocarcinoma cells through inhibition of the Wnt pathway.

  18. Bone Morphogenetic Protein (BMP-4 and BMP-7 regulate differentially Transforming Growth Factor (TGF-β1 in normal human lung fibroblasts (NHLF

    Directory of Open Access Journals (Sweden)

    Lloyd Clare M

    2010-06-01

    Full Text Available Abstract Background Airway remodelling is thought to be under the control of a complex group of molecules belonging to the Transforming Growth Factor (TGF-superfamily. The Bone Morphogenetic Proteins (BMPs belong to this family and have been shown to regulate fibrosis in kidney and liver diseases. However, the role of BMPs in lung remodelling remains unclear. BMPs may regulate tissue remodelling in asthma by controlling TGF-β-induced profibrotic functions in lung fibroblasts. Methods Cell cultures were exposed to TGF-β1 alone or in the presence of BMP-4 or BMP-7; control cultures were exposed to medium only. Cell proliferation was assessed by quantification of the incorporation of [3H]-thymidine. The expression of the mRNA encoding collagen type I and IV, tenascin C and fibronectin in normal human lung fibroblasts (NHLF was determined by real-time quantitative PCR and the main results were confirmed by ELISA. Cell differentiation was determined by the analysis of the expression of α-smooth muscle actin (α-SMA by western blot and immunohistochemistry. The effect on matrix metalloproteinase (MMP activity was assessed by zymography. Results We have demonstrated TGF-β1 induced upregulation of mRNAs encoding the extracellular matrix proteins, tenascin C, fibronectin and collagen type I and IV when compared to unstimulated NHLF, and confirmed these results at the protein level. BMP-4, but not BMP-7, reduced TGF-β1-induced extracellular matrix protein production. TGF-β1 induced an increase in the activity of the pro-form of MMP-2 which was inhibited by BMP-7 but not BMP-4. Both BMP-4 and BMP-7 downregulated TGF-β1-induced MMP-13 release compared to untreated and TGF-β1-treated cells. TGF-β1 also induced a myofibroblast-like transformation which was partially inhibited by BMP-7 but not BMP-4. Conclusions Our study suggests that some regulatory properties of BMP-7 may be tissue or cell type specific and unveil a potential regulatory role for

  19. A simple and effective protocol for fast isolation of human Tenon's fibroblasts from a single trabeculectomy biopsy - a comparison of cell behaviour in different culture media.

    Science.gov (United States)

    Przekora, Agata; Zarnowski, Tomasz; Ginalska, Grazyna

    2017-01-01

    Human Tenon's fibroblasts (HTFs) play a crucial role in wound healing. They cause postoperative scarring of the filtering bleb and are thus responsible for trabeculectomy failure. This study aimed to find an effective and fast protocol for HTF isolation from trabeculectomy biopsies. The protocol was compared with the commonly recommended HTF isolation procedure, which uses Dulbecco's modified Eagle's medium (DMEM). We used Eagle's minimum essential medium (EMEM) enriched with fibroblast growth factor (FGF), which selectively promoted the proliferation of HTF cells. A secondary goal was to compare HTF morphology, metabolism and growth during parallel cultivation of the isolated cells in FGF-enriched EMEM and DMEM. Standard procedures for HTF isolation from tissue biopsies require a 20- to 30-day culture of the explants to obtain the first monolayer. Our protocol yielded the first monolayer after approx. 15 days. More importantly, the majority of the cells were fibroblasts with only individual epithelium-derived cells present. Using FGF-enriched EMEM allowed 1.3 × 10 6 vimentin-positive fibroblasts to be obtained from a single biopsy within approx. 25 days. Using DMEM resulted in isolation failure and required exchange to FGF-enriched medium to recover the fibroblast culture. HTFs maintained in FGF-enriched EMEM also showed faster proliferation and a different type I collagen production ability compared to HTFs cultured in DMEM. Thus, FGF-enriched EMEM is recommended for fast propagation of HTFs unless the aim of the study is to assess the effect of a tested agent on proliferation ability or type I collagen production. Our fast protocol for HTF isolation allows easy setup of cell banks by researchers under laboratory conditions and could be very useful during testing of novel ophthalmologic anti-fibrotic agents in vitro. Molecular analysis of HTFs isolated from patients with known treatment histories may provide valuable information on the effects of some

  20. Lung fibroblasts may play an important role in clearing apoptotic bodies of bronchial epithelial cells generated by exposure to PHMG-P-containing solution.

    Science.gov (United States)

    Park, Eun-Jung; Park, Sung-Jin; Kim, Sanghwa; Lee, Kyuhong; Chang, Jaerak

    2018-04-01

    Polyhexamethylene guanidine (PHMG) has been widely used in the industry owing to its excellent biocidal, anti-corrosive, and anti-biofouling properties. In Korea, consumers exposed to PHMG-phosphate (PHMG-P)-containing humidifier disinfectant have begun to suffer from fibrotic lung injury-related symptoms for unknown reasons. However, no appropriate treatment has yet been found because the detail toxic mechanism has not been identified. Herein, we first studied the toxic mechanism of PHMG-P-containing solution using human normal bronchial epithelial cells (BEAS-2B cells). When exposed for 24 h, PHMG-P-containing solution rapidly decreased cell viability from around 6 h after exposure and significantly increased of the phosphatidylserine exposure and the LDH release. At 6 h of exposure, the material contained in the solution was found to be bound to the cell membrane and the inner wall of vacuoles, and damaged the cell membrane and organelles. In addition, a significant increase of IFN-γ was observed among cytokines, the expression of apoptosis-, autophagy-, and membrane and DNA damage-related proteins was also enhanced. Meanwhile, the level of intracellular ROS and the secretion of IL-8 and CXCL-1, which are chemokines for professional phagocytes, decreased. Thus, we treated dead BEAS-2B cells to lung fibroblasts (HFL-1), non-professional phagocytes, and then we observed that the dead cells rapidly attached to HFL-1 cells and were taken up. Additionally, increased secretion of IL-8 and CXCL-1 was observed in the cells. Based on these results, we suggest that pulmonary exposure to PHMG-P induces apoptosis of bronchial epithelial cells and lung fibroblasts might play an important role in the clearance of the apoptotic debris. Copyright © 2018 Elsevier B.V. All rights reserved.

  1. Active Oxygen Metabolites and Thromboxane in Phorbol Myristate Acetate Toxicity to the Isolated, Perfused Rat Lung.

    Science.gov (United States)

    Carpenter, Laurie Jean

    When administered intravenously or intratracheally to rats, rabbits and sheep, phorbol myristate acetate (PMA) produces changes in lung morphology and function are similar to those seen in humans with the adult respiratory distress syndrome (ARDS). Therefore, it is thought that information about the mechanism of ARDS development can be gained from experiments using PMA-treated animals. Currently, the mechanisms by which PMA causes pneumotoxicity are unknown. Results from other studies in rabbits and in isolated, perfused rabbit lungs suggest that PMA-induced lung injury is mediated by active oxygen species from neutrophils (PMN), whereas studies in sheep and rats suggest that PMN are not required for the toxic response. The role of PMN, active oxygen metabolites and thromboxane (TxA_2) in PMA-induced injury to isolated, perfused rat lungs (IPLs) was examined in this thesis. To determine whether PMN were required for PMA to produce toxicity to the IPL, lungs were perfused for 30 min with buffer containing various concentrations of PMA (in the presence or absence of PMN). When concentrations >=q57 ng/ml were added to medium devoid of added PMN, perfusion pressure and lung weight increased. When a concentration of PMA (14-28 ng/ml) that did not by itself cause lungs to accumulate fluid was added to the perfusion medium containing PMN (1 x 10 ^8), perfusion pressure increased, and lungs accumulated fluid. These results indicate that high concentrations of PMA produce lung injury which is independent of PMN, whereas injury induced by lower concentrations is PMN-dependent. To examine whether active oxygen species were involved in mediating lung injury induced by PMA and PMN, lungs were coperfused with the oxygen radical scavengers SOD and/or catalase. Coperfusion with either or both of these enzymes totally protected lungs against injury caused by PMN and PMA. These results suggest that active oxygen species (the hydroxyl radical in particular), mediate lung injury in

  2. Isolation of a cDNA for a Growth Factor of Vascular Endothelial Cells from Human Lung Cancer Cells: Its Identity with Insulin‐like Growth Factor II

    Science.gov (United States)

    Hagiwara, Koichi; Kobayashi, Tatsuo; Tobita, Masato; Kikyo, Nobuaki; Yazaki, Yoshio

    1995-01-01

    We have found growth‐promoting activity for vascular endothelial cells in the conditioned medium of a human lung cancer cell line, T3M‐11. Purification and characterization of the growth‐promoting activity have been carried out using ammonium sulfate precipitation and gel‐exclusion chromatography. The activity migrated as a single peak just after ribonuclease. It did not bind to a heparin affinity column. These results suggest that the activity is not a heparin‐binding growth factor (including fibroblast growth factors) or a vascular endothelial growth factor. To identify the molecule exhibiting the growth‐promoting activity, a cDNA encoding the growth factor was isolated through functional expression cloning in COS‐1 cells from a cDNA library prepared from T3M‐11 cells. The nucleotide sequence encoded by the cDNA proved to be identical with that of insulin‐like growth factor II. PMID:7730145

  3. Protective effect of enzymatic hydrolysates from highbush blueberry (Vaccinium corymbosum L.) against hydrogen peroxide-induced oxidative damage in Chinese hamster lung fibroblast cell line.

    Science.gov (United States)

    Senevirathne, Mahinda; Kim, Soo-Hyun; Jeon, You-Jin

    2010-06-01

    Blueberry was enzymatically hydrolyzed using selected commercial food grade carbohydrases (AMG, Celluclast, Termamyl, Ultraflo and Viscozyme) and proteases (Alcalase, Flavourzyme, Kojizyme, Neutrase and Protamex) to obtain water soluble compounds, and their protective effect was investigated against H(2)O(2)-induced damage in Chinese hamster lung fibroblast cell line (V79-4) via various published methods. Both AMG and Alcalase hydrolysates showed higher total phenolic content as well as higher cell viability and ROS scavenging activities, and hence, selected for further antioxidant assays. Both AMG and Alcalase hydrolysates also showed higher protective effects against lipid peroxidation, DNA damage and apoptotic body formation in a dose-dependent fashion. Thus, the results indicated that water soluble compounds obtained by enzymatic hydrolysis of blueberry possess good antioxidant activity against H(2)O(2)-induced cell damage in vitro.

  4. TCDD and a putative endogenous AhR ligand, ITE, elicit the same immediate changes in gene expression in mouse lung fibroblasts.

    Science.gov (United States)

    Henry, Ellen C; Welle, Stephen L; Gasiewicz, Thomas A

    2010-03-01

    The aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor, mediates toxicity of several classes of xenobiotics and also has important physiological roles in differentiation, reproduction, and immunity, although the endogenous ligand(s) mediating these functions is/are as yet unidentified. One candidate endogenous ligand, 2-(1'H-indolo-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE), is a potent AhR agonist in vitro, activates the murine AhR in vivo, but does not induce toxicity. We hypothesized that ITE and the toxic ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), may modify transcription of different sets of genes to account for their different toxicity. To test this hypothesis, primary mouse lung fibroblasts were exposed to 0.5muM ITE, 0.2nM TCDD, or vehicle for 4 h, and total gene expression was evaluated using microarrays. After this short-term and low-dose treatment, several hundred genes were changed significantly, and the response to ITE and TCDD was remarkably similar, both qualitatively and quantitatively. Induced gene sets included the expected battery of AhR-dependent xenobiotic-metabolizing enzymes, as well as several sets that reflect the inflammatory role of lung fibroblasts. Real time quantitative RT-qPCR assay of several selected genes confirmed these microarray data and further suggested that there may be kinetic differences in expression between ligands. These data suggest that ITE and TCDD elicit an analogous change in AhR conformation such that the initial transcription response is the same. Furthermore, if the difference in toxicity between TCDD and ITE is mediated by differences in gene expression, then it is likely that secondary changes enabled by the persistent TCDD, but not by the shorter lived ITE, are responsible.

  5. Acrolein-Exposed Normal Human Lung Fibroblasts in Vitro: Cellular Senescence, Enhanced Telomere Erosion, and Degradation of Werner’s Syndrome Protein

    Science.gov (United States)

    Jang, Jun-Ho; Bruse, Shannon; Huneidi, Salam; Schrader, Ronald M.; Monick, Martha M.; Lin, Yong; Carter, A. Brent; Klingelhutz, Aloysius J.

    2014-01-01

    Background: Acrolein is a ubiquitous environmental hazard to human health. Acrolein has been reported to activate the DNA damage response and induce apoptosis. However, little is known about the effects of acrolein on cellular senescence. Objectives: We examined whether acrolein induces cellular senescence in cultured normal human lung fibroblasts (NHLF). Methods: We cultured NHLF in the presence or absence of acrolein and determined the effects of acrolein on cell proliferative capacity, senescence-associated β-galactosidase activity, the known senescence-inducing pathways (e.g., p53, p21), and telomere length. Results: We found that acrolein induced cellular senescence by increasing both p53 and p21. The knockdown of p53 mediated by small interfering RNA (siRNA) attenuated acrolein-induced cellular senescence. Acrolein decreased Werner’s syndrome protein (WRN), a member of the RecQ helicase family involved in DNA repair and telomere maintenance. Acrolein-induced down-regulation of WRN protein was rescued by p53 knockdown or proteasome inhibition. Finally, we found that acrolein accelerated p53-mediated telomere shortening. Conclusions: These results suggest that acrolein induces p53-mediated cellular senescence accompanied by enhanced telomere attrition and WRN protein down-regulation. Citation: Jang JH, Bruse S, Huneidi S, Schrader RM, Monick MM, Lin Y, Carter AB, Klingelhutz AJ, Nyunoya T. 2014. Acrolein-exposed normal human lung fibroblasts in vitro: cellular senescence, enhanced telomere erosion, and degradation of Werner’s syndrome protein. Environ Health Perspect 122:955–962; http://dx.doi.org/10.1289/ehp.1306911 PMID:24747221

  6. Potential of the isolated lung technique for the examination of sildenafil absorption from lung-delivered poly (lactide-co-glycolide) microparticles

    DEFF Research Database (Denmark)

    Beck-Broichsitter, Moritz; Stoisiek, Katharina; Bohr, Adam

    2016-01-01

    Herein, we challenged the isolated lung (IL) technique to discriminate the performance of lung-delivered polymeric microparticles (MPs) having distinct drug release rates. For this purpose, sildenafil-loaded poly(lactide-co-glycolide) MPs were administered to the airspace of an IL model and the d...

  7. Activation of rho is involved in the mechanism of hydrogen-peroxide-induced lung edema in isolated perfused rabbit lung.

    Science.gov (United States)

    Chiba, Y; Ishii, Y; Kitamura, S; Sugiyama, Y

    2001-09-01

    Acute lung injury is attributed primarily to increased vascular permeability caused by reactive oxygen species derived from neutrophils, such as hydrogen peroxide (H2O2). Increased permeability is accompanied by the contraction and cytoskeleton reorganization of endothelial cells, resulting in intercellular gap formation. The Rho family of Ras-like GTPases is implicated in the regulation of the cytoskeleton and cell contraction. We examined the role of Rho in H2O2-induced pulmonary edema with the use of isolated perfused rabbit lungs. To our knowledge, this is the first study to examine the role of Rho in increased vascular permeability induced by H2O2 in perfused lungs. Vascular permeability was evaluated on the basis of the capillary filtration coefficient (Kfc, ml/min/cm H2O/100 g). We found that H2O2 (300 microM) increased lung weight, Kfc, and pulmonary capillary pressure. These effects of H2O2 were abolished by treatment with Y-27632 (50 microM), an inhibitor of the Rho effector p160 ROCK. In contrast, the muscular relaxant papaverine inhibited the H2O2-induced rise in pulmonary capillary pressure, but did not suppress the increases in lung weight and Kfc. These findings indicate that H2O2 causes pulmonary edema by elevating hydrostatic pressure and increasing vascular permeability. Y-27632 inhibited the formation of pulmonary edema by blocking both of these H2O2-induced effects. Our results suggest that Rho-related pathways have a part in the mechanism of H2O2-induced pulmonary edema. Copyright 2001 Academic Press.

  8. Ischemia-reperfusion injury in the isolated rat lung. Role of flow and endogenous leukocytes.

    Science.gov (United States)

    Seibert, A F; Haynes, J; Taylor, A

    1993-02-01

    Microvascular lung injury caused by ischemia-reperfusion (IR) may occur via leukocyte-dependent and leukocyte-independent pathways. Leukocyte-endothelial adhesion may be a rate-limiting step in IR lung injury. Leukocyte adhesion to microvascular endothelium occurs when the attractant forces between leukocyte and endothelium are greater than the kinetic energy of the leukocyte and the vascular wall shear rate. We hypothesized (1) that isolated, buffer-perfused rat lungs are not free of endogenous leukocytes, (2) that endogenous leukocytes contribute to IR-induced microvascular injury as measured by the capillary filtration coefficient (Kfc), and (3) that a reduction of perfusate flow rate would potentiate leukocyte-dependent IR injury. Sixty lungs were divided into four groups: (1) low-flow controls, (2) high-flow controls, (3) low-flow IR, and (4) high-flow IR. Microvascular injury was linearly related to baseline perfusate leukocyte concentrations at both low (r = 0.78) and high (r = 0.82) flow rates. Kfc in the high-flow IR group (0.58 +/- 0.03 ml/min/cm H2O/100 g) was less (p Kfc in the low-flow IR group (0.82 +/- 0.07), and in both groups Kfc values were significantly greater than low-flow (0.34 +/- 0.03) and high-flow (0.31 +/- 0.01) control Kfc values after 75 min. Retention of leukocytes in the lung, evaluated by a tissue myeloperoxidase assay, was greatest in the low-flow IR group. We conclude (1) that isolated, buffer-perfused rat lungs contain significant quantities of leukocytes and that these leukocytes contribute to IR lung injury, and (2) that IR-induced microvascular injury is potentiated by low flow.

  9. Red blood cells augment transport of reactive metabolites of monocrotaline from liver to lung in isolated and tandem liver and lung preparations

    Energy Technology Data Exchange (ETDEWEB)

    Pan, L.C.; Lame, M.W.; Morin, D.; Wilson, D.W.; Segall, H.J. (Department of Veterinary Pharmacology, University of California, Davis (United States))

    1991-09-01

    Monocrotaline (MCT) is a pyrrolizidine alkaloid that causes pulmonary hypertension in rats by mechanisms which remain largely unknown. MCT is thought to be activated in the liver to a reactive intermediate that is transported to the lung where it causes endothelial injury. The authors previous pharmacokinetic work demonstrated significant sequestration of radioactivity in red blood cells (RBCs) of rats treated with (14C)MCT. To determine whether this RBC sequestration might be important in the transport of reactive MCT metabolites, they compared the effect of inclusion of RBCs in the perfusion buffer on the extent of covalent binding of (14C)MCT to rat lungs in tandem liver-lung preparations. The potential effect of RBCs in stabilizing reactive intermediates was evaluated by preperfusion of isolated liver preparations with (14C)MCT with and without RBCs, separation and washing of the RBC fraction, and subsequent (90 min later) perfusion of washed RBCs or buffer alone in isolated perfused lungs. Covalent binding to lung tissues was determined by exhaustive methanol/chloroform extractions of unbound label from homogenized lung tissue followed by scintillation counting of residual 14C. Covalent binding was expressed as picomole MCT molecular weight equivalents/mg protein. Comparison of the relative capability of these isolated organ preparations for conversion of MCT to polar metabolites was done by extraction and HPLC analysis of perfusate at the end of the experiment. Isolated livers converted 65-85% of MCT to polar metabolites compared with less than 5% conversion in the isolated lungs. Inclusion of RBCs in the buffer of tandem lung liver preparations perfused with 400 microM (14C)MCT increased the covalent binding to the lung from 97 {plus minus} 25 (buffer alone) to 182 {plus minus} 36 (buffer + RBC) pmol/mg protein.

  10. Red blood cells augment transport of reactive metabolites of monocrotaline from liver to lung in isolated and tandem liver and lung preparations

    International Nuclear Information System (INIS)

    Pan, L.C.; Lame, M.W.; Morin, D.; Wilson, D.W.; Segall, H.J.

    1991-01-01

    Monocrotaline (MCT) is a pyrrolizidine alkaloid that causes pulmonary hypertension in rats by mechanisms which remain largely unknown. MCT is thought to be activated in the liver to a reactive intermediate that is transported to the lung where it causes endothelial injury. The authors previous pharmacokinetic work demonstrated significant sequestration of radioactivity in red blood cells (RBCs) of rats treated with [14C]MCT. To determine whether this RBC sequestration might be important in the transport of reactive MCT metabolites, they compared the effect of inclusion of RBCs in the perfusion buffer on the extent of covalent binding of [14C]MCT to rat lungs in tandem liver-lung preparations. The potential effect of RBCs in stabilizing reactive intermediates was evaluated by preperfusion of isolated liver preparations with [14C]MCT with and without RBCs, separation and washing of the RBC fraction, and subsequent (90 min later) perfusion of washed RBCs or buffer alone in isolated perfused lungs. Covalent binding to lung tissues was determined by exhaustive methanol/chloroform extractions of unbound label from homogenized lung tissue followed by scintillation counting of residual 14C. Covalent binding was expressed as picomole MCT molecular weight equivalents/mg protein. Comparison of the relative capability of these isolated organ preparations for conversion of MCT to polar metabolites was done by extraction and HPLC analysis of perfusate at the end of the experiment. Isolated livers converted 65-85% of MCT to polar metabolites compared with less than 5% conversion in the isolated lungs. Inclusion of RBCs in the buffer of tandem lung liver preparations perfused with 400 microM [14C]MCT increased the covalent binding to the lung from 97 ± 25 (buffer alone) to 182 ± 36 (buffer + RBC) pmol/mg protein

  11. Toxins secreted by Bacillus isolated from lung adenocarcinomas favor the penetration of toxic substances

    Directory of Open Access Journals (Sweden)

    Alexandra eMerlos

    2015-11-01

    Full Text Available The aim was to explore the eventual role of bacteria in the induction of lung cancer by smoking habits. Viable bacteria closely related to the genus Bacillus were detected at high frequencies in lung-cancer biopsies. Similar, if not identical, microbes were isolated from cigarettes and in smog. Bacteria present in cigarettes could be transferred to a physiological solution via a smoker device that mimicked their potential transfer during smoking those bacteria produce exotoxins able to open transmembrane pores. These channels can be used as a way to penetrate cells of benzopyrenes and other toxic substances present in tobacco products. We hypothesize that Bacillaceae present in tobacco play a key role in the development of lung cancer.

  12. Isolation and identification of a South African lentivirus from jaagsiekte lungs

    International Nuclear Information System (INIS)

    Payne, A.; York, D.F.; De Villiers, E-M.; Verwoerd, D.W.; Querat, G.; Barban, V.; Sauze, N.; Vigne, R.

    1986-01-01

    In the course of attempts to grow the jaagsiekte retrovirus in cell culture, a typical lentivirus was isolated for the first time in South Africa from adenomatous lungs. Morphologically the virus could not be distinguished from other lentiviruses, but serologically it was shown to be more closely related to visna virus than to caprine arthritis-encephalitis virus. However, a preliminary restriction enzyme analysis of the linear proviral DNA of this new lentivirus (SA-DMVV) revealed that it is significantly district from visna virus and CAEV and therefore may represent a third type of lentivirus. Antibodies to the virus were demonstrated in a number of sheep in various parts of the country, but a direct link to a disease condition was not found. Attempts to produce lung lesions by intratracheal injection of the virus have been unsuccessful to date but a transient arthritis was produced by intra-articular inoculation. Viral replication seems to be enhanced in jaagsiekte lungs

  13. Isolation of Blastomyces dermatitidis yeast from lung tissue during murine infection for in vivo transcriptional profiling.

    Science.gov (United States)

    Marty, Amber J; Wüthrich, Marcel; Carmen, John C; Sullivan, Thomas D; Klein, Bruce S; Cuomo, Christina A; Gauthier, Gregory M

    2013-07-01

    Blastomyces dermatitidis belongs to a group of thermally dimorphic fungi that grow as sporulating mold in the soil and convert to pathogenic yeast in the lung following inhalation of spores. Knowledge about the molecular events important for fungal adaptation and survival in the host remains limited. The development of high-throughput analytic tools such as RNA sequencing (RNA-Seq) has potential to provide novel insight on fungal pathogenesis especially if applied in vivo during infection. However, in vivo transcriptional profiling is hindered by the low abundance of fungal cells relative to mammalian tissue and difficulty in isolating fungal cells from the tissues they infect. For the purpose of obtaining B. dermatitidis RNA for in vivo transcriptional analysis by RNA-Seq, we developed a simple technique for isolating yeast from murine lung tissue. Using a two-step approach of filtration and centrifugation following lysis of murine lung cells, 91% of yeast cells causing infection were isolated from lung tissue. B. dermatitidis recovered from the lung yielded high-quality RNA with minimal murine contamination and was suitable for RNA-Seq. Approximately 87% of the sequencing reads obtained from the recovered yeast aligned with the B. dermatitidis genome. This was similar to 93% alignment for yeast grown in vitro. The use of near-freezing temperature along with short ex vivo time minimized transcriptional changes that would have otherwise occurred with higher temperature or longer processing time. In conclusion, we have developed a technique that recovers the majority of yeast causing pulmonary infection and yields high-quality fungal RNA with minimal contamination by mammalian RNA. Copyright © 2013 Elsevier Inc. All rights reserved.

  14. Isolated left-sided pulmonary artery agenesis with left lung hypoplasia: A report of two cases

    Directory of Open Access Journals (Sweden)

    V Govindaraj

    2017-01-01

    Full Text Available Unilateral absence of pulmonary artery or pulmonary artery agenesis (UAPA is a rare congenital malformation that can present as an isolated lesion or in association with other cardiac anomalies. Though congenital, presentation in adults are also reported. Most common presentation in adults is of exercise intolerance. The developing lung on the affected side is hypoplastic. Diagnosis of UAPA is established by imaging methods like CT and MRI . There is no specific treatment for this condition. Treatment depends on patients symptomatology, presence of pulmonary hypertension and collateral circulation. Presence of pulmonary hypertension carries a bad prognosis. We present two adult patients with isolated left sided unilateral pulmonary artery agenesis with ipsilateral lung hypoplasia. The diagnosis was confirmed by CT chest and perfusion scan.

  15. Optical measurement of isolated canine lung filtration coefficients at normal hematocrits.

    Science.gov (United States)

    Klaesner, J W; Pou, N A; Parker, R E; Finney, C; Roselli, R J

    1997-12-01

    In this study, lung filtration coefficient (Kfc) values were measured in eight isolated canine lung preparations at normal hematocrit values using three methods: gravimetric, blood-corrected gravimetric, and optical. The lungs were kept in zone 3 conditions and subjected to an average venous pressure increase of 10.24 +/- 0.27 (SE) cmH2O. The resulting Kfc (ml . min-1 . cmH2O-1 . 100 g dry lung wt-1) measured with the gravimetric technique was 0.420 +/- 0.017, which was statistically different from the Kfc measured by the blood-corrected gravimetric method (0.273 +/- 0.018) or the product of the reflection coefficient (sigmaf) and Kfc measured optically (0. 272 +/- 0.018). The optical method involved the use of a Cellco filter cartridge to separate red blood cells from plasma, which allowed measurement of the concentration of the tracer in plasma at normal hematocrits (34 +/- 1.5). The permeability-surface area product was measured using radioactive multiple indicator-dilution methods before, during, and after venous pressure elevations. Results showed that the surface area of the lung did not change significantly during the measurement of Kfc. These studies suggest that sigmafKfc can be measured optically at normal hematocrits, that this measurement is not influenced by blood volume changes that occur during the measurement, and that the optical sigmafKfc agrees with the Kfc obtained via the blood-corrected gravimetric method.

  16. Beta Adrenergic Regulation of Intrapulmonary Arteriovenous Anastomoses in Intact Rat and Isolated Rat Lungs

    Directory of Open Access Journals (Sweden)

    Melissa L. Bates

    2017-04-01

    Full Text Available Intrapulmonary arteriovenous anastomoses (IPAVA allow large diameter particles of venous origin to bypass the pulmonary capillary bed and embolize the systemic arterial circulation. IPAVA have been routinely observed in healthy humans with exercise, hypoxia, and catecholamine infusion, but the mechanism by which they are recruited is not well-defined. We hypothesized that beta-adrenergic receptor stimulation recruits IPAVA and that receptor blockade would limit hypoxia-induced IPAVA recruitment. To test our hypothesis, we evaluated the transpulmonary passage of microspheres in intact rats and isolated rats lung infused with the beta-adrenergic receptor agonist isoproterenol. We also evaluated IPAVA recruitment in intact rats with hypoxia and the beta-adrenergic receptor blocker propranolol. We found that IPAVA are recruited in the intact rat by isoproterenol and their recruitment by hypoxia can be minimized by propranolol, suggesting a role for the adrenergic system in the recruitment of IPAVA by hypoxia. IPAVA recruitment is completely abolished by ventilation with 100% oxygen. Isoproterenol also recruits IPAVA in isolated rat lungs. The fact that isoproterenol can recruit IPAVA in isolated lungs, without increased pulmonary flow, suggests that elevated cardiac output is not required for IPAVA recruitment.

  17. Radiosensitivity in cultured human fibroblasts

    International Nuclear Information System (INIS)

    Cox, R.; Masson, W.K.

    1980-01-01

    Caution is urged in the use of freshly isolated cultures of human diploid fibroblasts for quantitative studies of radiosensitivity. The distribution of x ray sensitivities of 'normal' human fibroblast cultures of foetal origin (10 subjects, skin or lung biopsy) and post-foetal origin (34 subjects, skin biopsy) are compared with the distribution in 12 patients with ataxia telangiectasia (probability of including any one of these in a normal post-foetal distribution is 0.01%). Cultures from nominally normal subjects showed a broad distribution of D 0 range of 98 +- 160 rad and assuming normal distribution, a mean +- one standard deviation of 122 +- 17 rad. Mean D 0 values for foetal origin cultures were 117 +- 12; values for post-foetal cultures D 0 were 124 +- 18. No systematic variation in D 0 was observed for age of donor, number of cell divisions in culture or for cloning efficiency. For ataxia telangiectasia D 0 values were 46 +- 7 rad. (U.K.)

  18. Identification of titanium dioxide nanoparticles in food products: induce intracellular oxidative stress mediated by TNF and CYP1A genes in human lung fibroblast cells.

    Science.gov (United States)

    Periasamy, Vaiyapuri Subbarayan; Athinarayanan, Jegan; Al-Hadi, Ahmed M; Juhaimi, Fahad Al; Mahmoud, Mohamed H; Alshatwi, Ali A

    2015-01-01

    Food grade TiO2 (E171) is a synthetic additive, and widely used as a coloring agent in many foods, pharmaceutical and personal care products. A few reports have highlighted that insoluble particulates (less than 200nm) of food grade TiO2 are found in many foods and confectionary products. However, information regarding the physico-chemical properties (i.e., size and shape)-based food grade TiO2 nanotoxicity related human health issues are limited. The main goal of this study is to examine the presence of nano-sized particulates and its structural characteristics of food grade- TiO2 materials and to assess the acute cellular uptake and metabolic stress induced by these particulates in human lung fibroblast (WI-38) cells. The results of transmission electron microscopy, energy dispersive X-ray spectroscopy, and X-ray diffraction studies indicated that about food grade TiO2 sample contains spherical shaped particulate forms in the nano-scale range, S>G0/G1) and changes in the TNF and CYP1A gene expression pattern are linked to cellular stress. Thus, food grade TiO2 as nano-scaled contaminants could not only be potential human health risk factors, suggesting that safety considerations with special respect to a few crucial factors such as size, and shape should be considered and regulated by food regulators. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. The uptake and metabolism of cystamine and taurine by isolated, ventilated and perfused rat and rabbit lungs

    International Nuclear Information System (INIS)

    Sharma, R.; Kodavanti, U.P.; Smith, L.L.; Mehendale, H.M.

    1991-01-01

    Cystamine has been reported to be taken up by the lung slices and metabolized to taurine via hypotaurine through enzymatic processes. The objective of these studies was to determine whether intact isolated, ventilated and perfused rat and rabbit lungs also posses similar characteristics. The lungs were isolated from male New Zealand white rabbits and S-D rats and perfused with 20 μM [ 14 C] cystamine (Sp. Act., 16.4 mCi/mmol) for 60 min and 30 min, respectively. Cystamine and its metabolites in lung as well as in perfusate were separated by TLC and quantitated using scintillation spectrometry. Similar experiments were also conducted with 20 μM taurine to investigate its fate in perfused lungs. Significant pulmonary uptake of cystamine and taurine occurred during perfusion. Cystamine was metabolized to [ 14 C] hypotaurine and [ 14 C] taurine. No further metabolism of taurine was evident in rat or rabbit lungs. Inclusion of 1 nM GSH did not significantly alter the ability of lungs to sequester cystamine, but the metabolism of hypotaurine to taurine was decreased. It was evident that cystamine was metabolized to taurine by perfused lungs in the same way as in lung slices

  20. Analysis of dermal fibroblasts isolated from neonatal and child cleft lip and adult skin: Developmental implications on reconstructive surgery

    Czech Academy of Sciences Publication Activity Database

    Živicová, V.; Lacina, L.; Mateu, R.; Smetana, K.; Kavkova, R.; Krejci, E.D.; Grim, M.; Kvasilová, A.; Borský, J.; Strnad, Hynek; Hradilová, Miluše; Šáchová, Jana; Kolář, Michal; Dvořánková, B.

    2017-01-01

    Roč. 40, č. 5 (2017), s. 1323-1334 ISSN 1107-3756 R&D Projects: GA ČR GA13-20293S; GA MŠk(CZ) LQ1604; GA MŠk(CZ) ED1.1.00/02.0109 Grant - others:GA MŠk(CZ) LM2015042 Institutional support: RVO:68378050 Keywords : dermal fibroblasts * myofibroblast * neonatal healing * trandforming growth factor-beta * cleft Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Cell biology Impact factor: 2.341, year: 2016

  1. Gene expression profiling following NRF2 and KEAP1 siRNA knockdown in human lung fibroblasts identifies CCL11/Eotaxin-1 as a novel NRF2 regulated gene

    Science.gov (United States)

    2012-01-01

    Background Oxidative Stress contributes to the pathogenesis of many diseases. The NRF2/KEAP1 axis is a key transcriptional regulator of the anti-oxidant response in cells. Nrf2 knockout mice have implicated this pathway in regulating inflammatory airway diseases such as asthma and COPD. To better understand the role the NRF2 pathway has on respiratory disease we have taken a novel approach to define NRF2 dependent gene expression in a relevant lung system. Methods Normal human lung fibroblasts were transfected with siRNA specific for NRF2 or KEAP1. Gene expression changes were measured at 30 and 48 hours using a custom Affymetrix Gene array. Changes in Eotaxin-1 gene expression and protein secretion were further measured under various inflammatory conditions with siRNAs and pharmacological tools. Results An anti-correlated gene set (inversely regulated by NRF2 and KEAP1 RNAi) that reflects specific NRF2 regulated genes was identified. Gene annotations show that NRF2-mediated oxidative stress response is the most significantly regulated pathway, followed by heme metabolism, metabolism of xenobiotics by Cytochrome P450 and O-glycan biosynthesis. Unexpectedly the key eosinophil chemokine Eotaxin-1/CCL11 was found to be up-regulated when NRF2 was inhibited and down-regulated when KEAP1 was inhibited. This transcriptional regulation leads to modulation of Eotaxin-1 secretion from human lung fibroblasts under basal and inflammatory conditions, and is specific to Eotaxin-1 as NRF2 or KEAP1 knockdown had no effect on the secretion of a set of other chemokines and cytokines. Furthermore, the known NRF2 small molecule activators CDDO and Sulphoraphane can also dose dependently inhibit Eotaxin-1 release from human lung fibroblasts. Conclusions These data uncover a previously unknown role for NRF2 in regulating Eotaxin-1 expression and further the mechanistic understanding of this pathway in modulating inflammatory lung disease. PMID:23061798

  2. Effects of macelignan isolated from Myristica fragrans (nutmeg) on expression of matrix metalloproteinase-1 and type I procollagen in UVB-irradiated human skin fibroblasts

    International Nuclear Information System (INIS)

    Lee, Kyung-Eun; Mun, Sukyeong; Pyun, Hee-Bong; Kim, Myung-Suk; Hwang, Jae-Kwan

    2012-01-01

    Exposure to ultraviolet (UV) light causes premature skin aging that is associated with upregulated matrix metalloproteinases (MMPs) and decreased collagen synthesis. Macelignan, a natural lignan compound isolated from Myristica fragrans HOUTT. (nutmeg), has been reported to possess antioxidant and antiinflammatory activities. This study assessed the effects of macelignan on photoaging and investigated its mechanisms of action in UV-irradiated human skin fibroblasts (Hs68) by reverse transcription-polymerase chain reaction, Western blot analysis, 2', 7'-dichlorofluorescein diacetate assay, and enzyme-linked immunosorbent assay. Our results show that macelignan attenuated UV-induced MMP-1 expression by suppressing phosphorylation of mitogen-activated protein kinases (MAPKs) induced by reactive oxygen species. Macelignan also increased type I procollagen expression and secretion through transforming growth factor β (TGF-β)/Smad signaling. These findings indicate that macelignan regulates the expression of MMP-1 and type I procollagen in UV-irradiated human skin fibroblasts by modulating MAPK and TGF-β/Smad signaling, suggesting its potential as an efficacious antiphotoaging agent. (author)

  3. Optical measurement of isolated canine lung filtration coefficients after alloxan infusion.

    Science.gov (United States)

    Klaesner, J W; Pou, N A; Parker, R E; Finney, C; Roselli, R J

    1998-04-01

    In this study, lung filtration coefficient (Kfc) was measured in eight isolated canine lung preparations by using three methods: standard gravimetric (Std), blood-corrected gravimetric (BC), and optical. The lungs were held in zone III conditions and were subjected to an average venous pressure increase of 8.79 +/- 0.93 (mean +/- SD) cmH2O. The permeability of the lungs was increased with an infusion of alloxan (75 mg/kg). The resulting Kfc values (in milliliters . min-1 . cmH2O-1 . 100 g dry lung weight-1) measured by using Std and BC gravimetric techniques before vs. after alloxan infusion were statistically different: Std, 0.527 +/- 0.290 vs. 1. 966 +/- 0.283; BC, 0.313 +/- 0.290 vs. 1.384 +/- 0.290. However, the optical technique did not show any statistical difference between pre- and postinjury with alloxan, 0.280 +/- 0.305 vs. 0.483 +/- 0. 297, respectively. The alloxan injury, quantified by using multiple-indicator techniques, showed an increase in permeability and a corresponding decrease in reflection coefficient for albumin (sigmaf). Because the optical method measures the product of Kfc and sigmaf, this study shows that albumin should not be used as an intravascular optical filtration marker when permeability is elevated. However, the optical technique, along with another means of measuring Kfc (such as BC), can be used to calculate the sigmaf of a tracer (in this study, sigmaf of 0.894 at baseline and 0.348 after injury). Another important finding of this study was that the ratio of baseline-to-injury Kfc values was not statistically different for Std and BC techniques, indicating that the percent contribution of slow blood-volume increases does not change because of injury.

  4. Nocardia kroppenstedtii sp. nov., an actinomycete isolated from a lung transplant patient with a pulmonary infection.

    LENUS (Irish Health Repository)

    Jones, Amanda L

    2014-03-01

    A novel actinomycete, strain N1286(T), isolated from a lung transplant patient with a pulmonary infection, was provisionally assigned to the genus Nocardia. The strain had chemotaxonomic and morphological properties typical of members of the genus Nocardia and formed a distinct phyletic line in the Nocardia 16S rRNA gene tree. Isolate N1286(T) was most closely related to Nocardia farcinica DSM 43665(T) (99.8% gene sequence similarity) but could be distinguished from the latter by the low level of DNA-DNA relatedness. These strains were also distinguishable on the basis of a broad range of phenotypic properties. It is concluded that strain N1286(T) represents a novel species of the genus Nocardia for which the name Nocardia kroppenstedtii sp. nov. is proposed. The type strain is N1286(T) ( = DSM 45810(T) = NCTC 13617(T)).

  5. Effect of re-expansion after short-period lung collapse on pulmonary capillary permeability and pro-inflammatory cytokine gene expression in isolated rabbit lungs.

    Science.gov (United States)

    Funakoshi, T; Ishibe, Y; Okazaki, N; Miura, K; Liu, R; Nagai, S; Minami, Y

    2004-04-01

    Re-expansion pulmonary oedema is a rare complication caused by rapid re-expansion of a chronically collapsed lung. Several cases of pulmonary oedema associated with one-lung ventilation (OLV) have been reported recently. Elevated levels of pro-inflammatory cytokines in pulmonary oedema fluid are suggested to play important roles in its development. Activation of cytokines after re-expansion of collapsed lung during OLV has not been thoroughly investigated. Here we investigated the effects of re-expansion of the collapsed lung on pulmonary oedema formation and pro-inflammatory cytokine expression. Lungs isolated from female white Japanese rabbits were perfused and divided into a basal (BAS) group (n=7, baseline measurement alone), a control (CONT) group (n=9, ventilated without lung collapse for 120 min) and an atelectasis (ATEL) group (n=9, lung collapsed for 55 min followed by re-expansion and ventilation for 65 min). Pulmonary vascular resistance (PVR) and the coefficient of filtration (Kfc) were measured at baseline and 60 and 120 min. At the end of perfusion, bronchoalveolar lavage fluid/plasma protein ratio (B/P), wet/dry lung weight ratio (W/D) and mRNA expressions of tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta and myeloperoxidase (MPO) were determined. TNF-alpha and IL-1beta mRNA were significantly up-regulated in lungs of the ATEL group compared with BAS and CONT, though no significant differences were noted in PVR, Kfc, B/P and W/D within and between groups. MPO increased at 120 min in CONT and ATEL groups. Pro-inflammatory cytokines were up-regulated upon re-expansion and ventilation after short-period lung collapse, though no changes were noted in pulmonary capillary permeability.

  6. Isoproterenol attenuates high vascular pressure-induced permeability increases in isolated rat lungs.

    Science.gov (United States)

    Parker, J C; Ivey, C L

    1997-12-01

    To separate the contributions of cellular and basement membrane components of the alveolar capillary barrier to the increased microvascular permeability induced by high pulmonary venous pressures (Ppv), we subjected isolated rat lungs to increases in Ppv, which increased capillary filtration coefficient (Kfc) without significant hemorrhage (31 cmH2O) and with obvious extravasation of red blood cells (43 cmH2O). Isoproterenol (20 microM) was infused in one group (Iso) to identify a reversible cellular component of injury, and residual blood volumes were measured to assess extravasation of red blood cells through ruptured basement membranes. In untreated lungs (High Ppv group), Kfc increased 6.2 +/- 1.3 and 38.3 +/- 15.2 times baseline during the 31 and 43 cmH2O Ppv states. In Iso lungs, Kfc was 36.2% (P Kfc increases at moderate Ppv, possibly because of an endothelial effect, but it did not affect red cell extravasation at higher vascular pressures.

  7. Detection of reactive oxygen species in isolated, perfused lungs by electron spin resonance spectroscopy

    Directory of Open Access Journals (Sweden)

    Schudt Christian

    2005-07-01

    Full Text Available Abstract Background The sources and measurement of reactive oxygen species (ROS in intact organs are largely unresolved. This may be related to methodological problems associated with the techniques currently employed for ROS detection. Electron spin resonance (ESR with spin trapping is a specific method for ROS detection, and may address some these technical problems. Methods We have established a protocol for the measurement of intravascular ROS release from isolated buffer-perfused and ventilated rabbit and mouse lungs, combining lung perfusion with the spin probe l-hydroxy-3-carboxy-2,2,5,5-tetramethylpyrrolidine (CPH and ESR spectroscopy. We then employed this technique to characterize hypoxia-dependent ROS release, with specific attention paid to NADPH oxidase-dependent superoxide formation as a possible vasoconstrictor pathway. Results While perfusing lungs with CPH over a range of inspired oxygen concentrations (1–21 %, the rate of CP• formation exhibited an oxygen-dependence, with a minimum at 2.5 % O2. Addition of superoxide dismutase (SOD to the buffer fluid illustrated that a minor proportion of this intravascular ROS leak was attributable to superoxide. Stimulation of the lungs by injection of phorbol-12-myristate-13-acetate (PMA into the pulmonary artery caused a rapid increase in CP• formation, concomitant with pulmonary vasoconstriction. Both the PMA-induced CPH oxidation and the vasoconstrictor response were largely suppressed by SOD. When the PMA challenge was performed at different oxygen concentrations, maximum superoxide liberation and pulmonary vasoconstriction occurred at 5 % O2. Using a NADPH oxidase inhibitor and NADPH-oxidase deficient mice, we illustrated that the PMA-induced superoxide release was attributable to the stimulation of NADPH oxidases. Conclusion The perfusion of isolated lungs with CPH is suitable for detection of intravascular ROS release by ESR spectroscopy. We employed this technique to

  8. Protection of ionizing radiation-induced cytogenetic damage by hydroalcoholic extract of Cynodon dactylon in Chinese hamster lung fibroblast cells and human peripheral blood lymphocytes.

    Science.gov (United States)

    Rao, Bola Sadashiva Satish; Upadhya, Dinesh; Adiga, Satish Kumar

    2008-01-01

    The radiomodulatory potential of hydroalcoholic extract of a medicinal plant Cynodon dactylon (family: Poaceae) against radiation-induced cytogenetic damage was analyzed using Chinese hamster lung fibroblast (V79) cells and human peripheral blood lymphocytes (HPBLs) growing in vitro. Induction of micronuclei was used as an index of cytogenetic damage, evaluated in cytokinesis blocked binucleate cells. The hydroalcoholic Cynodon dactylon extract (CDE) rendered protection against the radiation-induced DNA damage, as evidenced by the significant (p<0.001) reduction in micronucleated binucleate cells (MNBNC%) after various doses of CDE treatment in V79 cells and HPBLs. The optimum dose of CDE (40 and 50 microg/ml in HPBLs and V79 cells, respectively) with the greatest reduction in micronuclei was further used in combination with various doses of gamma radiation (0.5, 1, 2, 3, and 4 Gy) exposed 1 h after CDE treatment. A linear dose-dependent MNBNC% increase in radiation alone group was observed, while 40/50 microg/ml CDE significantly resulted in the reduction of MNBNC%, compared to the respective radiation alone groups. CDE resulted in a dose-dependent increase in free radical scavenging ability against various free radicals, viz., 2, 2-diphenyl-2-picryl-hydrazyl (DPPH); 2, 2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS); superoxide anion (O2*-); hydroxyl radical (OH*) and nitric oxide radical (NO*) generated in vitro. Also, an excellent (70%) inhibition of lipid peroxidation in vitro was observed at a dose of 300 microg/ml CDE, attaining the saturation point at higher doses. The present findings demonstrated the radioprotective effect of CDE, also rendering protection against radiation-induced genomic instability and DNA damage. The observed radioprotective effect may be partly attributed to the free radical scavenging and antilipid peroxidative potential of CDE.

  9. Biphasic effect of arsenite on cell proliferation and apoptosis is associated with the activation of JNK and ERK1/2 in human embryo lung fibroblast cells

    International Nuclear Information System (INIS)

    He Xiaoqing; Chen Rui; Yang Ping; Li Aiping; Zhou Jianwei; Liu Qizhan

    2007-01-01

    Biphasic dose-response relationship induced by environmental agents is often characterized with the effect of low-dose stimulation and high-dose inhibition. Some studies showed that arsenite may induce cell proliferation and apoptosis via biphasic dose-response relationship in human cells; however, mechanisms underlying this phenomenon are not well understood. In the present study, we aimed at investigating the relationship between biphasic effect of arsenite on cell proliferation and apoptosis and activation of JNK and ERK1/2 in human embryo lung fibroblast (HELF) cells. Our results demonstrated that cell proliferation may be stimulated at lower concentrations (0.1 and 0.5 μM) arsenite but inhibited at higher concentrations (5 and 10 μM). When cell apoptosis was used as the endpoint, the concentration-response curves were changed to U-shapes. During stimulation phospho-JNK levels were significantly increased at 3, 6, and 12 h after 0.1 or 0.5 μM arsenite exposure. Phospho-ERK1/2 levels were increased with different concentrations (0.1-10 μM) of arsenite at 6, 12, and 24 h. Blocking of JNK pathway with 20 μM SP600125 or ERK1/2 by 100 μM PD98059 significantly inhibited biphasic effect of arsenite in cells. Data in the present study suggest that activation of JNK and ERK1/2 may be involved in biphasic effect of arsenite when measuring cell proliferation and apoptosis in HELF cells. JNK activation seems to play a more critical role than ERK1/2 activation in the biphasic process

  10. Epigenetic modulation upon exposure of lung fibroblasts to TiO2 and ZnO nanoparticles: alterations in DNA methylation

    Directory of Open Access Journals (Sweden)

    Patil NA

    2016-09-01

    Full Text Available Nayana A Patil,1,2 WN Gade,2 Deepti D Deobagkar1 1Department of Zoology, Molecular Biology Research Laboratory, Centre of Advanced Studies, 2Department of Biotechnology, Proteomic Research Laboratory, Savitribai Phule Pune University, Pune, India Abstract: Titanium dioxide (TiO2 and zinc oxide (ZnO nanoparticles (NPs are promising candidates for numerous applications in consumer products. This will lead to increased human exposure, thus posing a threat to human health. Both these types of NPs have been studied for their cell toxicity, immunotoxicity, and genotoxicity. However, effects of these NPs on epigenetic modulations have not been studied. Epigenetics is an important link in the genotype and phenotype modulation and misregulation can often lead to lifestyle diseases. In this study, we have evaluated the DNA methylation-based epigenetic changes upon exposure to various concentrations of NPs. The investigation was designed to evaluate global DNA methylation, estimating the corresponding methyltransferase activity and expression of Dnmt gene using lung fibroblast (MRC5 cell line as lungs are the primary route of entry and target of occupational exposure to TiO2 and ZnO NPs. Enzyme-linked immunosorbent assay-based immunochemical assay revealed dose-related decrease in global DNA methylation and DNA methyltransferase activity. We also found direct correlation between the concentration of NPs, global methylation levels, and expression levels of Dnmt1, 3A, and 3B genes upon exposure. This is the first study to investigate effect of exposure to TiO2 and ZnO on DNA methylation levels in MRC5 cells. Epigenetic processes are known to play an important role in reprogramming and adaptation ability of an organism and can have long-term consequences. We suggest that changes in DNA methylation can serve as good biomarkers for early exposure to NPs since they occur at concentrations well below the sublethal levels. Our results demonstrate a clear

  11. Tezosentan reduces the microvascular filtration coefficient in isolated lungs from rats subjected to cecum ligation and puncture.

    Science.gov (United States)

    Kuklin, Vladimir; Sovershaev, Mikhail; Andreasen, Thomas; Skogen, Vegard; Ytrehus, Kirsti; Bjertnaes, Lars

    2005-01-01

    We recently demonstrated that the non-selective endothelin-1 (ET-1) receptor blocker tezosentan antagonizes ovine acute lung injury (ALI) following infusion of endotoxin or ET-1 by reducing the enhanced lung microvascular pressure, although we could not exclude the possibility of a simultaneous decline in microvascular permeability. In the present study, our aim was to find out if tezosentan reverses the rise in microvascular filtration coefficient (Kfc) in rat lungs that have been isolated and perfused 12 h after cecum ligation and puncture (CLP) or infusion of ET-1. Wistar rats (n = 42) were subjected to CLP. Postoperatively, rats were randomized to a CLP group (n = 7) and a CLP + tezosentan group (n = 7); the latter received tezosentan 30 mg/kg. A sham-operated group (n = 5) underwent laparotomy without CLP. Twelve hours postoperatively, the lungs were isolated and perfused with blood from similarly treated rats that also were used to assess plasma concentration of ET-1 and protein kinase Calpha (PKCalpha) in lung tissue. Additionally, isolated blood perfused lungs from healthy rats were randomized to a control group (n = 8), an ET-1 group (n = 7) subjected to pulmonary arterial injection of ET-1 10 nM, and an ET-1 + tezosentan group (n = 7) that received tezosentan 30 mg/kg. All lung preparations received papaverine 0.1 microg/kg added to the perfusate for vasoplegia. Pulmonary hemodynamic variables, Kfc and lung compliance (CL) were assessed. After CLP, the plasma concentration of ET-1 increased. Papaverine abolished the vasoconstrictor response to ET-1 and the pulmonary vascular pressures remained close to baseline throughout the experiments. Both CLP and injection of ET-1 caused significant changes in Kfc and CL that were prevented in tezosentan-treated rats. Compared to sham-operated animals, CLP increased the content of PKCalpha by 50% and 70% in the cytosolic and the membrane fractions of lung tissue homogenates, respectively. Tezosentan prevented the

  12. Action of the isolated canine diaphragm on the lower ribs at high lung volumes.

    Science.gov (United States)

    De Troyer, André; Wilson, Theodore A

    2014-10-15

    The normal diaphragm has an inspiratory action on the lower ribs, but subjects with chronic obstructive pulmonary disease commonly have an inward displacement of the lateral portions of the lower rib cage during inspiration. This paradoxical displacement, conventionally called 'Hoover's sign', has traditionally been attributed to the direct action of radially oriented diaphragmatic muscle fibres. In the present study, the inspiratory intercostal muscles in all interspaces in anaesthetized dogs were severed so that the diaphragm was the only muscle active during inspiration. The displacements of the lower ribs along the craniocaudal and laterolateral axes and the changes in pleural pressure (∆Ppl) and transdiaphragmatic pressure were measured during occluded breaths and mechanical ventilation at different lung volumes between functional residual capacity (FRC) and total lung capacity. From these data, the separate effects on rib displacement of ∆Ppl and of the force exerted by the diaphragm on the ribs were determined. Isolated spontaneous diaphragm contraction at FRC displaced the lower ribs cranially and outward, but this motion was progressively reversed into a caudal and inward motion as lung volume increased. However, although the force exerted by the diaphragm on the ribs decreased with increasing volume, it continued to displace the ribs cranially and outward. These observations suggest that Hoover's sign is usually caused by the decrease in the zone of apposition and, thus, by the dominant effect of ∆Ppl on the lower ribs, rather than an inward pull from the diaphragm. © 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.

  13. QUANTIFICATION OF SIDEROPHORE AND HEMOLYSIN FROM STACHYBOTRYS CHARTARUM STRAINS, INCLUDING A STRAIN ISOLATED FROM THE LUNG OF A CHILD WITH PULMONARY HEMORRHAGE AND HEMOSIDEROSIS

    Science.gov (United States)

    A strain of Stachybotrys chartarum was recently isolated from the lung of a pulmonary hemorrhage and hemosiderosis (PH) patient in Texas (designated the Houston strain). This is the first time that S. chartarum has been isolated from the lung of a PH patient. In this study, the ...

  14. Isolated lung perfusion with gemcitabine for the treatment of pulmonary metastases : Experimental study in a rat model

    NARCIS (Netherlands)

    Putte, B.P. van

    2003-01-01

    Isolated lung perfusion is an experimental surgical technique for the treatment of pulmonary metastases in order to improve the current 5-year survival of approximately 40 % after surgical resection of manually palpable metastases. Several drugs have been tested in animals models and five phase I

  15. Viral replication and lung lesions in BALB/c mice experimentally inoculated with avian metapneumovirus subgroup C isolated from chickens.

    Directory of Open Access Journals (Sweden)

    Li Wei

    Full Text Available Avian metapneumovirus (aMPV emerged as an important respiratory pathogen causing acute respiratory tract infection in avian species. Here we used a chicken aMPV subgroup C (aMPV/C isolate to inoculate experimentally BALB/c mice and found that the aMPV/C can efficiently replicate and persist in the lungs of mice for at least 21 days with a peak viral load at day 6 postinoculation. Lung pathological changes were characterized by increased inflammatory cells. Immunochemical assay showed the presence of viral antigens in the lungs and significant upregulation of pulmonary inflammatory cytokines and chemokines including MCP-1, MIP-1α, RANTES, IL-1β, IFN-γ, and TNF-α were detected following inoculation. These results indicate for the first time that chicken aMPV/C may replicate in the lung of mice. Whether aMPV/C has potential as zoonotic pathogen, further investigation will be required.

  16. Viral replication and lung lesions in BALB/c mice experimentally inoculated with avian metapneumovirus subgroup C isolated from chickens.

    Science.gov (United States)

    Wei, Li; Zhu, Shanshan; She, Ruiping; Hu, Fengjiao; Wang, Jing; Yan, Xu; Zhang, Chunyan; Liu, Shuhang; Quan, Rong; Li, Zixuan; Du, Fang; Wei, Ting; Liu, Jue

    2014-01-01

    Avian metapneumovirus (aMPV) emerged as an important respiratory pathogen causing acute respiratory tract infection in avian species. Here we used a chicken aMPV subgroup C (aMPV/C) isolate to inoculate experimentally BALB/c mice and found that the aMPV/C can efficiently replicate and persist in the lungs of mice for at least 21 days with a peak viral load at day 6 postinoculation. Lung pathological changes were characterized by increased inflammatory cells. Immunochemical assay showed the presence of viral antigens in the lungs and significant upregulation of pulmonary inflammatory cytokines and chemokines including MCP-1, MIP-1α, RANTES, IL-1β, IFN-γ, and TNF-α were detected following inoculation. These results indicate for the first time that chicken aMPV/C may replicate in the lung of mice. Whether aMPV/C has potential as zoonotic pathogen, further investigation will be required.

  17. Assessment of lung development in isolated congenital diaphragmatic hernia using signal intensity ratios on fetal MR imaging

    International Nuclear Information System (INIS)

    Balassy, Csilla; Kasprian, Gregor; Weber, Michael; Herold, Christian; Prayer, Daniela; Brugger, Peter C.; Csapo, Bence

    2010-01-01

    To investigate developmental changes in the apparently unaffected contralateral lung by using signal intensity ratios (SIR) and lung volumes (LV), and to search for correlation with clinical outcome. Twenty-five fetuses (22-37 weeks' gestation) were examined. Lung/liver signal intensity ratios (LLSIR) were assessed on T1-weighted and T2-weighted sequences for both lungs, then together with LV compared with age-matched controls of 91 fetuses by using the U test. Differences in LLSIRs and lung volumes were correlated with neonatal outcomes. LLSIRs in fetuses with congenital diaphragmatic hernia (CDH) were significantly higher in both lungs on T1-weighted images and significantly lower on T2-weighted images, compared with normals (p < 0.05), increasing on T2-weighted imaging and decreasing on T1-weighted imaging during gestation. Total LV were significantly smaller in the CDH group than in controls (p < 0.05). No significant differences in LLSIR of the two lungs were found. Outcomes correlated significantly with total LV, but not with LLSIR. Changes in LLSIR seem to reflect developmental impairment in CDH; however, they provide no additional information in predicting outcome. LV remains the best indicator on fetal MR imaging of neonatal survival in isolated, left-sided CDH. (orig.)

  18. Assessment of lung development in isolated congenital diaphragmatic hernia using signal intensity ratios on fetal MR imaging

    Energy Technology Data Exchange (ETDEWEB)

    Balassy, Csilla; Kasprian, Gregor; Weber, Michael; Herold, Christian; Prayer, Daniela [Medical University of Vienna, Department of Radiology, Vienna (Austria); Brugger, Peter C. [Medical University of Vienna, Centre of Anatomy and Cell Biology, Vienna (Austria); Csapo, Bence [Medical University of Vienna, Department of Obstetrics and Gyneocology, Vienna (Austria)

    2010-04-15

    To investigate developmental changes in the apparently unaffected contralateral lung by using signal intensity ratios (SIR) and lung volumes (LV), and to search for correlation with clinical outcome. Twenty-five fetuses (22-37 weeks' gestation) were examined. Lung/liver signal intensity ratios (LLSIR) were assessed on T1-weighted and T2-weighted sequences for both lungs, then together with LV compared with age-matched controls of 91 fetuses by using the U test. Differences in LLSIRs and lung volumes were correlated with neonatal outcomes. LLSIRs in fetuses with congenital diaphragmatic hernia (CDH) were significantly higher in both lungs on T1-weighted images and significantly lower on T2-weighted images, compared with normals (p < 0.05), increasing on T2-weighted imaging and decreasing on T1-weighted imaging during gestation. Total LV were significantly smaller in the CDH group than in controls (p < 0.05). No significant differences in LLSIR of the two lungs were found. Outcomes correlated significantly with total LV, but not with LLSIR. Changes in LLSIR seem to reflect developmental impairment in CDH; however, they provide no additional information in predicting outcome. LV remains the best indicator on fetal MR imaging of neonatal survival in isolated, left-sided CDH. (orig.)

  19. Isolation of a mannose/N-acetylglucosamine receptor from rabbit lung

    International Nuclear Information System (INIS)

    Lennartz, M.R.; Wileman, T.E.; Stahl, P.D.

    1986-01-01

    The presence of a mannose receptor on alveolar macrophages was first described in 1978 and later extended to other macrophage populations. Recently the novel ligand, mannose-conjugated lactoperoxidase, was used to identify this receptor as a 175kD protein. A 175kD protein exhibiting mannose and N-acetylglucosamine (GlcNAc)-binding properties was isolated from rabbit lung membranes. Membranes were washed with high salt, mannose and EDTA to remove endogenously bound ligand and were subsequently extracted with 1% Triton-X 100. The extract was subjected to affinity chromatography on Mannose-Sepharose followed by GlcNAc-Agarose. Triton was exchanged for 1% CHAPS while the protein was bound to GlcNAc-Agarose, allowing the eluate to be concentrated without denaturation. The eluted protein bound [ 125 I]mannose-BSA in a mannan-inhibitable fashion. Microgram quantities of protein were isolated in this fashion. SDS-PAGE revealed a major protein band at 175kD. Amino acid analysis indicates low concentrations of methionine. Results from concanavalin A binding studies and endoglycosidase F digestion suggest that the mannose receptor is a glycoprotein containing N-linked oligosaccharides

  20. Fibroblastic rheumatism

    Directory of Open Access Journals (Sweden)

    Jyoti Ranjan Parida

    2017-01-01

    Full Text Available Fibroblastic rheumatism (FR is a rare dermoarthopathy reported from different parts of the world since 1980. Although the exact cause is unknown, few reports implicate infection may be a triggering event. Patients usually present with multiple skin nodules and polyarthropathy with progressive skin contractures. Laboratory parameters including acute phase reactants are usually normal. The confirmatory diagnosis is based on histopathologic study of skin nodules, which demonstrate fibroblastic proliferation, thickened collagen fibers, dermal fibrosis, and decreased number of elastic fibers. Immunoreactivity for b-catenin, smooth muscle actin, and the monoclonal antibody HHF35 show myofibroblastic differentiation. Treatments with oral prednisolone and other disease-modifying drugs such as methotrexate, infliximab, and interferon have been tried with variable success. In general, skin lesions respond more aptly than joint symptoms indicating that skin fibroblast is more amenable to treatment than synovial fibroblasts. Awareness regarding this orphan disease among clinicians and pathologists will help in more reporting of such cases and finding out optimal treatment regimen.

  1. Effect of selenium and vitamin E deficiencies on the fate of arachidonic acid in rat isolated lungs

    International Nuclear Information System (INIS)

    Uotila, P.; Puustinen, T.

    1985-01-01

    The fate of exogenous 14 C-arachidonic acid ( 14 C-AA) was investigated in the isolated lungs of rats fed selenium and vitamin E deficient diet or diets supplemented with selenium and/or vitamin E. When 80 nmol of 14 C-AA was infused into the pulmonary circulation most of the infused 14 C-AA was found in different phospholipid and neutral lipid fractions of the perfused lungs. Only less than ten percent of the infused radioactivity was recovered in the perfusion effluent. The amount of arachidonate metabolites in the perfusion effluent was negligible, and most of the radioactivity in the perfusion effluent consisted of unmetabolized arachidonate. Selenium deficiency had no significant effect on the distribution of 14 C-AA in different lung lipid fractions. However, in the lungs of vitamin E deficient rats the amount of radioactivity was slightly increased in the neutral lipid fraction, which was due to the increased amount of 14 C-AA in the diacylglycerols. The amount of radioactivity was increased especially in the 1,3-diacylglycerols. The amount of radioactivity was increased especially in the 1,3-diacylglycerols. The amount of 14 C-AA in the triacylglycerols and in different phospholipids was not significantly changed. The present study might indicate that selenium deficiency has no significant effect on the fate of exogenous arachidonic acid in isolated rat lungs, and that vitamin E deficiency would slightly increase the amount of arachidonic acid in the diacylglycerols

  2. High-fidelity simulation of lung isolation with double-lumen endotracheal tubes and bronchial blockers in anesthesiology resident training.

    Science.gov (United States)

    Failor, Erin; Bowdle, Andrew; Jelacic, Srdjan; Togashi, Kei

    2014-08-01

    Demonstrate the feasibility of using the AirSim Bronchi airway simulator to teach residents how to manage lung isolation with double-lumen endotracheal tubes and bronchial blockers and evaluate their performance with a detailed checklist. Prospective observational study. University anesthesiology residency training program. Anesthesiology residents taking a cardiothoracic anesthesiology rotation. Residents were instructed in 7 tasks using the AirSim Bronchi: The use of the fiberoptic bronchoscope, methods for placing left and right double-lumen endotracheal tubes and 3 bronchial blockers (Univent, Arndt, and Cohen), and application of continuous positive airway pressure (CPAP) to the unventilated lung. Two to 3 weeks later, checklists and a detailed scoring system were used to assess performance. Residents rated the curriculum and their own confidence in performing the tasks using a 5-point Likert scale. Thirteen residents completed the curriculum. Their median Likert scale ratings of the curriculum based on a questionnaire with 6 items ranged from 4 to 5 of 5. Resident confidence scores for each lung isolation technique improved after the simulation training, with the median gain ranging from 0.5 to 1.5 Likert levels depending on the task. The largest improvement occurred with the bronchial blockers (psimulator in a novel simulation curriculum to teach lung-isolation techniques to anesthesiology residents and evaluated performance using a detailed checklist scoring system. This curriculum is a promising educational tool. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Apoptosis Effect of Girinimbine Isolated from Murraya koenigii on Lung Cancer Cells In Vitro

    Directory of Open Access Journals (Sweden)

    Syam Mohan

    2013-01-01

    Full Text Available Murraya koenigii Spreng has been traditionally claimed as a remedy for cancer. The current study investigated the anticancer effects of girinimbine, a carbazole alkaloid isolated from Murraya koenigii Spreng, on A549 lung cancer cells in relation to apoptotic mechanistic pathway. Girinimbine was isolated from Murraya koenigii Spreng. The antiproliferative activity was assayed using MTT and the apoptosis detection was done by annexin V and lysosomal stability assays. Multiparameter cytotoxicity assays were performed to investigate the change in mitochondrial membrane potential and cytochrome c translocation. ROS, caspase, and human apoptosis proteome profiler assays were done to investigate the apoptotic mechanism of cell death. The MTT assay revealed that the girinimbine induces cell death with an IC50 of 19.01 μM. A significant induction of early phase of apoptosis was shown by annexin V and lysosomal stability assays. After 24 h treatment with 19.01 μM of girinimbine, decrease in the nuclear area and increase in mitochondrial membrane potential and plasma membrane permeability were readily visible. Moreover the translocation of cytochrome c also was observed. Girinimbine mediates its antiproliferative and apoptotic effects through up- and downregulation of apoptotic and antiapoptotic proteins. There was a significant involvement of both intrinsic and extrinsic pathways. Moreover, the upregulation of p53 as well as the cell proliferation repressor proteins, p27 and p21, and the significant role of insulin/IGF-1 signaling were also identified. Moreover the caspases 3 and 8 were found to be significantly activated. Our results taken together indicated that girinimbine may be a potential agent for anticancer drug development.

  4. Degradation of type IV collagen by neoplastic human skin fibroblasts

    International Nuclear Information System (INIS)

    Sheela, S.; Barrett, J.C.

    1985-01-01

    An assay for the degradation of type IV (basement membrane) collagen was developed as a biochemical marker for neoplastic cells from chemically transformed human skin fibroblasts. Type IV collagen was isolated from basement membrane of Syrian hamster lung and type I collagen was isolated from rat tails; the collagens were radioactively labelled by reductive alkylation. The abilities of normal (KD) and chemically transformed (Hut-11A) human skin fibroblasts to degrade the collagens were studied. A cell-associated assay was performed by growing either normal or transformed cells in the presence of radioactively labelled type IV collagen and measuring the released soluble peptides in the medium. This assay also demonstrated that KD cells failed to synthesize an activity capable of degrading type IV collagen whereas Hut-11A cells degraded type IV collagen in a linear manner for up to 4 h. Human serum at very low concentrations, EDTA and L-cysteine inhibited the enzyme activity, whereas protease inhibitors like phenylmethyl sulfonyl fluoride, N-ethyl maleimide or soybean trypsin inhibitor did not inhibit the enzyme from Hut-11A cells. These results suggest that the ability to degrade specifically type IV collagen may be an important marker for neoplastic human fibroblasts and supports a role for this collagenase in tumor cell invasion

  5. Interleukin-1 beta Attenuates Myofibroblast Formation and Extracellular Matrix Production in Dermal and Lung Fibroblasts Exposed to Transforming Growth Factor-beta 1

    NARCIS (Netherlands)

    Mia, Masum M.; Boersema, Miriam; Bank, Ruud A.

    2014-01-01

    One of the most potent pro-fibrotic cytokines is transforming growth factor (TGF beta). TGF beta is involved in the activation of fibroblasts into myofibroblasts, resulting in the hallmark of fibrosis: the pathological accumulation of collagen. Interleukin-1 beta (IL1 beta) can influence the

  6. Common-path Fourier domain optical coherence tomography of irradiated human skin and ventilated isolated rabbit lungs

    Science.gov (United States)

    Popp, A.; Wendel, M.; Knels, L.; Knuschke, P.; Mehner, M.; Koch, T.; Boller, D.; Koch, P.; Koch, E.

    2005-08-01

    A compact common path Fourier domain optical coherence tomography (FD-OCT) system based on a broadband superluminescence diode is used for biomedical imaging. The epidermal thickening of human skin after exposure to ultraviolet radiation is measured to proof the feasibility of FD-OCT for future substitution of invasive biopsies in a long term study on natural UV skin protection. The FD-OCT system is also used for imaging lung parenchyma. FD-OCT images of a formalin fixated lung show the same alveolar structure as scanning electron microscopy images. In the ventilated and blood-free perfused isolated rabbit lung FD-OCT is used for real-time cross-sectional image capture of alveolar mechanics throughout tidal ventilation. The alveolar mechanics changing from alternating recruitment-derecruitment at zero positive end-expiratory pressure (PEEP) to persistent recruitment after applying a PEEP of 5 cm H2O is observed in the OCT images.

  7. Dehydrodiconiferyl Alcohol Isolated from Cucurbita moschata Shows Anti-adipogenic and Anti-lipogenic Effects in 3T3-L1 Cells and Primary Mouse Embryonic Fibroblasts*

    Science.gov (United States)

    Lee, Junghun; Kim, Donghyun; Choi, Jonghyun; Choi, Hyounjeong; Ryu, Jae-Ha; Jeong, Jinhyun; Park, Eun-Jin; Kim, Seon-Hee; Kim, Sunyoung

    2012-01-01

    A water-soluble extract from the stems of Cucurbita moschata, code named PG105, was previously found to contain strong anti-obesity activities in a high fat diet-induced obesity mouse model. One of its biological characteristics is that it inhibits 3T3-L1 adipocyte differentiation. To isolate the biologically active compound(s), conventional solvent fractionation was performed, and the various fractions were tested for anti-adipogenic activity using Oil Red O staining method. A single spot on thin layer chromatography of the chloroform fraction showed a potent anti-adipogenic activity. When purified, the structure of its major component was resolved as dehydrodiconiferyl alcohol (DHCA), a lignan, by NMR and mass spectrometry analysis. In 3T3-L1 cells, synthesized DHCA significantly reduced the expression of several adipocyte marker genes, including peroxisome proliferator-activated receptor γ (Pparg), CCAAT/enhancer-binding protein α (Cebpa), fatty acid-binding protein 4 (Fabp4), sterol response element-binding protein-1c (Srebp1c), and stearoyl-coenzyme A desaturase-1 (Scd), and decreased lipid accumulation without affecting cell viability. DHCA also suppressed the mitotic clonal expansion of preadipocytes (an early event of adipogenesis), probably by suppressing the DNA binding activity of C/EBPβ, and lowered the production level of cyclinA and cyclin-dependent kinase 2 (Cdk2), coinciding with the decrease in DNA synthesis and cell division. In addition, DHCA directly inhibited the expression of SREBP-1c and SCD-1. Similar observations were made, using primary mouse embryonic fibroblasts. Taken together, our data indicate that DHCA may contain dual activities, affecting both adipogenesis and lipogenesis. PMID:22262865

  8. Inflammatory Response and Barrier Dysfunction by Different e-Cigarette Flavoring Chemicals Identified by Gas Chromatography-Mass Spectrometry in e-Liquids and e-Vapors on Human Lung Epithelial Cells and Fibroblasts.

    Science.gov (United States)

    Gerloff, Janice; Sundar, Isaac K; Freter, Robert; Sekera, Emily R; Friedman, Alan E; Robinson, Risa; Pagano, Todd; Rahman, Irfan

    2017-03-01

    Recent studies suggest that electronic cigarette (e-cig) flavors can be harmful to lung tissue by imposing oxidative stress and inflammatory responses. The potential inflammatory response by lung epithelial cells and fibroblasts exposed to e-cig flavoring chemicals in addition to other risk-anticipated flavor enhancers inhaled by e-cig users is not known. The goal of this study was to evaluate the release of the proinflammatory cytokine (interleukin-8 [IL-8]) and epithelial barrier function in response to different e-cig flavoring chemicals identified in various e-cig e-liquid flavorings and vapors by chemical characterization using gas chromatography-mass spectrometry analysis. Flavorings, such as acetoin (butter), diacetyl, pentanedione, maltol (malt), ortho-vanillin (vanilla), coumarin, and cinnamaldehyde in comparison with tumor necrosis factor alpha (TNFα), were used in this study. Human bronchial epithelial cells (Beas2B), human mucoepidermoid carcinoma epithelial cells (H292), and human lung fibroblasts (HFL-1) were treated with each flavoring chemical for 24 hours. The cells and conditioned media were then collected and analyzed for toxicity (viability %), lung epithelial barrier function, and proinflammatory cytokine IL-8 release. Cell viability was not significantly affected by any of the flavoring chemicals tested at a concentration of 10 μM to 1 mM. Acetoin and diacetyl treatment induced IL-8 release in Beas2B cells. Acetoin- and pentanedione-treated HFL-1 cells produced a differential, but significant response for IL-8 release compared to controls and TNFα. Flavorings, such as ortho-vanillin and maltol, induced IL-8 release in Beas2B cells, but not in H292 cells. Of all the flavoring chemicals tested, acetoin and maltol were more potent inducers of IL-8 release than TNFα in Beas2B and HFL-1 cells. Flavoring chemicals rapidly impaired epithelial barrier function in human bronchial epithelial cells (16-HBE) as measured by electric cell surface

  9. Isoflurane administration before ischemia and during reperfusion attenuates ischemia/reperfusion-induced injury of isolated rabbit lungs.

    Science.gov (United States)

    Liu, R; Ishibe, Y; Ueda, M; Hang, Y

    1999-09-01

    To investigate the effects of isoflurane on ischemia/ reperfusion (IR)-induced lung injury, we administered isoflurane before ischemia or during reperfusion. Isolated rabbit lungs were divided into the following groups: control (n = 6), perfused and ventilated for 120 min without ischemia; ISO-control (n = 6), 1 minimum alveolar anesthetic concentration (MAC) isoflurane was administered for 30 min before 120 min continuous perfusion; IR (n = 6), ischemia for 60 min, followed by 60 min reperfusion; IR-ISO1 and IR-ISO2, ischemia followed by reperfusion and 1 MAC (n = 6) or 2 MAC (n = 6) isoflurane for 60 min; ISO-IR (n = 6), 1 MAC isoflurane was administered for 30 min before ischemia, followed by IR. During these maneuvers, we measured total pulmonary vascular resistance (Rt), coefficient of filtration (Kfc), and lung wet to dry ratio (W/D). The results indicated that administration of isoflurane during reperfusion inhibited an IR-induced increase in Kfc and W/D ratio. Furthermore, isoflurane at 2 MAC, but not 1 MAC, significantly inhibited an IR-induced increase in Rt. The administration of isoflurane before ischemia significantly attenuated the increase in IR-induced Kfc, W/D, and Rt. Our results suggest that the administration of isoflurane before ischemia and during reperfusion protects against ischemia-reperfusion-induced injury in isolated rabbit lungs.

  10. Porphyromonas bronchialis sp. nov. Isolated from Intraoperative Bronchial Fluids of a Patient with Non-Small Cell Lung Cancer.

    Science.gov (United States)

    Sato, Takuichi; Tomida, Junko; Naka, Takashi; Fujiwara, Nagatoshi; Hasegawa, Ayako; Hoshikawa, Yasushi; Matsuyama, Junko; Ishida, Naoko; Kondo, Takashi; Tanaka, Kaori; Takahashi, Nobuhiro; Kawamura, Yoshiaki

    2015-09-01

    Porphyromonas strains, including Porphyromonas-like strains, have been isolated from oral and various other systemic infections. The characterization of such strains is a crucial issue, because such information contributes to both the taxonomy of anaerobic bacteria and the clinical aspects of infectious diseases. We previously isolated four Porphyromonas-like strains from intraoperative bronchial fluids of a patient with non-small cell lung cancer. This study aimed to characterize the genetic, biochemical and chemotaxonomic aspects of these isolates. Each strain only grew under anaerobic conditions and their colony morphology was convex, 0.1-1.0 mm in diameter, light gray, and slightly glistening colony, with no black or brown pigmentation on blood agar plates after five-day incubation. The pigmentation was helpful to differentiate the isolates from other Porphyromonas, as most of Porphyromonas species show the pigmentation. In the 16S rRNA gene phylogenetic analysis (98% sequence identity of isolates indicates the same species), the four isolates were closely related to one another (99.7-100.0%), but not related to Porphyromonas (P.) catoniae, the closest species (96.9%). In addition, the DNA-DNA hybridization data revealed less than 16% similarity values between a representative isolate and the P. catoniae, indicating that the strains were genetically independent. Biochemically, the isolates could be differentiated from closely related species, i.e., P. catoniae, P. gingivalis, P. gulae, and P. pogonae, with trypsin activity (negative only in the isolates) and leucine arylamidase activity (positive only in the isolates). We therefore propose a new species to include these isolates: Porphyromonas bronchialis sp. nov.

  11. Effects of Constant Flow vs. Constant Pressure Perfusion on Fluid Filtration in Severe Hypothermic Isolated Blood-Perfused Rat Lungs.

    Science.gov (United States)

    Halsøy, Kathrine; Kondratiev, Timofey; Tveita, Torkjel; Bjertnaes, Lars J

    2016-01-01

    Victims of severe accidental hypothermia are prone to fluid extravasation but rarely develop lung edema. We hypothesize that combined hypothermia-induced increase in pulmonary vascular resistance (PVR) and a concomitant fall in cardiac output protect the lungs against edema development. Our aim was to explore in hypothermic-isolated blood-perfused rat lungs whether perfusion at constant pressure influences fluid filtration differently from perfusion at constant flow. Isolated blood-perfused rat lungs were hanging freely in a weight transducer for measuring weight changes (ΔW). Fluid filtration coefficient (Kfc), was determined by transiently elevating left atrial pressure (Pla) by 5.8 mmHg two times each during normothermia (37°C) and during hypothermia (15°C). The lung preparations were randomized to two groups. One group was perfused with constant flow (Constant flow group) and the other group with constant pulmonary artery pressure (Constant PPA group). Microvascular pressure (Pmv) was determined before and during elevation of Pla (ΔPmv) by means of the double occlusion technique. Kfc was calculated with the formula Kfc = ΔW/ΔPmv/min. All Kfc values were normalized to predicted lung weight (P LW ), which was based on body weight (BW) according to the formula: P LW  = 0.0053 BW - 0.48 and presented as Kfc PLW in mg/min/mmHg/g. At cessation, bronchoalveolar lavage (BAL) fluid/perfusate protein concentration (B/P) ratio was determined photometrically. Data were analyzed with parametric or non-parametric tests as appropriate. p  Kfc PLW and B/P ratio increased significantly by more than 10-fold during hypothermia concerted by visible signs of edema in the trachea. Hemoglobin and hematocrit increased within the Constant flow group and between the groups at cessation of the experiments. In hypothermic rat lungs perfused at constant flow, fluid filtration coefficient per gram P LW and B/P ratio increased more than 10-fold concerted by increased

  12. Mucoid Pseudomonas aeruginosa isolates maintain the biofilm formation capacity and the gene expression profiles during the chronic lung infection of CF patients

    DEFF Research Database (Denmark)

    Lee, Bao le ri; Schjerling, Charlotte K.; Kirkby, Nikolai

    2011-01-01

    Phenotypic and genotypic diversifications of Pseudomonas aeruginosa in the airways of patients with cystic fibrosis (CF) promote long-term survival of bacteria during chronic lung infection. Twelve clonally related, sequential mucoid and non-mucoid paired P. aeruginosa isolates obtained from three......-mucoid isolates observed in this particular P. aeruginosa clone reflects different adaptation strategies used by these two phenotypes in the different niches of the CF lung environment....

  13. Antifibrotic effects of crocetin in scleroderma fibroblasts and in bleomycin-induced sclerotic mice

    Directory of Open Access Journals (Sweden)

    Yinghua Song

    2013-10-01

    Full Text Available OBJECTIVE: To investigate the antifibrotic effects of crocetin in scleroderma fibroblasts and in sclerotic mice. METHODS: Skin fibroblasts that were isolated from three systemic scleroderma (SSc patients and three healthy subjects were treated with crocetin (0.1, 1 or 10 μM. Cell proliferation was measured with an MTT assay. Alpha-smooth muscle actin was detected via an immunohistochemical method. Alpha 1 (I procollagen (COL1A1, alpha 1 (III procollagen (COL3A1, matrix metalloproteinase (MMP-1 and tissue inhibitor of matrix metalloproteinase (TIMP-1 mRNA levels were measured using real-time PCR. SSc mice were established by the subcutaneous injection of bleomycin. Crocetin (50 mg/kg/d was injected intraperitoneally for 14 days. Dermal thickness and lung fibrosis were assessed with Masson's trichrome staining. Plasma ET-1 was detected with an enzyme-linked immunosorbent assay (ELISA. Skin and lung ET-1 and COL1A1 mRNA levels were measured via real-time PCR. RESULTS: Crocetin inhibited the proliferation of SSc and normal fibroblasts, an effect that increased with crocetin concentration and incubation time. Crocetin decreased the expression of α-SMA and the levels of mRNA for COL1A1, COL3A1 and matrix metalloproteinase-1, while crocetin increased TIMP-1 mRNA levels in both SSc and normal fibroblasts. Skin and lung fibrosis was induced, and the levels of ET-1 in the plasma, skin and lungs were elevated in bleomycin-injected mice. Crocetin alleviated the thickening of the dermis and lung fibrosis; decreased COL1A1 mRNA levels in the skin and lung; and simultaneously decreased ET-1 concentrations in the plasma and ET-1 mRNA levels in the skin and lungs of the bleomycin-induced sclerotic mice, especially during the early phase (weeks 1-3. CONCLUSION: Crocetin inhibits cell proliferation, differentiation and collagen production in SSc fibroblasts. Crocetin alleviates skin and lung fibrosis in a bleomycin-induced SSc mouse model, in part due to a

  14. Fibroblast-matrix interplay: Nintedanib and pirfenidone modulate the effect of IPF fibroblast-conditioned matrix on normal fibroblast phenotype.

    Science.gov (United States)

    Epstein Shochet, Gali; Wollin, Lutz; Shitrit, David

    2018-03-12

    Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with poor prognosis. Activated fibroblasts are the key effector cells in fibrosis, producing excessive amounts of collagen and extracellular matrix (ECM) proteins. Whether the ECM conditioned by IPF fibroblasts determines the phenotype of naïve fibroblasts is difficult to explore. IPF-derived primary fibroblasts were cultured on Matrigel and then cleared using ammonium hydroxide, creating an IPF-conditioned matrix (CM). Normal fibroblast CM served as control. Normal fibroblasts were cultured on both types of CM, and cell count, cell distribution and markers of myofibroblast differentiation; transforming growth factor beta (TGFβ) signalling; and ECM expression were assessed. The effects of the anti-fibrotic drugs nintedanib and pirfenidone at physiologically relevant concentrations were also explored. Normal fibroblasts cultured on IPF-CM arranged in large aggregates as a result of increased proliferation and migration. Moreover, increased levels of pSmad3, pSTAT3 (phospho signal transducer and activator of transcription 3), alpha smooth muscle actin (αSMA) and Collagen1a were found, suggesting a differentiation towards a myofibroblast-like phenotype. SB505124 (10 μmol/L) partially reversed these alterations, suggesting a TGFβ contribution. Furthermore, nintedanib at 100 nmol/L and, to a lesser extent, pirfenidone at 100 μmol/L prevented the IPF-CM-induced fibroblast phenotype alterations, suggesting an attenuation of the ECM-fibroblast interplay. IPF fibroblasts alter the ECM, thus creating a CM that further propagates an IPF-like phenotype in normal fibroblasts. This assay demonstrated differences in drug activities for approved IPF drugs at clinically relevant concentrations. Thus, the matrix-fibroblast phenotype interplay might be a relevant assay to explore drug candidates for IPF treatment. © 2018 Asian Pacific Society of Respirology.

  15. The NF-κB family member RelB regulates microRNA miR-146a to suppress cigarette smoke-induced COX-2 protein expression in lung fibroblasts.

    Science.gov (United States)

    Zago, Michela; Rico de Souza, Angela; Hecht, Emelia; Rousseau, Simon; Hamid, Qutayba; Eidelman, David H; Baglole, Carolyn J

    2014-04-21

    Diseases due to cigarette smoke exposure, including chronic obstructive pulmonary disease (COPD) and lung cancer, are associated with chronic inflammation typified by the increased expression of cyclooxygenase-2 (COX-2) protein. RelB is an NF-κB family member that suppresses cigarette smoke induction of COX-2 through an unknown mechanism. The ability of RelB to regulate COX-2 expression may be via miR-146a, a miRNA that attenuates COX-2 in lung fibroblasts. In this study we tested whether RelB attenuation of cigarette smoke-induced COX-2 protein is due to miR-146a. Utilizing pulmonary fibroblasts deficient in RelB expression, together with siRNA knock-down of RelB, we show the essential role of RelB in diminishing smoke-induced COX-2 protein expression despite robust activation of the canonical NF-κB pathway and subsequent induction of Cox-2 mRNA. RelB did not regulate COX-2 protein expression at the level of mRNA stability. Basal levels of miR-146a were significantly lower in Relb-deficient cells and cigarette smoke increased miR-146a expression only in Relb-expressing cells. Inhibition of miR-146a had no effects on Relb expression or induction of Cox-2 mRNA by cigarette smoke but significantly increased COX-2 protein. These data highlight the potential of a RelB-miR-146a axis as a novel regulatory pathway that attenuates inflammation in response to respiratory toxicants. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  16. Stereotactic body radiation therapy for isolated hilar and mediastinal non-small cell lung cancers.

    Science.gov (United States)

    Horne, Zachary D; Richman, Adam H; Dohopolski, Michael J; Clump, David A; Burton, Steven A; Heron, Dwight E

    2018-01-01

    The seminal phase II trial for pulmonary stereotactic body radiation therapy (SBRT) suggested that SBRT to central lesions resulted in unacceptable toxicity. Alternative dose-fractionation schemes have been proposed which may improve safety without compromise of efficacy. We report our institutional outcomes of SBRT for hilar/mediastinal non-small cell lung cancer (NSCLC). A retrospective review was conducted of patients with NSCLC in a hilar or mediastinal nodal station which was treated with SBRT. Patients presented with a lesion involving the hilum or mediastinum from primary or oligorecurrent NSCLC. Kaplan-Meier with log-rank testing and Cox analysis were utilized for outcomes analysis. From 2008-2015, 40 patients with median age of 70 were treated with SBRT for primary/oligorecurrent hilar/mediastinal NSCLC with median follow-up of 16.4 months. 85% presented with oligorecurrent disease at a median of 22.4 months following definitive therapy. The aortico-pulmonary window was the target in 40%, the hilum in 25%, lower paratracheal in 20%, subcarinal in 10%, and prevascular in 5%. The median dose was 48Gy in 4 fractions (range: 35-48Gy in 4-5 fractions). Median overall (OS) and progression-free (PFS) survivals were 22.7 and 13.1 months, respectively. Two-year local control was 87.7% and not significantly different between hilar and mediastinal targets. Median PFS was significantly improved in patients with hilar vs mediastinal nodal targets: 33.3 vs 8.4 months, respectively (p=0.031). OS was not statistically different between hilar and mediastinal targets (p=0.359). On multivariable analysis, hilar vs mediastinal target predicted for PFS (HR 3.045 95%CI [1.044-8.833], p=0.042), as did shorter time to presentation in patients with oligorecurrence (HR 0.983 [95%CI 0.967-1.000], p=0.049). Acute grade 3+ morbidity was seen in 3 patients (hemoptysis, pericardial/pleural effusion, heart failure) and late grade 3+ morbidity (hemoptysis) in 1 patient. Hilar

  17. [Rd7 genotyping of M. tuberculosis strains isolated from patients with lung tuberculosis in different areas of Kazakhstan].

    Science.gov (United States)

    Demkin, V V; Korneva, I N; Riazanova, Iu A; Muminov, T A; Beĭsembaeva, Sh A; Zhakipbaeva, B T; Shopaeva, G A; Dauletbakova, A M

    2008-01-01

    A three-primer PCR assay was designed for detection of possible deletions in the RD7 region of the Mycobacterium tuberculosis complex chromosome. The assay produced amplicons of different size depending on the presence or absence of the deletions. The PCR assay was applied to 176 isolates from patients with lung tuberculosis collected in different areas of Kazakhstan in summer 2004. The isolates were initially characterized by culture and biochemical tests. The RD7 genotyping results demonstrated no polymorphism and the absence of deletions in the RD7 genome region. Some strains were additionally characterized using PCR-RFLP analysis of gyrB and hsp64 genes. The RFLP-patterns obtained corresponded to the M. tuberculosis genotypes. The results of this work are consistent with certain previous studies, indicating population stability of the RD7 region in M. tuberculosis strains. Species characterization of the isolates shows that M. tuberculosis sensu stricto is the principal causative agent of human lung tuberculosis in Kazakhstan.

  18. Podoplanin-positive cancer-associated fibroblast recruitment within cancer stroma is associated with a higher number of single nucleotide variants in cancer cells in lung adenocarcinoma.

    Science.gov (United States)

    Nakasone, Shoko; Mimaki, Sachiyo; Ichikawa, Tomohiro; Aokage, Keiju; Miyoshi, Tomohiro; Sugano, Masato; Kojima, Motohiro; Fujii, Satoshi; Kuwata, Takeshi; Ochiai, Atsushi; Tsuboi, Masahiro; Goto, Koichi; Tsuchihara, Katsuya; Ishii, Genichiro

    2018-05-01

    Podoplanin-positive cancer-associated fibroblasts (CAFs) play an essential role in tumor progression. However, it is still unclear whether specific genomic alterations of cancer cells are required to recruit podoplanin-positive CAFs. The aim of this study was to investigate the relationship between the mutation status of lung adenocarcinoma cells and the presence of podoplanin-positive CAFs. Ninety-seven lung adenocarcinomas for which whole exome sequencing data were available were enrolled. First, we analyzed the clinicopathological features of the cases, and then, evaluated the relationship between genetic features of cancer cells (major driver mutations and the number of single nucleotide variants, SNVs) and the presence of podoplanin-positive CAFs. The presence of podoplanin-positive CAFs was associated with smoking history, solid predominant subtype, and lymph node metastasis. We could not find any significant correlations between major genetic mutations (EGFR, KRAS, TP53, MET, ERBB2, BRAF, and PIC3CA) in cancer cells and the presence of podoplanin-positive CAFs. However, cases with podoplanin-positive CAFs had a significantly higher number of SNVs in cancer cells than the podoplanin-negative CAFs cases (median 84 vs 37, respectively; p = 0.001). This was also detected in a non-smoker subgroup (p = 0.037). Multivariate analyses revealed that the number of SNVs in cancer cells was the only statistically significant independent predictor for the presence of podoplanin-positive CAFs (p = 0.044). In lung adenocarcinoma, the presence of podoplanin-positive CAFs was associated with higher numbers of SNVs in cancer cells, suggesting a relationship between accumulations of SNVs in cancer cells and the generation of a tumor-promoting microenvironment.

  19. An Isolated Pulmonary Hematoma Mimicking a Lung Tumor as the Initial Finding of Vascular Ehlers-Danlos Syndrome

    International Nuclear Information System (INIS)

    Kang, Eun Ju; Lee, Ki Nam; Choi, Pil Jo; Ki, Chang Seok

    2012-01-01

    The vascular type of Ehlers-Danlos syndrome (vEDS) is an uncommon inherited disorder characterized by abnormalities in type III collagen, presenting itself as arterial dissection or rupture. We report a case of an isolated pulmonary hematoma mimicking a lung tumor in an 18-year-old man which turned out to be the initial finding of vEDS. Pneumothorax and hemothorax occurred repeatedly for 15 months following the surgical removal of the mass, and were treated by repeated left upper and lower lobectomy and thoracotomy. The diagnosis of vEDS was confirmed by pathologic and genetic studies.

  20. An Isolated Pulmonary Hematoma Mimicking a Lung Tumor as the Initial Finding of Vascular Ehlers-Danlos Syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Eun Ju; Lee, Ki Nam; Choi, Pil Jo [Dept. of Radiology, Dong-A University Medicine Center, Dong-A University College of Medicine, Busan (Korea, Republic of); Ki, Chang Seok [Dept. of Radiology, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of)

    2012-09-15

    The vascular type of Ehlers-Danlos syndrome (vEDS) is an uncommon inherited disorder characterized by abnormalities in type III collagen, presenting itself as arterial dissection or rupture. We report a case of an isolated pulmonary hematoma mimicking a lung tumor in an 18-year-old man which turned out to be the initial finding of vEDS. Pneumothorax and hemothorax occurred repeatedly for 15 months following the surgical removal of the mass, and were treated by repeated left upper and lower lobectomy and thoracotomy. The diagnosis of vEDS was confirmed by pathologic and genetic studies.

  1. Sustained activation of toll-like receptor 9 induces an invasive phenotype in lung fibroblasts: possible implications in idiopathic pulmonary fibrosis.

    Science.gov (United States)

    Kirillov, Varvara; Siler, Jonathan T; Ramadass, Mahalakshmi; Ge, Lingyin; Davis, James; Grant, Geraldine; Nathan, Steven D; Jarai, Gabor; Trujillo, Glenda

    2015-04-01

    Idiopathic pulmonary fibrosis (IPF) is characterized by excessive scarring of the lung parenchyma, resulting in a steady decline of lung function and ultimately respiratory failure. The disease course of IPF is extremely variable, with some patients exhibiting stability of symptoms for prolonged periods of time, whereas others exhibit rapid progression and loss of lung function. Viral infections have been implicated in IPF and linked to disease severity; however, whether they directly contribute to progression is unclear. We previously classified patients as rapid and slow progressors on the basis of clinical features and expression of the pathogen recognition receptor, Toll-like receptor 9 (TLR9). Activation of TLR9 in vivo exacerbated IPF in mice and induced differentiation of myofibroblasts in vitro, but the mechanism of TLR9 up-regulation and progression of fibrosis are unknown. Herein, we investigate whether transforming growth factor (TGF)-β, a pleiotropic cytokine central to IPF pathogenesis, regulates TLR9 in lung myofibroblasts. Results showed induction of TLR9 expression by TGF-β in lung myofibroblasts and a distinct profibrotic myofibroblast phenotype driven by stimulation with the TLR9 agonist, CpG-DNA. Chronic TLR9 stimulation resulted in stably differentiated α-smooth muscle actin(+)/platelet-derived growth factor receptor α(+)/CD44(+)/matrix metalloproteinase-14(+)/matrix metalloproteinase-2(+) myofibroblasts, which secrete inflammatory cytokines, invade Matrigel toward platelet-derived growth factor, and resist hypoxia-induced apoptosis. These results suggest a mechanism by which TGF-β and TLR9 responses in myofibroblasts collaborate to drive rapid progression of IPF. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  2. Molecular analysis of clinical isolates previously diagnosed as Mycobacterium intracellulare reveals incidental findings of "Mycobacterium indicus pranii" genotypes in human lung infection.

    Science.gov (United States)

    Kim, Su-Young; Park, Hye Yun; Jeong, Byeong-Ho; Jeon, Kyeongman; Huh, Hee Jae; Ki, Chang-Seok; Lee, Nam Yong; Han, Seung-Jung; Shin, Sung Jae; Koh, Won-Jung

    2015-09-30

    Mycobacterium intracellulare is a major cause of Mycobacterium avium complex lung disease in many countries. Molecular studies have revealed several new Mycobacteria species that are closely related to M. intracellulare. The aim of this study was to re-identify and characterize clinical isolates from patients previously diagnosed with M. intracellulare lung disease at the molecular level. Mycobacterial isolates from 77 patients, initially diagnosed with M. intracellulare lung disease were re-analyzed by multi-locus sequencing and pattern of insertion sequences. Among the 77 isolates, 74 (96 %) isolates were designated as M. intracellulare based on multigene sequence-based analysis. Interestingly, the three remaining strains (4 %) were re-identified as "Mycobacterium indicus pranii" according to distinct molecular phylogenetic positions in rpoB and hsp65 sequence-based typing. In hsp65 sequevar analysis, code 13 was found in the majority of cases and three unreported codes were identified. In 16S-23S rRNA internal transcribed spacer (ITS) sequevar analysis, all isolates of both species were classified within the Min-A ITS sequevar. Interestingly, four of the M. intracellulare isolates harbored IS1311, a M. avium-specific element. Two of three patients infected with "M. indicus pranii" had persistent positive sputum cultures after antibiotic therapy, indicating the clinical relevance of this study. This analysis highlights the importance of precise identification of clinical isolates genetically close to Mycobacterium species, and suggests that greater attention should be paid to nontuberculous mycobacteria lung disease caused by "M. indicus pranii".

  3. Lung

    International Nuclear Information System (INIS)

    DeNardo, G.L.; Blankenship, W.J.; Burdine, J.A. Jr.; DeNardo, S.J.

    1975-01-01

    At present no simple statement can be made relative to the role of radionuclidic lung studies in the pediatric population. It is safe to assume that they will be used with increasing frequency for research and clinical applications because of their sensitivity and ready applicability to the pediatric patient. Methods comparable to those used in adults can be used in children older than 4 years. In younger children, however, a single injection of 133 Xe in solution provides an index of both regional perfusion and ventilation which is easier to accomplish. This method is particularly valuable in infants and neonates because it is rapid, requires no patient cooperation, results in a very low radiation dose, and can be repeated in serial studies. Radionuclidic studies of ventilation and perfusion can be performed in almost all children if the pediatrician and the nuclear medicine specialist have motivation and ingenuity. S []ontaneous pulmonary vascular occlusive disease which occurs in infants and pulmonary emboli in children are easily detected using radionuclides. The pathophysiologic defects of pulmonary agenesis, bronchopulmonary sequestration, and foreign body aspiration may be demonstrated by these techniques. These techniques also appear to be useful in following patients with bronchial asthma, cystic fibrosis, congenital emphysema, and postinfection pulmonary abnormalities. (auth)

  4. [Anesthetic care for fibrobronchoscopy in patients with chronic terminal respiratory insufficiency undergoing evaluation for isolated lung transplantation].

    Science.gov (United States)

    Solca, M; Elena, A; Croci, M; Damia, G

    1993-01-01

    During the first 18 month operation of the isolated lung transplantation program at or Institution, eight patients with terminal chronic respiratory failure underwent fiberoptic bronchoscopy and broncho-alveolar lavage as part of their evaluation for isolated lung transplantation. Four patients had severe obstructive, three restrictive, and one mixed, obstructive and restrictive, disease; all of them were on continuous supplemental oxygen. Procedures were performed under topical anaesthesia, with either light sedation or simple monitored anaesthesia care. Monitoring included non-invasive blood pressure measurement, pulse oximeter and precordial stethoscope. No adverse events were recorded, except in one case, when pulse oximeter reading precipitously dropped below 80%, to a minimum of 68-69%. The procedures was terminated short of its completion, and the patient was briefly assisted with manual bag ventilation on oxygen 100%. Pulse oximeter quickly returned to normal levels (above 90%), and the patient promptly recovered, without complications. The importance of monitored anaesthesia care during fiberoptic bronchoscopy (a usually benign procedure) in critically ill patients is greatly emphasized.

  5. Platelets stimulate fibroblast-mediated contraction of collagen gels

    Directory of Open Access Journals (Sweden)

    Lundahl Joachim

    2003-10-01

    Full Text Available Abstract Background Platelets are thought to play a role in a variety of inflammatory conditions in the lung, some of which may lead to fibrosis. In the current study we tested the hypothesis that whole platelets and platelet lysate can mediate remodelling of extracellular matrix in vitro by affecting fibroblast-mediated contraction of a collagen gel. We also sought to determine to what extent platelet-derived growth factor (PDGF and transforming growth factor-β (TGF-β contribute to this effect. Methods Washed platelets, isolated from healthy blood donors, and platelet lysate (freezing and thawing, were cast together with human lung fibroblasts in three-dimensional collagen gels. The gels were then released and cultured for four days. PDGF and TGF-β1 concentrations were measured in culture supernatants by ELISA. Results Both platelets and platelet lysate augmented fibroblast-mediated gel contraction in a time and concentration dependent manner (19.9% ± 0.1 (mean ± SEM of initial area vs. 48.0% ± 0.4 at 48 hours; P 1 and PDGF-AA/AB were released in co-culture. PDGF-AA/AB had a maximum release at 24 hours whereas TGF-β1 release increased with longer culture periods. Neutralising antibodies to these mediators partially inhibited platelet-induced gel contraction. Conclusion We conclude that platelets may promote remodelling of extracellular matrix in vitro and that PDGF and TGF-β partially mediate this effect, also indicating a role for other mediators. The findings may be an important mechanism in regulating repair processes after injury.

  6. An Actinobacillus pleuropneumoniae PCR typing system based on the apx and omlA genes - evaluation of isolates from lungs and tonsils of pigs

    DEFF Research Database (Denmark)

    Gram, T.; Ahrens, Peter; Andreasen, Morten

    2000-01-01

    . The PCR typing system was tested on 102 field strains of A. pleuropneumoniae isolated from lungs of diseased pigs. The serotyping results of the investigated field strains were in agreement with the apr and omlA gene patterns found in the reference strains of the bacteria, with the exception of the oml...... gene patterns and in 89% of the isolates using the omlA gene. The same serotype specific apx/omlA gene pattern was thus found in the majority of the tonsil isolates and in isolates from diseased lungs. Most of the differences in the omlA gene were found in 18 tonsil isolates of serotype 12. The oml...

  7. Low pulmonary artery flush perfusion pressure combined with high positive end-expiratory pressure reduces oedema formation in isolated porcine lungs

    International Nuclear Information System (INIS)

    Schumann, Stefan; Schließmann, Stephan J; Wagner, Giskard; Goebel, Ulrich; Priebe, Hans-Joachim; Guttmann, Josef; Kirschbaum, Andreas

    2010-01-01

    Flush perfusion of the pulmonary artery with organ protection solution is a standard procedure before lung explantation. However, rapid flush perfusion may cause pulmonary oedema which is deleterious in the lung transplantation setting. In this study we tested the hypotheses that high pulmonary perfusion pressure contributes to the development of pulmonary oedema and positive end-expiratory pressure (PEEP) counteracts oedema formation. We expected oedema formation to increase weight and decrease compliance of the lungs on the basis of a decrease in alveolar volume as fluid replaces alveolar air spaces. The pulmonary artery of 28 isolated porcine lungs was perfused with a low-potassium dextrane solution at low (mean 27 mmHg) or high (mean 40 mmHg) pulmonary artery pressure (PAP) during mechanical ventilation at low (4 cmH 2 O) or high (8 cmH 2 O) PEEP, respectively. Following perfusion and storage, relative increases in lung weight were smaller (p < 0.05) during perfusion at low PAP (62 ± 32% and 42 ± 26%, respectively) compared to perfusion at high PAP (133 ± 54% and 87 ± 30%, respectively). Compared to all other PAP–PEEP combinations, increases in lung weight were smallest (44 ± 9% and 27 ± 12%, respectively), nonlinear intratidal lung compliance was largest (46% and 17% respectively, both p < 0.05) and lung histology showed least infiltration of mononuclear cells in the alveolar septa, and least alveolar destruction during the combination of low perfusion pressure and high PEEP. The findings suggest that oedema formation during pulmonary artery flush perfusion in isolated and ventilated lungs can be reduced by choosing low perfusion pressure and high PEEP. PAP–PEEP titration to minimize pulmonary oedema should be based on lung mechanics and PAP monitoring

  8. Therapeutic Potential of Andrographolide Isolated from the Leaves of Andrographis paniculata Nees for Treating Lung Adenocarcinomas

    Directory of Open Access Journals (Sweden)

    Yu-Tang Tung

    2013-01-01

    Full Text Available Andrographolide is one of the major diterpene lactones found in Andrographis paniculata Nees and exhibits remarkable inhibitory effects on various cancers. In this study, the antipulmonary cancer effects of andrographolide were studied in a lung tumor mouse model induced by human vascular endothelial growth factor A165 (hVEGF-A165. These results demonstrated that andrographolide significantly reduced the expression of hVEGF-A165 compared with a mock group in the Clara cells of the lungs. In addition, andrographolide also decreased tumor formation by reducing VEGF, EGFR, Cyclin A, and Cyclin B expression on the transcriptional and translational levels. These results indicated that andrographolide treatment on the overexpression of VEGF can arrest the cell cycle, which induced pulmonary tumors in transgenic mice. In conclusion, the antiangiogenesis and chemotherapeutic potential of andrographolide may provide a cure for pulmonary tumors in the future.

  9. Isolation of Blastomyces dermatitidis yeast from lung tissue during murine infection for in vivo transcriptional profiling

    OpenAIRE

    Marty, Amber J.; Wüthrich, Marcel; Carmen, John C.; Sullivan, Thomas D.; Klein, Bruce S.; Cuomo, Christina A.; Gauthier, Gregory M.

    2013-01-01

    B. dermatitidis belongs to a group of thermally dimorphic fungi that grow as sporulating mold in the soil and convert to pathogenic yeast in the lung following inhalation of spores. Knowledge about the molecular events important for fungal adaptation and survival in the host remains limited. The development of high-throughput analytic tools such as RNA sequencing (RNA-Seq) has potential to provide novel insight on fungal pathogenesis especially if applied in vivo during infection. However, in...

  10. Endoscopic ultrasound with fine needle aspiration and biopsy in lung cancer and isolated mediastinal lymphadenopathy.

    LENUS (Irish Health Repository)

    Nadarajan, P

    2010-03-01

    Endoscopic ultrasound with fine-needle aspiration and biopsy (EUS-FNAB) is well established in diagnosing and staging lung cancer in patients with mediastinal adenopathy. EUS-FNAB is highly sensitive, less invasive and has lower complication rates when compared to surgical staging of mediastinal nodes. In this study we describe our experience of EUS-FNAB in lung cancer and other causes of mediastinal lymphadenopathy. EUS-FNAB was performed for assessment of PET positive mediastinal lymph nodes between January 2007 and March 2009 in AMNCH. The endpoints of our study were sensitivity and specificity of EUS-FNAB, morbidity and length of hospital stay. Thirty four patients underwent EUS-FNAB during the study period for both diagnosis and staging. Thirty patients had positive lymph node invasion and 4 had no evidence of malignant invasion. In these 4 patients negative cytology was confirmed on mediastinoscopy giving EUS-FNAB a sensitivity and specificity of 100%. EUS-FNAB upstaged the disease in 12 patients. EUS-FNAB is a reliable tool for mediastinal staging in lung cancer, significantly reducing the need for surgical staging procedures in patients with suspected mediastinal involvement.

  11. Identification of leukotriene D4 specific binding sites in the membrane preparation isolated from guinea pig lung

    International Nuclear Information System (INIS)

    Mong, S.; Wu, H.L.; Clark, M.A.; Stadel, J.M.; Gleason, J.G.; Crooke, S.T.

    1984-01-01

    A radioligand binding assay has been established to study leukotriene specific binding sites in the guinea pig and rabbit tissues. Using high specific activity [ 3 H]-leukotriene D4 [( 3 H]-LTD4), in the presence or absence of unlabeled LTD4, the diastereoisomer of LTD4 (5R,6S-LTD4), leukotriene E4 (LTE4) and the end-organ antagonist, FPL 55712, the authors have identified specific binding sites for [ 3 H]-LTD4 in the crude membrane fraction isolated from guinea pig lung. The time required for [ 3 H]-LTD4 binding to reach equilibrium was approximately 20 to 25 min at 37 degrees C in the presence of 10 mM Tris-HCl buffer (pH 7.5) containing 150 mM NaCl. The binding of [ 3 H]-LTD4 to the specific sites was saturable, reversible and stereospecific. The maximal number of binding sites (Bmax), derived from Scatchard analysis, was approximately 320 +/- 200 fmol per mg of crude membrane protein. The dissociation constants, derived from kinetic and saturation analyses, were 9.7 nM and 5 +/- 4 nM, respectively. The specific binding sites could not be detected in the crude membrane fraction prepared from guinea pig ileum, brain and liver, or rabbit lung, trachea, ileum and uterus. In radioligand competition experiments, LTD4, FPL 55712 and 5R,6S-LTD4 competed with [ 3 H]-LTD4. The metabolic inhibitors of arachidonic acid and SKF 88046, an antagonist of the indirectly-mediated actions of LTD4, did not significantly compete with [ 3 H]-LTD4 at the specific binding sites. These correlations indicated that these specific binding sites may be the putative leukotriene receptors in the guinea-pig lung

  12. Cyclic-AMP mediated drugs: differential or global reduction of eicosanoid synthesis in the isolated rat lung?

    Directory of Open Access Journals (Sweden)

    Mark J. Post

    1992-01-01

    Full Text Available In this study the question was addressed whether cAMP mediated drugs induce a differential reduction of branches of the arachidonic acid metabolism rather than a global reduction of eicosanoid synthesis. The isolated lungs of actively sensitized rats were employed to study prostaglandin and leukotriene release in the presence and absence of the cAMP mediated drugs theophylline, milrinone, sulmazole, isobutyl-methylxanthine and salbutamol. The release of eicosanoids as measured by RIA was predominantly basal and continuous, with a mild antigen induced stimulation only for TXB2 and the leukotrienes. All drugs reduced eicosanoid release globally. It is concluded that cAMP mediated drugs interfere with arachidonic acid metabolism at a site proximal to the branching into lipoxygenase and cyclo-oxygenase pathways.

  13. MicroRNA expression in lung tissue and blood isolated from pigs suffering from bacterial pneumonia

    DEFF Research Database (Denmark)

    Skovgaard, Kerstin; Wendt, Karin Tarp; Heegaard, Peter M. H.

    . No pathological changes were seen in lungs from control animals. All AP infected animals had a significantly higher level of mRNA coding for the acute-phase protein SAA-2 in the liver compared to the control group. Whole Blood samples were collected in PAXgene Blood RNA Tubes (PrenalytiX) before (control...... samples before and after infection were labelled with Hy5™ and Hy3™ fluorescent label, respectively, using the miRCURY™ LNA Array power labelling kit (Exiqon, Denmark). Sample were mixed pair-wise and hybridized to miRCURY™ LNA array version 11.0 (Exiqon, Denmark), which contains capture probes targeting...

  14. Chromosome aberration induction in human diploid fibroblast and epithelial cells

    International Nuclear Information System (INIS)

    Scott, D.

    1986-01-01

    The relative sensitivity of cultured human fibroblasts and epithelial cells to radiation-induced chromosomal aberrations was investigated. Lung fibroblast and kidney epithelial cells from the same fetus were compared, as were skin fibroblasts and epithelial keratinocytes from the same foreskin sample. After exposure of proliferating fetal cells to 1.5 Gy X-rays there was a very similar aberration yield in the fibroblasts and epithelial cells. Observations of either little or no difference in chromosomal sensitivity between human fibroblasts and epithelial cells give added confidence that quantitative cytogenetic data obtained from cultured fibroblasts are relevant to the question of sensitivity of epithelial cells which are the predominant cell type in human cancers. (author)

  15. Gadolinium prevents high airway pressure-induced permeability increases in isolated rat lungs.

    Science.gov (United States)

    Parker, J C; Ivey, C L; Tucker, J A

    1998-04-01

    To determine the initial signaling event in the vascular permeability increase after high airway pressure injury, we compared groups of lungs ventilated at different peak inflation pressures (PIPs) with (gadolinium group) and without (control group) infusion of 20 microM gadolinium chloride, an inhibitor of endothelial stretch-activated cation channels. Microvascular permeability was assessed by using the capillary filtration coefficient (Kfc), a measure of capillary hydraulic conductivity. Kfc was measured after ventilation for 30-min periods with 7, 20, and 30 cmH2O PIP with 3 cmH2O positive end-expiratory pressure and with 35 cmH2O PIP with 8 cmH2O positive end-expiratory pressure. In control lungs, Kfc increased significantly to 1.8 and 3.7 times baseline after 30 and 35 cmH2O PIP, respectively. In the gadolinium group, Kfc was unchanged from baseline (0.060 +/- 0.010 ml . min-1 . cmH2O-1 . 100 g-1) after any PIP ventilation period. Pulmonary vascular resistance increased significantly from baseline in both groups before the last Kfc measurement but was not different between groups. These results suggest that microvascular permeability is actively modulated by a cellular response to mechanical injury and that stretch-activated cation channels may initiate this response through increases in intracellular calcium concentration.

  16. Proliferation and mitogenic response to PDGF-BB of fibroblasts isolated from chronic venous leg ulcers is ulcer-age dependent

    DEFF Research Database (Denmark)

    Agren, M S; Steenfos, H H; Dabelsteen, S

    1999-01-01

    Several pathophysiologic mechanisms have been proposed to explain slow-healing leg ulcers, but little is known about the growth behavior of cells in these wounds. Platelet-derived growth factor-BB applied topically to chronic wounds has shown beneficial effects, although the effects have been less...... pronounced than would have been expected based on studies on acute wounds. The objective of this study was to compare fibroblasts in culture obtained from chronic wounds (non-healing chronic venous leg ulcers), acute wounds and normal dermis regarding growth, mitogenic response to platelet-derived growth...... chronic wounds have approached or even reached the end of their lifespan (phase III). This might provide one explanation for the non-healing state and therapy resistance to topical platelet-derived growth factor-BB of some venous leg ulcers....

  17. The Effect of the Isolated Aorticopulmonary Lymph Node on Survival in Lung Cancer

    Directory of Open Access Journals (Sweden)

    Serdar Ozkan

    2014-12-01

    Full Text Available Aim: This study aims to investigate investigate the effects of aorticopulmonary LN metastasis and other N1 and N2 LN involvements on survival rates especially for left upper lobe tumors. Material and Method: 111 cases who underwent surgery due to NSCLC and were diagnosed with lymph node metastasis secondary to the postoperative pathological examination, were examined retrospectively. The cases on whom complete resection and mediastinal lymph node dissection were applied and who were diagnosed with postoperative mediastinal LN metastasis were examined with regard to the effects of some prognostic factors on survival. Results: 13 of the cases who were followed up for 21.41 months on average lost their lives. In the general survival analysis, it was found that isolated aorticopulmonary LN metastasis did not affect survival differently from other N2 diseases. Discussion: This paper claims that in cases with NSCLC located on the left upper lobe, isolated aorticopulmonary LN involvement does not have a negative effect on survival different from other N2 stations but further studies need for support this idea. Therefore, these cases should not be considered as inoperable and complete resection should be performed on the appropriate patients.

  18. 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.

    Science.gov (United States)

    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

  19. In vitro culture and characterization of human lung cancer circulating tumor cells isolated by size exclusion from an orthotopic nude-mouse model expressing fluorescent protein.

    Science.gov (United States)

    Kolostova, Katarina; Zhang, Yong; Hoffman, Robert M; Bobek, Vladimir

    2014-09-01

    In the present study, we demonstrate an animal model and recently introduced size-based exclusion method for circulating tumor cells (CTCs) isolation. The methodology enables subsequent in vitro CTC-culture and characterization. Human lung cancer cell line H460, expressing red fluorescent protein (H460-RFP), was orthotopically implanted in nude mice. CTCs were isolated by a size-based filtration method and successfully cultured in vitro on the separating membrane (MetaCell®), analyzed by means of time-lapse imaging. The cultured CTCs were heterogeneous in size and morphology even though they originated from a single tumor. The outer CTC-membranes were blebbing in general. Abnormal mitosis resulting in three daughter cells was frequently observed. The expression of RFP ensured that the CTCs originated from lung tumor. These readily isolatable, identifiable and cultivable CTCs can be used to characterize individual patient cancers and for screening of more effective treatment.

  20. Dysregulated proinflammatory and fibrogenic phenotype of fibroblasts in cystic fibrosis.

    Directory of Open Access Journals (Sweden)

    François Huaux

    Full Text Available Morbi-mortality in cystic fibrosis (CF is mainly related to chronic lung infection and inflammation, uncontrolled tissue rearrangements and fibrosis, and yet the underlying mechanisms remain largely unknown. We evaluated inflammatory and fibrosis responses to bleomycin in F508del homozygous and wild-type mice, and phenotype of fibroblasts explanted from mouse lungs and skin. The effect of vardenafil, a cGMP-specific phosphodiesterase type 5 inhibitor, was tested in vivo and in culture. Responses of proinflammatory and fibrotic markers to bleomycin were enhanced in lungs and skin of CF mice and were prevented by treatment with vardenafil. Purified lung and skin fibroblasts from CF mice proliferated and differentiated into myofibroblasts more prominently and displayed higher sensitivity to growth factors than those recovered from wild-type littermates. Under inflammatory stimulation, mRNA and protein expression of proinflammatory mediators were higher in CF than in wild-type fibroblasts, in which CFTR expression reached similar levels to those observed in other non-epithelial cells, such as macrophages. Increased proinflammatory responses in CF fibroblasts were reduced by half with submicromolar concentrations of vardenafil. Proinflammatory and fibrogenic functions of fibroblasts are upregulated in CF and are reduced by vardenafil. This study provides compelling new support for targeting cGMP signaling pathway in CF pharmacotherapy.

  1. Phage T4 endonuclease V stimulates DNA repair replication in isolated nuclei from ultraviolet-irradiated human cells, including xeroderma pigmentosum fibroblasts

    International Nuclear Information System (INIS)

    Smith, C.A.; Hanawalt, P.C.

    1978-01-01

    The repair mode of DNA replication has been demonstrated in isolated nuclei from uv-irradiated human cells. Nuclei are incubated in a mixture containing [ 3 H]thymidine triphosphate and bromodeoxyuridine triphosphate in a 1:5 ratio. The 3 H at the density of parental DNA in alkaline CsCl density gradients is then a measure of repair. In nuclei prepared from WI38 cells 30 min after irradiation, repair replication is uv-dependent and proceeds at approximately the in vivo rate for 5 min. Repair replication is reduced in irradiated nuclei or in nuclei prepared immediately after irradiation. It is Mg 2+ -dependent and stimulated by added ATP and deoxyribonucleoside triphosphates. No repair replication is observed in nuclei from xeroderma pigmentosum (complementation group A) cells. However, upon addition of coliphage T4 endonuclease V, which specifically nicks DNA containing pyrimidine dimers, repair replication is observed in nuclei from irradiated xeroderma pigmentosum cells and is stimulated in WI38 nuclei. The reaction then persists for an hour and is dependent upon added ATP and deoxyribonucleoside triphosphates. The repair label is in stretches of roughly 35 nucleotides, as it is in intact cells. Added pancreatic DNase does not promote uv-dependent repair synthesis. Our results support the view that xeroderma pigmentosum (group A) cells are defective in the incision step of the DNA excision repair pathway, and demonstrate the utility of this system for probing DNA repair mechanisms

  2. Growth Patterns of Fetal Lung Volumes in Healthy Fetuses and Fetuses With Isolated Left-Sided Congenital Diaphragmatic Hernia.

    Science.gov (United States)

    Ruano, Rodrigo; Britto, Ingrid Schwach Werneck; Sananes, Nicolas; Lee, Wesley; Sangi-Haghpeykar, Haleh; Deter, Russell L

    2016-06-01

    To evaluate fetal lung growth using 3-dimensional sonography in healthy fetuses and those with congenital diaphragmatic hernia (CDH). Right and total lung volumes were serially evaluated by 3-dimensional sonography in 66 healthy fetuses and 52 fetuses with left-sided CDH between 20 and 37 weeks' menstrual age. Functions fitted to these parameters were compared for 2 groups: (1) healthy versus those with CDH; and (2) fetuses with CHD who survived versus those who died. Fetal right and total lung volumes as well as fetal observed-to-expected right and total lung volume ratios were significantly lower in fetuses with CDH than healthy fetuses (Pvolume ratios did not vary with menstrual age in healthy fetuses or in those with CDH (independent of outcome). Lung volume rates were lower in fetuses with left-sided CDH compared to healthy fetuses, as well as in fetuses with CDH who died compared to those who survived. The observed-to-expected right and total lung volume ratios were relatively constant throughout menstrual age in fetuses with left-sided CDH, suggesting that the origin of their lung growth abnormalities occurred before 20 weeks and did not progress. The observed-to-expected ratios may be useful in predicting the outcome in fetuses with CDH independent of menstrual age. © 2016 by the American Institute of Ultrasound in Medicine.

  3. Slow loaded breathing training improves blood pressure, lung capacity and arm exercise endurance for older people with treated and stable isolated systolic hypertension.

    Science.gov (United States)

    Ublosakka-Jones, Chulee; Tongdee, Phailin; Pachirat, Orathai; Jones, David A

    2018-03-28

    Hypertension and reduced lung function are important features of aging. Slow loaded breathing training reduces resting blood pressure and the question is whether this can also improve lung function. Thirty-two people (67 ± 5 years, 16 male) with controlled isolated systolic hypertension undertook an eight weeks randomised controlled training trial with an inspiratory load of 25% maximum inspiratory pressure (MIP) at 6 breaths per minute (slow loaded breathing; SLB) or deep breathing control (CON). Outcome measures were resting blood pressure (BP) and heart rate; MIP; lung capacity; chest and abdominal expansion; arm cranking exercise endurance at 50% heart rate reserve. Home based measurement of resting systolic BP decreased by 20 mm Hg (15 to 25) (Mean and 95%CI) for SLB and by 5 mm Hg (1 to 7) for CON. Heart rate and diastolic BP also decreased significantly for SLB but not CON. MIP increased by 15.8 cm H 2 O (11.8 to 19.8) and slow vital capacity by 0.21 L (0.15 to 0.27) for SLB but not for CON. Chest and abdominal expansion increased by 2.3 cm (2.05 to 2.55) and 2.5 cm (2.15 to 2.85), respectively for SLB and by 0.5 cm (0.26 to 0.74) and 1.7 cm (1.32 to 2.08) for CON. Arm exercise time increased by 4.9 min (3.65 to 5.15) for SLB with no significant change for CON. Slow inspiratory muscle training is not only effective in reducing resting BP, even in older people with well controlled isolated systolic hypertension but also increases inspiratory muscle strength, lung capacity and arm exercise duration. Copyright © 2018. Published by Elsevier Inc.

  4. Anti-fibrotic effect of pirfenidone in muscle derived-fibroblasts from Duchenne muscular dystrophy patients.

    Science.gov (United States)

    Zanotti, Simona; Bragato, Cinzia; Zucchella, Andrea; Maggi, Lorenzo; Mantegazza, Renato; Morandi, Lucia; Mora, Marina

    2016-01-15

    Tissue fibrosis, characterized by excessive deposition of extracellular matrix proteins, is the end point of diseases affecting the kidney, bladder, liver, lung, gut, skin, heart and muscle. In Duchenne muscular dystrophy (DMD), connective fibrotic tissue progressively substitutes muscle fibers. So far no specific pharmacological treatment is available for muscle fibrosis. Among promising anti-fibrotic molecules, pirfenidone has shown anti-fibrotic and anti-inflammatory activity in animal and cell models, and has already been employed in clinical trials. Therefore we tested pirfenidone anti-fibrotic properties in an in vitro model of muscle fibrosis. We evaluated effect of pirfenidone on fibroblasts isolated from DMD muscle biopsies. These cells have been previously characterized as having a pro-fibrotic phenotype. We tested cell proliferation and migration, secretion of soluble collagens, intracellular levels of collagen type I and fibronectin, and diameter of 3D fibrotic nodules. We found that pirfenidone significantly reduced proliferation and cell migration of control and DMD muscle-derived fibroblasts, decreased extracellular secretion of soluble collagens by control and DMD fibroblasts, as well as levels of collagen type I and fibronectin, and, in DMD fibroblasts only, reduced synthesis and deposition of intracellular collagen. Furthermore, pirfenidone was able to reduce the diameter of fibrotic-nodules in our 3D model of in vitro fibrosis. These pre-clinical results indicate that pirfenidone has potential anti-fibrotic effects also in skeletal muscle fibrosis, urging further studies in in vivo animal models of muscular dystrophy in order to translate the drug into the treatment of muscle fibrosis in DMD patients. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Synchronous Oligometastatic Non-Small Cell Lung Cancer and Isolated Renal Cell Carcinoma: A Case Report and Literature Review.

    Science.gov (United States)

    Nguyen, Timothy K; Louie, Alexander V

    2015-10-27

    A 58-year-old gentleman presenting with a progressive headache, visual disturbance, decreased appetite, and weight loss was found to have a localized clear cell carcinoma of the kidney and synchronous Stage IV non-small cell lung cancer with a solitary brain metastasis. This case illustrates the challenges in distinguishing between primary and metastatic disease in a patient with both renal cell carcinoma and lung cancer. We highlight the uncertainties in the diagnosis and management of this unique clinical scenario and the potential implications on prognosis.

  6. The fibroblast surface markers FAP, anti-fibroblast, and FSP are expressed by cells of epithelial origin and may be altered during epithelial-to-mesenchymal transition.

    Science.gov (United States)

    Kahounová, Zuzana; Kurfürstová, Daniela; Bouchal, Jan; Kharaishvili, Gvantsa; Navrátil, Jiří; Remšík, Ján; Šimečková, Šárka; Študent, Vladimír; Kozubík, Alois; Souček, Karel

    2017-04-06

    The identification of fibroblasts and cancer-associated fibroblasts from human cancer tissue using surface markers is difficult, especially because the markers used currently are usually not expressed solely by fibroblasts, and the identification of fibroblast-specific surface molecules is still under investigation. It was aimed to compare three commercially available antibodies in the detection of different surface epitopes of fibroblasts (anti-fibroblast, fibroblast activation protein α, and fibroblast surface protein). The specificity of their expression, employing fibroblast cell lines and tumor-derived fibroblasts from breast and prostate tissues was investigated. Both the established fibroblast cell line HFF-1 and ex vivo primary fibroblasts isolated from breast and prostate cancer tissues expressed the tested surface markers to different degrees. Surprisingly, those markers were expressed also by permanent cell lines of epithelial origin, both benign and cancer-derived (breast-cell lines MCF 10A, HMLE and prostate-cell lines BPH-1, DU 145, and PC-3). The expression of fibroblast activation protein α increased on the surface of previously described models of epithelial cells undergoing epithelial-to-mesenchymal transition in response to treatment with TGF-β1. To prove the co-expression of the fibroblast markers on cells of epithelial origin, we used freshly dissociated human prostate and breast cancer tissues. The results confirmed the co-expression of anti-fibroblast and fibroblast surface protein on CD31/CD45-negative/EpCAM-positive epithelial cells. In summary, our data support the findings that the tested fibroblast markers are not fibroblast specific and may be expressed also by cells of epithelial origin (e.g., cells undergoing EMT). Therefore, the expression of these markers should be interpreted with caution, and the combination of several epitopes for both positive (anti-fibroblast or fibroblast activation protein α) and negative (Ep

  7. Fibroblast growth factor 23

    African Journals Online (AJOL)

    Dr Olaleye

    Systemic phosphate homeostasis is maintained through several hormonal mechanisms which involve fibroblast growth factor 23 (FGF-23), α-klotho, vitamin D and parathyroid hormone. FGF-23 is known to be the major regulator of phosphate balance (Mirams et al., 2004). FGF-23 is a phosphaturic hormone, which is.

  8. Cryopreservation of canine ovarian and testicular fibroblasts.

    Science.gov (United States)

    Yu, Il-Jeoung; Leibo, S P; Songsasen, Nucharin; Dresser, Betsy L; Kim, In-Shik

    2009-01-01

    To derive a practical procedure to store canine somatic cells, fibroblasts isolated from testicular or ovarian tissues were cryopreserved in 1.2 M ethylene glycol or in 1.2 M dimethylsulfoxide prepared in Dulbecco's Modified Eagle Medium as cryoprotectants, and were frozen either in plastic straws or vials. Thawed cells were cultured for 24 hr at 38.5 degree C in a humidified atmosphere of 5 percent CO2 95 percent air, and then their membrane integrity was assayed with a double fluorescent stain, Fertilight. In addition, frozen-thawed fibroblasts were cultured for 4 days, and then their functional survival was measured after staining small colonies with trypan blue. After freezing and thawing, membrane integrity of testicular fibroblasts was 55-70 percent and functional survival ranged from 20-40 percent. With frozen-thawed ovarian cells, the average membrane integrity was 55-75 percent and the average functional survival was 35-40 percent. When frozen in ethylene glycol, functional survival of ovarian fibroblasts was significantly higher than that of testicular cells (P less than 0.05). These methods should prove useful to preserve cells collected from canids in the wild.

  9. Production of N-acyl-L-homoserine lactones by P. aeruginosa isolates from chronic lung infections associated with cystic fibrosis

    DEFF Research Database (Denmark)

    Geisenberger, O; Givskov, M; Riedel, K

    2000-01-01

    The N-acyl-L-homoserine lactones (AHLs) produced by sequential Pseudomonas aeruginosa isolates from chronically infected patients with cystic fibrosis were analyzed by thin-layer chromatography. It is demonstrated that both the amounts and the types of molecules synthesized by isolates from...

  10. Walking along the Fibroblast Growth Factor 10 Route: A Key Pathway to Understand the Control and Regulation of Epithelial and Mesenchymal Cell-Lineage Formation during Lung Development and Repair after Injury.

    Science.gov (United States)

    El Agha, Elie; Bellusci, Saverio

    2014-01-01

    Basic research on embryonic lung development offers unique opportunities to make important discoveries that will impact human health. Developmental biologists interested in the molecular control of branching morphogenesis have intensively studied the developing lung, with its complex and seemingly stereotyped ramified structure. However, it is also an organ that is linked to a vast array of clinical problems in humans such as bronchopulmonary dysplasia in premature babies and emphysema, chronic obstructive pulmonary disease, fibrosis, and cancer in adults. Epithelial stem/progenitor cells reside in niches where they interact with specific extracellular matrices as well as with mesenchymal cells; the latter are still poorly characterized. Interactions of epithelial stem/progenitor cells with their microenvironments are usually instructive, controlling quiescence versus activation, proliferation, differentiation, and migration. During the past 18 years, Fgf10 has emerged not only as a marker for the distal lung mesenchyme during early lung development, but also as a key player in branching morphogenesis and a critical component of the niche for epithelial stem cells. In this paper, we will present the current knowledge regarding the lineage tree in the lung, with special emphasis on cell-lineage decisions in the lung mesenchyme and the role of Fgf10 in this context.

  11. Walking along the Fibroblast Growth Factor 10 Route: A Key Pathway to Understand the Control and Regulation of Epithelial and Mesenchymal Cell-Lineage Formation during Lung Development and Repair after Injury

    Directory of Open Access Journals (Sweden)

    Elie El Agha

    2014-01-01

    Full Text Available Basic research on embryonic lung development offers unique opportunities to make important discoveries that will impact human health. Developmental biologists interested in the molecular control of branching morphogenesis have intensively studied the developing lung, with its complex and seemingly stereotyped ramified structure. However, it is also an organ that is linked to a vast array of clinical problems in humans such as bronchopulmonary dysplasia in premature babies and emphysema, chronic obstructive pulmonary disease, fibrosis, and cancer in adults. Epithelial stem/progenitor cells reside in niches where they interact with specific extracellular matrices as well as with mesenchymal cells; the latter are still poorly characterized. Interactions of epithelial stem/progenitor cells with their microenvironments are usually instructive, controlling quiescence versus activation, proliferation, differentiation, and migration. During the past 18 years, Fgf10 has emerged not only as a marker for the distal lung mesenchyme during early lung development, but also as a key player in branching morphogenesis and a critical component of the niche for epithelial stem cells. In this paper, we will present the current knowledge regarding the lineage tree in the lung, with special emphasis on cell-lineage decisions in the lung mesenchyme and the role of Fgf10 in this context.

  12. Improved throughput traction microscopy reveals pivotal role for matrix stiffness in fibroblast contractility and TGF-β responsiveness

    Science.gov (United States)

    Marinković, Aleksandar; Mih, Justin D.; Park, Jin-Ah; Liu, Fei

    2012-01-01

    Lung fibroblast functions such as matrix remodeling and activation of latent transforming growth factor-β1 (TGF-β1) are associated with expression of the myofibroblast phenotype and are directly linked to fibroblast capacity to generate force and deform the extracellular matrix. However, the study of fibroblast force-generating capacities through methods such as traction force microscopy is hindered by low throughput and time-consuming procedures. In this study, we improved at the detail level methods for higher-throughput traction measurements on polyacrylamide hydrogels using gel-surface-bound fluorescent beads to permit autofocusing and automated displacement mapping, and transduction of fibroblasts with a fluorescent label to streamline cell boundary identification. Together these advances substantially improve the throughput of traction microscopy and allow us to efficiently compute the forces exerted by lung fibroblasts on substrates spanning the stiffness range present in normal and fibrotic lung tissue. Our results reveal that lung fibroblasts dramatically alter the forces they transmit to the extracellular matrix as its stiffness changes, with very low forces generated on matrices as compliant as normal lung tissue. Moreover, exogenous TGF-β1 selectively accentuates tractions on stiff matrices, mimicking fibrotic lung, but not on physiological stiffness matrices, despite equivalent changes in Smad2/3 activation. Taken together, these results demonstrate a pivotal role for matrix mechanical properties in regulating baseline and TGF-β1-stimulated contraction of lung fibroblasts and suggest that stiff fibrotic lung tissue may promote myofibroblast activation through contractility-driven events, whereas normal lung tissue compliance may protect against such feedback amplification of fibroblast activation. PMID:22659883

  13. Fibroblast growth factor 10 haploinsufficiency causes chronic obstructive pulmonary disease.

    Science.gov (United States)

    Klar, Joakim; Blomstrand, Peter; Brunmark, Charlott; Badhai, Jitendra; Håkansson, Hanna Falk; Brange, Charlotte Sollie; Bergendal, Birgitta; Dahl, Niklas

    2011-10-01

    Genetic factors influencing lung function may predispose to chronic obstructive pulmonary disease (COPD). The fibroblast growth factor 10 (FGF10) signalling pathway is critical for lung development and lung epithelial renewal. The hypothesis behind this study was that constitutive FGF10 insufficiency may lead to pulmonary disorder. Therefore investigation of the pulmonary functions of patients heterozygous for loss of function mutations in the FGF10 gene was performed. The spirometric measures of lung function from patients and non-carrier siblings were compared and both groups were related to matched reference data for normal human lung function. The patients show a significant decrease in lung function parameters when compared to control values. The average FEV1/IVC quota (FEV1%) for the patients is 0.65 (80% of predicted) and reversibility test using Terbutalin resulted in a 3.7% increase in FEV1. Patients with FGF10 haploinsufficiency have lung function parameters indicating COPD. A modest response to Terbutalin confirms an irreversible obstructive lung disease. These findings support the idea that genetic variants affecting the FGF10 signalling pathway are important determinants of lung function that may ultimately contribute to COPD. Specifically, the results show that FGF10 haploinsufficiency affects lung function measures providing a model for a dosage sensitive effect of FGF10 in the development of COPD.

  14. Lung-derived growth factors: possible paracrine effectors of fetal lung development

    International Nuclear Information System (INIS)

    Montes, A.M.

    1985-01-01

    A potential role for paracrine secretions in lung organogenesis has been hypothesized (Alescio and Piperno, 1957). These studies present direct support for the paracrine model by demonstrating the presence of locally produced mitogenic/maturational factors in fetal rat lung tissue. Conditioned serum free medium (CSFM) from nineteen-day fetal rat lung cultures was shown to contain several bioactive peptides as detected by 3 H-Thymidine incorporation into chick embryo and rat lung fibroblasts, as well as 14 C-choline incorporation into surfactant in mixed cell cultures. Using ion-exchange chromatography and Sephadex gel filtration, a partially purified mitogen, 11-III, was obtained. The partially purified 11-III stimulates mitosis in chick embryo fibroblasts and post-natal rat lung fibroblasts. Multiplication in fetal rat lung fibroblasts cultures is stimulated only when these are pre-incubated with a competence factor or unprocessed CSFM. This suggests the existence of an endogenously produced competence factor important in the regulation of fetal lung growth. Preparation 11-III does not possess surfactant stimulating activity as assessed by 3 H-choline incorporation into lipids in predominantly type-II cell cultures. These data demonstrate the presence of a maturational/mitogenic factor, influencing type-II mixed cell cultures. In addition, 11-III had been shown to play an autocrine role stimulating the proliferation of fetal lung fibroblasts. Finally, these data suggest the existence of a local produced competence factor

  15. Replication of cultured lung epithelial cells

    International Nuclear Information System (INIS)

    Guzowski, D.; Bienkowski, R.

    1986-01-01

    The authors have investigated the conditions necessary to support replication of lung type 2 epithelial cells in culture. Cells were isolated from mature fetal rabbit lungs (29d gestation) and cultured on feeder layers of mitotically inactivated 3T3 fibroblasts. The epithelial nature of the cells was demonstrated by indirect immunofluorescent staining for keratin and by polyacid dichrome stain. Ultrastructural examination during the first week showed that the cells contained myofilaments, microvilli and lamellar bodies (markers for type 2 cells). The following changes were observed after the first week: increase in cell size; loss of lamellar bodies and appearance of multivesicular bodies; increase in rough endoplasmic reticulum and golgi; increase in tonafilaments and well-defined junctions. General cell morphology was good for up to 10 wk. Cells cultured on plastic surface degenerated after 1 wk. Cell replication was assayed by autoradiography of cultures exposed to ( 3 H)-thymidine and by direct cell counts. The cells did not replicate during the first week; however, between 2-10 wk the cells incorporated the label and went through approximately 6 population doublings. They have demonstrated that lung alveolar epithelial cells can replicate in culture if they are maintained on an appropriate substrate. The coincidence of ability to replicate and loss of markers for differentiation may reflect the dichotomy between growth and differentiation commonly observed in developing systems

  16. Acinetobacter baumannii and A. pittii clinical isolates lack adherence and cytotoxicity to lung epithelial cells in vitro.

    Science.gov (United States)

    Lázaro-Díez, María; Navascués-Lejarza, Teresa; Remuzgo-Martínez, Sara; Navas, Jesús; Icardo, José Manuel; Acosta, Felix; Martínez-Martínez, Luis; Ramos-Vivas, José

    2016-09-01

    The molecular and genetic basis of Acinetobacter baumannii and Acinetobacter pittii virulence remains poorly understood, and there is still lack of knowledge in host cell response to these bacteria. In this study, we have used eleven clinical Acinetobacter strains (A. baumannii n = 5; A. pittii n = 6) to unravel bacterial adherence, invasion and cytotoxicity to human lung epithelial cells. Our results showed that adherence to epithelial cells by Acinetobacter strains is scarce and cellular invasion was not truly detected. In addition, all Acinetobacter strains failed to induce any cytotoxic effect on A549 cells. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  17. One-and-a-half ventricular repair for isolated right ventricle metastatic tumor resection after lobectomy for lung cancer.

    Science.gov (United States)

    Shiose, Akira; Desai, Parag; Criner, Gerard J; Pai, Sheela; Steiner, Robert M; Kaiser, Larry R; Guy, T Sloane; Toyoda, Yoshiya

    2014-01-01

    A 77-year-old woman presented with shortness of breath 1 year after a right upper lobectomy for lung cancer. She showed a possible intracardiac metastasis on positron emission tomography scan. There was no other evidence of recurrence. The large right ventricular mass was associated with the right ventricle free wall, the apex, the papillary muscle, and the chordae to the tricuspid valve. After mass resection of the right ventricle, a one-and-a-half ventricular repair was performed with tricuspid valve replacement and defect closure. The patient was discharged on postoperative day 14 without complications and has been well for the first 3 months after the surgery.

  18. Matrix metalloproteinase inhibition reduces contraction by dupuytren fibroblasts.

    Science.gov (United States)

    Townley, William A; Cambrey, Alison D; Khaw, Peng T; Grobbelaar, Adriaan O

    2008-11-01

    Dupuytren's disease is a common fibroproliferative condition of the hand characterized by fibrotic lesions (nodules and cords), leading to disability through progressive digital contracture. Although the etiology of the disease is poorly understood, recent evidence suggests that abnormal matrix metalloproteinase (MMP) activity may play a role in cell-mediated collagen contraction and tissue scarring. The aim of this study was to investigate the efficacy of ilomastat, a broad-spectrum MMP inhibitor, in an in vitro model of Dupuytren fibroblast-mediated contraction. Nodule-derived and cord-derived fibroblasts were isolated from Dupuytren patients; carpal ligament-derived fibroblasts acted as control. Stress-release fibroblast-populated collagen lattices (FPCLs) were used as a model of contraction. FPCLs were allowed to develop mechanical stress (48 hours) during treatment with ilomastat (0-100 micromol/L), released, and allowed to contract over a 48-hour period. Contraction was estimated by measuring lattice area compared with untreated cells or treatment with a control peptide. MMP-1, MMP-2, and MT1-MMP levels were assessed by zymography, Western blotting, and enzyme-linked immunosorbent assay. Nodule-derived fibroblasts contracted lattices (69% +/- 2) to a greater extent than did cord-derived (55% +/- 3) or carpal ligament-derived (55% +/- 1) fibroblasts. Exposure to ilomastat led to significant inhibition of lattice contraction by all fibroblasts, although a reduction in lattice contraction by nodule-derived fibroblasts was most prominent (84% +/- 8). In addition, treatment with ilomastat led to a concomitant suppression of MMP-1 and MMP-2 activity, whereas MT1-MMP activity was found to be upregulated. Our results demonstrate that inhibition of MMP activity results in a reduction in extracellular matrix contraction by Dupuytren fibroblasts and suggest that MMP activity may be a critical target in preventing recurrent contracture caused by this disease.

  19. Treatment and Prognosis of Isolated Local Relapse after Stereotactic Body Radiotherapy for Clinical Stage I Non-Small-Cell Lung Cancer: Importance of Salvage Surgery.

    Science.gov (United States)

    Hamaji, Masatsugu; Chen, Fengshi; Matsuo, Yukinori; Ueki, Nami; Hiraoka, Masahiro; Date, Hiroshi

    2015-11-01

    Many efforts have been made to detect local relapse (LR) in the follow-up after stereotactic body radiotherapy (SBRT) for non-small-cell lung cancer (NSCLC) although limited data are available on its treatment and prognosis. We aimed to characterize treatment options and clarify long-term outcomes of isolated LR after SBRT for patients with clinical stage I NSCLC. We reviewed our institutional database in search of patients with isolated LR after SBRT for clinical stage I NSCLC at our institution between 1999 and 2013. Patient characteristics were compared with Mann-Whitney U test, χ2 test, or Fisher's exact test as appropriate. Survival outcomes were estimated with Kaplan-Meier method. Potential prognostic factors were investigated using Cox proportional hazard model. Of 308 patients undergoing SBRT for clinical stage I NSCLC, 49 patients were identified to have isolated LR. Twelve patients underwent salvage surgery, none underwent radiotherapy, and eight patients received chemotherapy, whereas 29 patients received best supportive care. No patient characteristic except operability was significantly related with patient selection for LR treatments. Five-year overall survival (OS) rate of the whole cohort was 47.9% from SBRT and 25.7% from LR. Salvage surgery was associated with improved OS after LR (p = 0.014), and 5-year OS for patients undergoing salvage surgery was 79.5% from LR. It was confirmed that our patient selection for salvage surgery for isolated LR was associated with favorable survival outcomes. Operability based on multidisciplinary conferences, rather than measurable patient characteristics, is essential for appropriate patient selection for salvage surgery.

  20. Establishment of human induced pluripotent stem cell lines from normal fibroblast TIG-1.

    Science.gov (United States)

    Kumazaki, Tsutomu; Kurata, Sayaka; Matsuo, Taira; Mitsui, Youji; Takahashi, Tomoko

    2011-06-01

    Normal human cells have a replicative life span and therefore senesce. Usually, normal human cell strains are differentiated cells and reach a terminally differentiated state after a number of cell divisions. At present, definitive differences are not known between replicative senescence and terminal differentiation. TIG-1 is a human fibroblast strain established from fetal lung and has been used extensively in studies of cellular senescence, and numerous data were accumulated at the molecular level. Recently, a method for generating induced pluripotent stem cells (iPSCs) was developed. Using the method, we introduced four reprogramming genes to TIG-1 fibroblasts and succeeded in isolating colonies that had embryonic stem cell (ESC)-like morphologies. They showed alkaline phosphatase activity and expressed ESC markers, as shown by immunostaining of OCT4, SOX2, SSEA4, and TRA-1-81 as well as reverse-transcription polymerase chain reaction (RT-PCR) for OCT4 and NANOG transcripts. Thus, we succeeded in establishing iPSC clones from TIG-1. The iPSC clones could differentiate to cells originated from all three germ-cell layers, as shown by RT-PCR, for messenger RNA (mRNA) expression of α-fetoprotein (endoderm), MSX1 (mesoderm) and microtubule-associated protein 2 (ectoderm), and by immunostaining for α-fetoprotein (endoderm), α-smooth muscle actin (mesoderm), and β-III-tubulin (ectoderm). The iPSCs formed teratoma containing the structures developed from all three germ-cell layers in severe combined immune-deficiency mice. Thus, by comparing the aging process of parental TIG-1 cells and the differentiation process of iPSC-derived fibrocytes to fibroblasts, we can reveal the exact differences in processes between senescence and terminal differentiation.

  1. PHENOMENON OF DEMIKHOV. "TRANSPLANTATION OF VITAL ORGANS IN EXPERIMENT" (1960. HOMOPLASTIC ORGAN TRANSPLANTATION: OF AN ISOLATED HEART, ISOLATED LUNG, KIDNEY, HEAD, HALF OF THE TRUNK, AND THE WHOLE TRUNK

    Directory of Open Access Journals (Sweden)

    S. P. Glyantsev

    2017-01-01

    Full Text Available The  article  (the third one of five presents the analysis of the final part  of the third chapter on homoplastic organ transplantation from V.P.  Demikhov's monograph "Transplantation of  vital  organs  in  experiment"  (MedGIz Publ., Moscow,  1960. The article covers the data presented by V.P. Demikhov on his creating the following animal models:  of the isolated  heart  and lung,  kidneys, head, half of the trunk, and the whole  trunk. The isolated  heart  models where  the heart  was positioned in place of the biological one were not successful. The longest  period  of the dog-recipient survival was  15 hours. The  dogs with transplanted lobes of  the  lung  survived up  to 6 days  (there  are no evidence of  a whole lung  transplant in the book; animal-recipients of kidneys survived up to 19 days.  V.P. Demikhov considered that  the graft deaths might be caused  by the biological  tissue  incompatibility between  the donor  and  recipient, and  he stressed the  need  for  its suppression and  the  importance of  choosing the  donor  and  recipient compatible by their  blood group for  successful homoplastic transplantation. By  transplanting the puppies' heads  onto  the cervical  vessels  of  the dogs, starting the experiments from 1954, V.P. Demikhov attempted to prove the role of the central nervous system in the organ engraftment, which was one of the targets  in those experiments. The integration of two bodies, combining their vascular systems, and mixing the blood had two goals: to create the conditions for overcoming the transplantation immunity and to make an animal model  of organ transplant in combination with retaining the organ neurohumoral links. The model of the cross circulation also allowed the formulation and implementation of the organ bank concept in the experiment.

  2. Global gene expression patterns in the post-pneumonectomy lung of adult mice

    Directory of Open Access Journals (Sweden)

    Ingenito Edward P

    2009-10-01

    Full Text Available Abstract Background Adult mice have a remarkable capacity to regenerate functional alveoli following either lung resection or injury that exceeds the regenerative capacity observed in larger adult mammals. The molecular basis for this unique capability in mice is largely unknown. We examined the transcriptomic responses to single lung pneumonectomy in adult mice in order to elucidate prospective molecular signaling mechanisms used in this species during lung regeneration. Methods Unilateral left pneumonectomy or sham thoracotomy was performed under general anesthesia (n = 8 mice per group for each of the four time points. Total RNA was isolated from the remaining lung tissue at four time points post-surgery (6 hours, 1 day, 3 days, 7 days and analyzed using microarray technology. Results The observed transcriptomic patterns revealed mesenchymal cell signaling, including up-regulation of genes previously associated with activated fibroblasts (Tnfrsf12a, Tnc, Eln, Col3A1, as well as modulation of Igf1-mediated signaling. The data set also revealed early down-regulation of pro-inflammatory cytokine transcripts and up-regulation of genes involved in T cell development/function, but few similarities to transcriptomic patterns observed during embryonic or post-natal lung development. Immunohistochemical analysis suggests that early fibroblast but not myofibroblast proliferation is important during lung regeneration and may explain the preponderance of mesenchymal-associated genes that are over-expressed in this model. This again appears to differ from embryonic alveologenesis. Conclusion These data suggest that modulation of mesenchymal cell transcriptome patterns and proliferation of S100A4 positive mesenchymal cells, as well as modulation of pro-inflammatory transcriptome patterns, are important during post-pneumonectomy lung regeneration in adult mice.

  3. Isolation and Characterization of Cancer Stem Cells of the Non-Small-Cell Lung Cancer (A549) Cell Line.

    Science.gov (United States)

    Halim, Noor Hanis Abu; Zakaria, Norashikin; Satar, Nazilah Abdul; Yahaya, Badrul Hisham

    2016-01-01

    Cancer is a major health problem worldwide. The failure of current treatments to completely eradicate cancer cells often leads to cancer recurrence and dissemination. Studies have suggested that tumor growth and spread are driven by a minority of cancer cells that exhibit characteristics similar to those of normal stem cells, thus these cells are called cancer stem cells (CSCs). CSCs are believed to play an important role in initiating and promoting cancer. CSCs are resistant to currently available cancer therapies, and understanding the mechanisms that control the growth of CSCs might have great implications for cancer therapy. Cancer cells are consist of heterogeneous population of cells, thus methods of identification, isolation, and characterisation of CSCs are fundamental to obtain a pure CSC populations. Therefore, this chapter describes in detail a method for isolating and characterizing a pure population of CSCs from heterogeneous population of cancer cells and CSCs based on specific cell surface markers.

  4. Spatial and Single-Cell Transcriptional Profiling Identifies Functionally Distinct Human Dermal Fibroblast Subpopulations.

    Science.gov (United States)

    Philippeos, Christina; Telerman, Stephanie B; Oulès, Bénédicte; Pisco, Angela O; Shaw, Tanya J; Elgueta, Raul; Lombardi, Giovanna; Driskell, Ryan R; Soldin, Mark; Lynch, Magnus D; Watt, Fiona M

    2018-04-01

    Previous studies have shown that mouse dermis is composed of functionally distinct fibroblast lineages. To explore the extent of fibroblast heterogeneity in human skin, we used a combination of comparative spatial transcriptional profiling of human and mouse dermis and single-cell transcriptional profiling of human dermal fibroblasts. We show that there are at least four distinct fibroblast populations in adult human skin, not all of which are spatially segregated. We define markers permitting their isolation and show that although marker expression is lost in culture, different fibroblast subpopulations retain distinct functionality in terms of Wnt signaling, responsiveness to IFN-γ, and ability to support human epidermal reconstitution when introduced into decellularized dermis. These findings suggest that ex vivo expansion or in vivo ablation of specific fibroblast subpopulations may have therapeutic applications in wound healing and diseases characterized by excessive fibrosis. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  5. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2014-08-01

    Full Text Available Background Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. Methods In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. Results Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1-fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity Conclusions Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  6. Cathepsin K in Lymphangioleiomyomatosis: LAM Cell-Fibroblast Interactions Enhance Protease Activity by Extracellular Acidification.

    Science.gov (United States)

    Dongre, Arundhati; Clements, Debbie; Fisher, Andrew J; Johnson, Simon R

    2017-08-01

    Lymphangioleiomyomatosis (LAM) is a rare disease in which LAM cells and fibroblasts form lung nodules and it is hypothesized that LAM nodule-derived proteases cause cyst formation and tissue damage. On protease gene expression profiling in whole lung tissue, cathepsin K gene expression was 40-fold overexpressed in LAM compared with control lung tissue (P ≤ 0.0001). Immunohistochemistry confirmed cathepsin K protein was expressed in LAM but not control lungs. Cathepsin K gene expression and protein and protease activity were detected in LAM-associated fibroblasts but not the LAM cell line 621-101. In lung nodules, cathepsin K immunoreactivity predominantly co-localized with LAM-associated fibroblasts. In vitro, fibroblast extracellular cathepsin K activity was minimal at pH 7.5 but significantly enhanced at pH 7 and 6. 621-101 cells reduced extracellular pH with acidification dependent on 621-101 mechanistic target of rapamycin activity and net hydrogen ion exporters, particularly sodium bicarbonate co-transporters and carbonic anhydrases, which were also expressed in LAM lung tissue. In LAM cell-fibroblast co-cultures, acidification paralleled cathepsin K activity, and both were reduced by sodium bicarbonate co-transporter (P ≤ 0.0001) and carbonic anhydrase inhibitors (P = 0.0021). Our findings suggest that cathepsin K activity is dependent on LAM cell-fibroblast interactions, and inhibitors of extracellular acidification may be potential therapies for LAM. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  7. The hallmarks of fibroblast ageing.

    Science.gov (United States)

    Tigges, Julia; Krutmann, Jean; Fritsche, Ellen; Haendeler, Judith; Schaal, Heiner; Fischer, Jens W; Kalfalah, Faiza; Reinke, Hans; Reifenberger, Guido; Stühler, Kai; Ventura, Natascia; Gundermann, Sabrina; Boukamp, Petra; Boege, Fritz

    2014-06-01

    Ageing is influenced by the intrinsic disposition delineating what is maximally possible and extrinsic factors determining how that frame is individually exploited. Intrinsic and extrinsic ageing processes act on the dermis, a post-mitotic skin compartment mainly consisting of extracellular matrix and fibroblasts. Dermal fibroblasts are long-lived cells constantly undergoing damage accumulation and (mal-)adaptation, thus constituting a powerful indicator system for human ageing. Here, we use the systematic of ubiquitous hallmarks of ageing (Lopez-Otin et al., 2013, Cell 153) to categorise the available knowledge regarding dermal fibroblast ageing. We discriminate processes inducible in culture from phenomena apparent in skin biopsies or primary cells from old donors, coming to the following conclusions: (i) Fibroblasts aged in culture exhibit most of the established, ubiquitous hallmarks of ageing. (ii) Not all of these hallmarks have been detected or investigated in fibroblasts aged in situ (in the skin). (iii) Dermal fibroblasts aged in vitro and in vivo exhibit additional features currently not considered ubiquitous hallmarks of ageing. (iv) The ageing process of dermal fibroblasts in their physiological tissue environment has only been partially elucidated, although these cells have been a preferred model of cell ageing in vitro for decades. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  8. The increase of microRNA-21 during lung fibrosis and its contribution to epithelial-mesenchymal transition in pulmonary epithelial cells.

    Science.gov (United States)

    Yamada, Mitsuhiro; Kubo, Hiroshi; Ota, Chiharu; Takahashi, Toru; Tando, Yukiko; Suzuki, Takaya; Fujino, Naoya; Makiguchi, Tomonori; Takagi, Kiyoshi; Suzuki, Takashi; Ichinose, Masakazu

    2013-09-24

    The excess and persistent accumulation of fibroblasts due to aberrant tissue repair results in fibrotic diseases such as idiopathic pulmonary fibrosis. Recent reports have revealed significant changes in microRNAs during idiopathic pulmonary fibrosis and evidence in support of a role for microRNAs in myofibroblast differentiation and the epithelial-mesenchymal transition in the context of fibrosis. It has been reported that microRNA-21 is up-regulated in myofibroblasts during fibrosis and promotes transforming growth factor-beta signaling by inhibiting Smad7. However, expression changes in microRNA-21 and the role of microRNA-21 in epithelial-mesenchymal transition during lung fibrosis have not yet been defined. Lungs from saline- or bleomycin-treated C57BL/6 J mice and lung specimens from patients with idiopathic pulmonary fibrosis were analyzed. Enzymatic digestions were performed to isolate single lung cells. Lung epithelial cells were isolated by flow cytometric cell sorting. The expression of microRNA-21 was analyzed using both quantitative PCR and in situ hybridization. To induce epithelial-mesenchymal transition in culture, isolated mouse lung alveolar type II cells were cultured on fibronectin-coated chamber slides in the presence of transforming growth factor-β, thus generating conditions that enhance epithelial-mesenchymal transition. To investigate the role of microRNA-21 in epithelial-mesenchymal transition, we transfected cells with a microRNA-21 inhibitor. Total RNA was isolated from the freshly isolated and cultured cells. MicroRNA-21, as well as mRNAs of genes that are markers of alveolar epithelial or mesenchymal cell differentiation, were quantified using quantitative PCR. The lung epithelial cells isolated from the bleomycin-induced lung fibrosis model system had decreased expression of epithelial marker genes, whereas the expression of mesenchymal marker genes was increased. MicroRNA-21 was significantly upregulated in isolated lung epithelial

  9. The effects of acoustic vibration on fibroblast cell migration.

    Science.gov (United States)

    Mohammed, Taybia; Murphy, Mark F; Lilley, Francis; Burton, David R; Bezombes, Frederic

    2016-12-01

    Cells are known to interact and respond to external mechanical cues and recent work has shown that application of mechanical stimulation, delivered via acoustic vibration, can be used to control complex cell behaviours. Fibroblast cells are known to respond to physical cues generated in the extracellular matrix and it is thought that such cues are important regulators of the wound healing process. Many conditions are associated with poor wound healing, so there is need for treatments/interventions, which can help accelerate the wound healing process. The primary aim of this research was to investigate the effects of mechanical stimulation upon the migratory and morphological properties of two different fibroblast cells namely; human lung fibroblast cells (LL24) and subcutaneous areolar/adipose mouse fibroblast cells (L929). Using a speaker-based system, the effects of mechanical stimulation (0-1600Hz for 5min) on the mean cell migration distance (μm) and actin organisation was investigated. The results show that 100Hz acoustic vibration enhanced cell migration for both cell lines whereas acoustic vibration above 100Hz was found to decrease cell migration in a frequency dependent manner. Mechanical stimulation was also found to promote changes to the morphology of both cell lines, particularly the formation of lamellipodia and filopodia. Overall lamellipodia was the most prominent actin structure displayed by the lung cell (LL24), whereas filopodia was the most prominent actin feature displayed by the fibroblast derived from subcutaneous areolar/adipose tissue. Mechanical stimulation at all the frequencies used here was found not to affect cell viability. These results suggest that low-frequency acoustic vibration may be used as a tool to manipulate the mechanosensitivity of cells to promote cell migration. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. An unusual case of isolated, serial metastases of gallbladder carcinoma involving the chest wall, axilla, breast and lung parenchyma

    Directory of Open Access Journals (Sweden)

    Matthew J. Iott

    2013-02-01

    Full Text Available In the English literature, only 9 cases of adenocarcinoma of the gallbladder with cutaneous metastasis have been reported so far. One case of multiple cutaneous metastases along with deposits in the breast tissue has been reported. We present a case of incidental metastatic gallbladder carcinoma with no intra-abdominal disease presenting as a series of four isolated cutaneous right chest wall, axillary nodal, breast and pulmonary metastases following resection and adjuvant chemoradiation for her primary tumor. In spite of the metastatic disease coupled with the aggressive nature of the cancer, this patient reported that her energy level had returned to baseline with a good appetite and a stable weight indicating a good performance status and now is alive at 25 months since diagnosis. Her serially-presented, oligometastatic diseases were well-controlled by concurrent chemoradiation and stereotactic radiation therapy. We report this case study because of its rarity and for the purpose of complementing current literature with an additional example of cutaneous metastasis from adenocarcinoma of the gallbladder.

  11. The Role of PPARs in Lung Fibrosis

    Directory of Open Access Journals (Sweden)

    Heather F. Lakatos

    2007-01-01

    wound healing. PPARβ/δ agonists inhibit lung fibroblast proliferation and enhance the antifibrotic properties of PPARγ agonists. PPARγ ligands oppose the profibrotic effect of TGF-β, which induces differentiation of fibroblasts to myofibroblasts, a critical effector cell in fibrosis. PPARγ ligands, including the thiazolidinedione class of antidiabetic drugs, effectively inhibit lung fibrosis in vitro and in animal models. The clinical availability of potent and selective PPARα and PPARγ agonists should facilitate rapid development of successful treatment strategies based on current and ongoing research.

  12. Three-dimensional characterization of fibroblast foci in idiopathic pulmonary fibrosis

    Science.gov (United States)

    Jones, Mark G.; Fabre, Aurélie; Schneider, Philipp; Cinetto, Francesco; Sgalla, Giacomo; Jogai, Sanjay; Alzetani, Aiman; Marshall, Ben G.; O’Reilly, Katherine M.A.; Warner, Jane A.; Lackie, Peter M.; Davies, Donna E.; Hansell, David M.; Nicholson, Andrew G.; Sinclair, Ian; Brown, Kevin K.; Richeldi, Luca

    2016-01-01

    In idiopathic pulmonary fibrosis (IPF), the fibroblast focus is a key histological feature representing active fibroproliferation. On standard 2D pathologic examination, fibroblast foci are considered small, distinct lesions, although they have been proposed to form a highly interconnected reticulum as the leading edge of a “wave” of fibrosis. Here, we characterized fibroblast focus morphology and interrelationships in 3D using an integrated micro-CT and histological methodology. In 3D, fibroblast foci were morphologically complex structures, with large variations in shape and volume (range, 1.3 × 104 to 9.9 × 107 μm3). Within each tissue sample numerous multiform foci were present, ranging from a minimum of 0.9 per mm3 of lung tissue to a maximum of 11.1 per mm3 of lung tissue. Each focus was an independent structure, and no interconnections were observed. Together, our data indicate that in 3D fibroblast foci form a constellation of heterogeneous structures with large variations in shape and volume, suggesting previously unrecognized plasticity. No evidence of interconnectivity was identified, consistent with the concept that foci represent discrete sites of lung injury and repair. PMID:27275013

  13. Lung cancer

    International Nuclear Information System (INIS)

    Aisner, J.

    1985-01-01

    This book contains 13 chapters. Some of the chapter titles are: The Pathology of Lung Cancer; Radiotherapy for Non-Small-Cell Cancer of the Lung; Chemotherapy for Non-Small-Cell Lung Cancer; Immunotherapy in the Management of Lung Cancer; Preoperative Staging and Surgery for Non-Small-Cell Lung Cancer; and Prognostic Factors in Lung Cancer

  14. Hyperoxia decreases glycolytic capacity, glycolytic reserve and oxidative phosphorylation in MLE-12 cells and inhibits complex I and II function, but not complex IV in isolated mouse lung mitochondria.

    Directory of Open Access Journals (Sweden)

    Kumuda C Das

    Full Text Available High levels of oxygen (hyperoxia are frequently used in critical care units and in conditions of respiratory insufficiencies in adults, as well as in infants. However, hyperoxia has been implicated in a number of pulmonary disorders including bronchopulmonary dysplasia (BPD and adult respiratory distress syndrome (ARDS. Hyperoxia increases the generation of reactive oxygen species (ROS in the mitochondria that could impair the function of the mitochondrial electron transport chain. We analyzed lung mitochondrial function in hyperoxia using the XF24 analyzer (extracellular flux and optimized the assay for lung epithelial cells and mitochondria isolated from lungs of mice. Our data show that hyperoxia decreases basal oxygen consumption rate (OCR, spare respiratory capacity, maximal respiration and ATP turnover in MLE-12 cells. There was significant decrease in glycolytic capacity and glycolytic reserve in MLE-12 cells exposed to hyperoxia. Using mitochondria isolated from lungs of mice exposed to hyperoxia or normoxia we have shown that hyperoxia decreased the basal, state 3 and state3 μ (respiration in an uncoupled state respirations. Further, using substrate or inhibitor of a specific complex we show that the OCR via complex I and II, but not complex IV was decreased, demonstrating that complexes I and II are specific targets of hyperoxia. Further, the activities of complex I (NADH dehydrogenase, NADH-DH and complex II (succinate dehydrogenase, SDH were decreased in hyperoxia, but the activity of complex IV (cytochrome oxidase, COX remains unchanged. Taken together, our study show that hyperoxia impairs glycolytic and mitochondrial energy metabolism in in tact cells, as well as in lungs of mice by selectively inactivating components of electron transport system.

  15. Fibroblast growth factor receptors in breast cancer.

    Science.gov (United States)

    Wang, Shuwei; Ding, Zhongyang

    2017-05-01

    Fibroblast growth factor receptors are growth factor receptor tyrosine kinases, exerting their roles in embryogenesis, tissue homeostasis, and development of breast cancer. Recent genetic studies have identified some subtypes of fibroblast growth factor receptors as strong genetic loci associated with breast cancer. In this article, we review the recent epidemiological findings and experiment results of fibroblast growth factor receptors in breast cancer. First, we summarized the structure and physiological function of fibroblast growth factor receptors in humans. Then, we discussed the common genetic variations in fibroblast growth factor receptors that affect breast cancer risk. In addition, we also introduced the potential roles of each fibroblast growth factor receptors isoform in breast cancer. Finally, we explored the potential therapeutics targeting fibroblast growth factor receptors for breast cancer. Based on the biological mechanisms of fibroblast growth factor receptors leading to the pathogenesis in breast cancer, targeting fibroblast growth factor receptors may provide new opportunities for breast cancer therapeutic strategies.

  16. Usefulness of fibroblast culture for testing of cattle tissues polluted with heavy metals

    International Nuclear Information System (INIS)

    Weglarz, L.; Drozdz, M.Wa.; Wardas, M.; Kula, B.; Pawlaczyk-Szpilowa, M.

    1990-01-01

    Cattle tissues (liver, kidney, brain, and lung) that had been polluted with heavy metals were tested for their ability to alter fibroblast culture growth, cellular protein and DNA content, and fibroblast DNA synthesis. At 72 hr of incubation a significant increase in cellular DNA and [14C]thymidine incorporation was noted in the primary cultures as well as in the subcultures compared to controls. Fibroblast cultures also displayed growth inhibition and reduction in protein content. The measurement of basic biochemical parameters of the fibroblast culture may represent a sensitive means of assessing rapidly the activity of heavy metals deposited in the tissues of cattle as a result of their grazing on polluted soil

  17. The mechanical memory of lung myofibroblasts.

    Science.gov (United States)

    Balestrini, Jenna L; Chaudhry, Sidharth; Sarrazy, Vincent; Koehler, Anne; Hinz, Boris

    2012-04-01

    Fibroblasts differentiate into the highly synthetic and contractile myofibroblast phenotype when exposed to substrates with an elastic modulus corresponding to pathologically stiff fibrotic tissue. Cellular responses to changes in substrate stiffness are typically analyzed after hours or days, which does not enable the monitoring of myofibroblast persistence, a hallmark of fibrosis. To determine long-lasting effects on the fibrotic behavior of lung fibroblasts, we followed a novel approach of explanting and repeatedly passaging fibroblasts on silicone substrates with stiffness representing various states of lung health. Fibrotic activity was determined by assaying for myofibroblast proliferation, cell contractility, expression of α-smooth muscle actin, extracellular matrix and active TGFβ1. As predicted, myofibroblast activity was low on healthy soft substrates and increased with increasing substrate stiffness. However, explanting and mechanically priming lung fibroblasts for 3 weeks on pathologically stiff substrates resulted in sustained myofibroblast activity even after the cells were returned to healthy soft cultures for 2 weeks. Such primed cells retained higher fibrotic activity than cells that had been exclusively cultured on soft substrates, and were not statistically different from cells continuously passaged on stiff surfaces. Inversely, priming lung fibroblasts for 3 weeks on soft substrates partially protected from myofibroblast activation after the shift to stiff substrates. Hence, mechano-sensed information relating to physical conditions of the local cellular environment could permanently induce fibrotic behavior of lung fibroblasts. This priming effect has important implications for the progression and persistence of aggressive fibrotic diseases such as idiopathic pulmonary fibrosis. This journal is © The Royal Society of Chemistry 2012

  18. Inhibition of fibroblast growth factor receptor with AZD4547 mitigates juvenile nasopharyngeal angiofibroma.

    Science.gov (United States)

    Le, Tran; New, Jacob; Jones, Joel W; Usman, Shireen; Yalamanchali, Sreeya; Tawfik, Ossama; Hoover, Larry; Bruegger, Dan E; Thomas, Sufi Mary

    2017-10-01

    Juvenile nasopharyngeal angiofibroma (JNA) is a benign tumor that presents in adolescent males. Although surgical excision is the mainstay of treatment, recurrences complicate treatment. There is a need to develop less invasive approaches for management. JNA tumors are composed of fibroblasts and vascular endothelial cells. We identified fibroblast growth factor receptor (FGFR) and vascular endothelial growth factor (VEGF) expression in JNA-derived fibroblasts. FGFR influences fibroblast proliferation and VEGF is necessary for angiogenesis. We hypothesized that targeting FGFR would mitigate JNA fibroblast proliferation, invasion, and migration, and that targeting the VEGF receptor would attenuate endothelial tubule formation. After informed consent, fibroblasts from JNA explants of 3 patients were isolated. Fibroblasts were treated with FGFR inhibitor AZD4547, 0 to 25 μg/mL for 72 hours and proliferation was quantified using CyQuant assay. Migration and invasion of JNA were assessed using 24-hour transwell assays with subsequent fixation and quantification. Mitigation of FGFR and downstream signaling was evaluated by immunoblotting. Tubule formation was assessed in human umbilical vein endothelial cells (HUVECs) treated with vehicle control (dimethylsulfoxide [DMSO]) or semaxanib (SU5416) as well as in serum-free media (SFM) or JNA conditioned media (CM). Tubule length was compared between treatment groups. Compared to control, AZD4547 inhibited JNA fibroblast proliferation, migration, and invasion through inhibition of FGFR and downstream signaling, specifically phosphorylation of - p44/42 mitogen activated protein kinase (p44/42 MAPK). JNA fibroblast CM significantly increased HUVEC tubule formation (p = 0.0039). AZD4547 effectively mitigates FGFR signaling and decreases JNA fibroblast proliferation, migration, and invasion. SU5416 attenuated JNA fibroblast-induced tubule formation. AZD4547 may have therapeutic potential in the treatment of JNA. © 2017 ARS

  19. MicroRNA-124 controls the proliferative, migratory, and inflammatory phenotype of pulmonary vascular fibroblasts.

    Science.gov (United States)

    Wang, Daren; Zhang, Hui; Li, Min; Frid, Maria G; Flockton, Amanda R; McKeon, B Alexandre; Yeager, Michael E; Fini, Mehdi A; Morrell, Nicholas W; Pullamsetti, Soni S; Velegala, Sivareddy; Seeger, Werner; McKinsey, Timothy A; Sucharov, Carmen C; Stenmark, Kurt R

    2014-01-03

    Pulmonary hypertensive remodeling is characterized by excessive proliferation, migration, and proinflammatory activation of adventitial fibroblasts. In culture, fibroblasts maintain a similar activated phenotype. The mechanisms responsible for generation/maintenance of this phenotype remain unknown. We hypothesized that aberrant expression of microRNA-124 (miR-124) regulates this activated fibroblast phenotype and sought to determine the signaling pathways through which miR-124 exerts effects. We detected significant decreases in miR-124 expression in fibroblasts isolated from calves and humans with severe pulmonary hypertension. Overexpression of miR-124 by mimic transfection significantly attenuated proliferation, migration, and monocyte chemotactic protein-1 expression of hypertensive fibroblasts, whereas anti-miR-124 treatment of control fibroblasts resulted in their increased proliferation, migration, and monocyte chemotactic protein-1 expression. Furthermore, the alternative splicing factor, polypyrimidine tract-binding protein 1, was shown to be a direct target of miR-124 and to be upregulated both in vivo and in vitro in bovine and human pulmonary hypertensive fibroblasts. The effects of miR-124 on fibroblast proliferation were mediated via direct binding to the 3' untranslated region of polypyrimidine tract-binding protein 1 and subsequent regulation of Notch1/phosphatase and tensin homolog/FOXO3/p21Cip1 and p27Kip1 signaling. We showed that miR-124 directly regulates monocyte chemotactic protein-1 expression in pulmonary hypertension/idiopathic pulmonary arterial hypertension fibroblasts. Furthermore, we demonstrated that miR-124 expression is suppressed by histone deacetylases and that treatment of hypertensive fibroblasts with histone deacetylase inhibitors increased miR-124 expression and decreased proliferation and monocyte chemotactic protein-1 production. Stable decreases in miR-124 expression contribute to an epigenetically reprogrammed, highly

  20. Endoglin negatively regulates transforming growth factor beta1-induced profibrotic responses in intestinal fibroblasts.

    LENUS (Irish Health Repository)

    Burke, J P

    2012-02-01

    BACKGROUND: Fibroblasts isolated from strictures in Crohn\\'s disease (CD) exhibit reduced responsiveness to stimulation with transforming growth factor (TGF) beta1. TGF-beta1, acting through the smad pathway, is critical to fibroblast-mediated intestinal fibrosis. The membrane glycoprotein, endoglin, is a negative regulator of TGF-beta1. METHODS: Intestinal fibroblasts were cultured from seromuscular biopsies of patients undergoing intestinal resection for CD strictures or from control patients. Endoglin expression was assessed using confocal microscopy, flow cytometry and western blot. The effect of small interfering (si) RNA-mediated knockdown and plasmid-mediated overexpression of endoglin on fibroblast responsiveness to TGF-beta1 was assessed by examining smad phosphorylation, smad binding element (SBE) promoter activity, connective tissue growth factor (CTGF) expression and ability to contract collagen. RESULTS: Crohn\\'s stricture fibroblasts expressed increased constitutive cell-surface and whole-cell endoglin relative to control cells. Endoglin co-localized with filamentous actin. Fibroblasts treated with siRNA directed against endoglin exhibited enhanced TGF-beta1-mediated smad-3 phosphorylation, and collagen contraction. Cells transfected with an endoglin plasmid did not respond to TGF-beta1 by exhibiting SBE promoter activity or producing CTGF. CONCLUSION: Fibroblasts from strictures in CD express increased constitutive endoglin. Endoglin is a negative regulator of TGF-beta1 signalling in the intestinal fibroblast, modulating smad-3 phosphorylation, SBE promoter activity, CTGF production and collagen contraction.

  1. Cancer-associated fibroblasts from human NSCLC survive ablative doses of radiation but their invasive capacity is reduced

    Directory of Open Access Journals (Sweden)

    Hellevik Turid

    2012-04-01

    Full Text Available Abstract Background Cancer-Associated Fibroblasts (CAFs are significant components of solid malignancies and play central roles in cancer sustainability, invasion and metastasis. In this study we have investigated the invasive capacity and matrix remodelling properties of human lung CAFs after exposure to ablative doses of ionizing radiation (AIR, equivalent to single fractions delivered by stereotactic ablative radiotherapy (SART for medically inoperable stage-I/II non-small-cell lung cancers. Methods CAFs were isolated from lung tumour specimens from 16 donors. Initially, intrinsic radiosensitivity was evaluated by checking viability and extent of DNA-damage response (DDR at different radiation doses. The migrative and invasive capacities of CAFs were thereafter determined after a sub-lethal single radiation dose of 18 Gy. To ascertain the mechanisms behind the altered invasive capacity of cells, expression of matrix metalloproteinases (MMPs and their endogenous inhibitors (TIMPs were measured in the conditioned media several days post-irradiation, along with expression of cell surface integrins and dynamics of focal contacts by vinculin-staining. Results Exposing CAFs to 1 × 18 Gy resulted in a potent induction of multiple nuclear DDR foci (> 9/cell with little resolution after 120 h, induced premature cellular senescence and inhibition of the proliferative, migrative and invasive capacity. AIR promoted MMP-3 and inhibited MMP-1 appearance to some extent, but did not affect expression of other major MMPs. Furthermore, surface expression of integrins α2, β1 and α5 was consistently enhanced, and a dramatic augmentation and redistribution of focal contacts was observed. Conclusions Our data indicate that ablative doses of radiation exert advantageous inhibitory effects on the proliferative, migratory and invasive capacity of lung CAFs. The reduced motility of irradiated CAFs might be a consequence of stabilized focal contacts via integrins.

  2. Constituents from the roots of Taraxacum platycarpum and their effect on proliferation of human skin fibroblasts.

    Science.gov (United States)

    Warashina, Tsutomu; Umehara, Kaoru; Miyase, Toshio

    2012-01-01

    A MeOH extract from the roots of Taraxacum platycarpum has shown significant effects on the proliferation of normal human skin fibroblasts. Chemical analysis of the extract resulted in the isolation of 26 compounds, including eight new triterpenes, one new sesquiterpene glycoside, and seventeen known compounds. The structure of each new compound was established using NMR spectroscopy. Some triterpenes had a significant effect on the proliferation of normal human skin fibroblasts.

  3. Microprobe analysis of human fibroblasts

    International Nuclear Information System (INIS)

    Allan, G.L.; Zhu, J.; Legge, G.J.F.

    1985-01-01

    The Melbourne Proton Microprobe has been used to study the copper content in human skin fibroblast cells derived from patients with the genetic disease Menkes Syndrome. Both normal and diseased cells have been studied to investigate any elemental differences occurring between the two cell types. This paper details the preparatory techniques necessary for individual cell analysis and presents the elemental information with a new three dimensional contour mapping technique. These maps are used to highlight elemental differences between normal and mutant fibroblasts. The work also confirms the expected copper excess found in the Menkes cell and indicates that the microprobe can be used for rapid identification of a Menkes carrier

  4. Merkel Cell Polyomavirus Infection of Animal Dermal Fibroblasts.

    Science.gov (United States)

    Liu, Wei; Krump, Nathan A; MacDonald, Margo; You, Jianxin

    2018-02-15

    Merkel cell polyomavirus (MCPyV) is the first polyomavirus to be associated with human cancer. Mechanistic studies attempting to fully elucidate MCPyV's oncogenic mechanisms have been hampered by the lack of animal models for MCPyV infection. In this study, we examined the ability of MCPyV-GFP pseudovirus (containing a green fluorescent protein [GFP] reporter construct), MCPyV recombinant virions, and several MCPyV chimeric viruses to infect dermal fibroblasts isolated from various model animals, including mouse ( Mus musculus ), rabbit ( Oryctolagus cuniculus ), rat ( Rattus norvegicus ), chimpanzee ( Pan troglodytes ), rhesus macaque ( Macaca mulatta ), patas monkey ( Erythrocebus patas ), common woolly monkey ( Lagothrix lagotricha ), red-chested mustached tamarin ( Saguinus labiatus ), and tree shrew ( Tupaia belangeri ). We found that MCPyV-GFP pseudovirus was able to enter the dermal fibroblasts of all species tested. Chimpanzee dermal fibroblasts were the only type that supported vigorous MCPyV gene expression and viral replication, and they did so to a level beyond that of human dermal fibroblasts. We further demonstrated that both human and chimpanzee dermal fibroblasts produce infectious MCPyV virions that can successfully infect new cells. In addition, rat dermal fibroblasts supported robust MCPyV large T antigen expression after infection with an MCPyV chimeric virus in which the entire enhancer region of the MCPyV early promoter has been replaced with the simian virus 40 (SV40) analog. Our results suggest that viral transcription and/or replication events represent the major hurdle for MCPyV cross-species transmission. The capacity of rat dermal fibroblasts to support MCPyV early gene expression suggests that the rat is a candidate model organism for studying viral oncogene function during Merkel cell carcinoma (MCC) oncogenic progression. IMPORTANCE MCPyV plays an important role in the development of a highly aggressive form of skin cancer, Merkel

  5. Case report 511: Fibroblastic rheumatism

    International Nuclear Information System (INIS)

    Hernandez, R.J.; Martel, W.; Headington, J.T.; Kaufman, R.A.; Cincinnati Univ., OH

    1989-01-01

    We report a ten-year-old child with the newly described entity of fibroblastic rheumatism. This child developed rapid, progressive, symmetrical polyarthritis, similar to the radiographic appearance of juvenile rheumatoid arthritis, except for the rapidity of progression. The polyarthritis was preceded by the development of skin nodules with characteristic histological changes. (orig./GDG)

  6. Lung Emergencies

    Science.gov (United States)

    ... The Marfan Foundation Marfan & Related Disorders What is Marfan Syndrome? What are Related Disorders? What are the Signs? ... Emergencies Lung Emergencies Surgeries Lung Emergencies People with Marfan syndrome can be at increased risk of sudden lung ...

  7. Nutrition for Lung Cancer

    Science.gov (United States)

    ... Become An Advocate Volunteer Ways To Give Lung Cancer www.lung.org > Lung Health and Diseases > Lung Disease Lookup > ... Cancer Learn About Lung Cancer What Is Lung Cancer Lung Cancer Basics Causes & Risk Factors Lung Cancer Staging ...

  8. Diallylthiosulfinate (Allicin), a Volatile Antimicrobial from Garlic (Allium sativum), Kills Human Lung Pathogenic Bacteria, Including MDR Strains, as a Vapor.

    Science.gov (United States)

    Reiter, Jana; Levina, Natalja; van der Linden, Mark; Gruhlke, Martin; Martin, Christian; Slusarenko, Alan J

    2017-10-12

    Garlic ( Allium sativum ) has potent antimicrobial activity due to allicin (diallylthiosulfinate) synthesized by enzyme catalysis in damaged garlic tissues. Allicin gives crushed garlic its characteristic odor and its volatility makes it potentially useful for combating lung infections. Allicin was synthesized (>98% pure) by oxidation of diallyl disulfide by H₂O₂ using formic acid as a catalyst and the growth inhibitory effect of allicin vapor and allicin in solution to clinical isolates of lung pathogenic bacteria from the genera Pseudomonas , Streptococcus , and Staphylococcus , including multi-drug resistant (MDR) strains, was demonstrated. Minimal inhibitory (MIC) and minimal bactericidal concentrations (MBC) were determined and compared to clinical antibiotics using standard European Committee on Antimicrobial Susceptibility Testing (EUCAST) procedures. The cytotoxicity of allicin to human lung and colon epithelial and murine fibroblast cells was tested in vitro and shown to be ameliorated by glutathione (GSH). Similarly, the sensitivity of rat precision-cut lung slices (PCLS) to allicin was decreased by raising the [GSH] to the approximate blood plasma level of 1 mM. Because allicin inhibited bacterial growth as a vapor, it could be used to combat bacterial lung infections via direct inhalation. Since there are no volatile antibiotics available to treat pulmonary infections, allicin, particularly at sublethal doses in combination with oral antibiotics, could make a valuable addition to currently available treatments.

  9. Diallylthiosulfinate (Allicin, a Volatile Antimicrobial from Garlic (Allium sativum, Kills Human Lung Pathogenic Bacteria, Including MDR Strains, as a Vapor

    Directory of Open Access Journals (Sweden)

    Jana Reiter

    2017-10-01

    Full Text Available Garlic (Allium sativum has potent antimicrobial activity due to allicin (diallylthiosulfinate synthesized by enzyme catalysis in damaged garlic tissues. Allicin gives crushed garlic its characteristic odor and its volatility makes it potentially useful for combating lung infections. Allicin was synthesized (>98% pure by oxidation of diallyl disulfide by H2O2 using formic acid as a catalyst and the growth inhibitory effect of allicin vapor and allicin in solution to clinical isolates of lung pathogenic bacteria from the genera Pseudomonas, Streptococcus, and Staphylococcus, including multi-drug resistant (MDR strains, was demonstrated. Minimal inhibitory (MIC and minimal bactericidal concentrations (MBC were determined and compared to clinical antibiotics using standard European Committee on Antimicrobial Susceptibility Testing (EUCAST procedures. The cytotoxicity of allicin to human lung and colon epithelial and murine fibroblast cells was tested in vitro and shown to be ameliorated by glutathione (GSH. Similarly, the sensitivity of rat precision-cut lung slices (PCLS to allicin was decreased by raising the [GSH] to the approximate blood plasma level of 1 mM. Because allicin inhibited bacterial growth as a vapor, it could be used to combat bacterial lung infections via direct inhalation. Since there are no volatile antibiotics available to treat pulmonary infections, allicin, particularly at sublethal doses in combination with oral antibiotics, could make a valuable addition to currently available treatments.

  10. Effect of eosinophils activated with Alternaria on the production of extracellular matrix from nasal fibroblasts.

    Science.gov (United States)

    Shin, Seung-Heon; Ye, Mi-Kyung; Choi, Sung-Yong; Kim, Yee-Hyuk

    2016-06-01

    Eosinophils and fibroblasts are known to play major roles in the pathogenesis of nasal polyps. Fungi are commonly found in nasal secretion and are associated with airway inflammation. To investigate whether activated eosinophils by airborne fungi can influence the production of extracellular matrix (ECM) from nasal fibroblasts. Inferior turbinate and nasal polyp fibroblasts were stimulated with Alternaria or Aspergillus, respectively, for 24 hours and ECM messenger RNA (mRNA) and protein expressions were measured. Eosinophils isolated from healthy volunteers were stimulated with Alternaria or Aspergillus for 4 hours then superoxide, eosinophil peroxidase, and transforming growth factor β1 were measured. Then activated eosinophils were cocultured with nasal fibroblasts for 24 hours, and ECM mRNA expressions were measured. Alternaria strongly enhanced ECM mRNA expression and protein production from nasal fibroblasts. Alternaria also induced the production of superoxide, eosinophil peroxidase, and transforming growth factor β1 from eosinophils, and activated eosinophils enhanced ECM mRNA expression when they were cocultured without the Transwell insert system. Eosinophils activated with Alternaria enhanced ECM mRNA expression from nasal polyp fibroblasts. Alternaria plays an important role in tissue fibrosis in the pathogenesis of nasal polyps by directly or indirectly influencing the production of ECM from nasal fibroblasts. Copyright © 2016 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  11. Semaphorin 4A enhances lung fibrosis through activation of Akt via ...

    Indian Academy of Sciences (India)

    2015-11-28

    Nov 28, 2015 ... In the present study, we show that treatment of Sema4A on normal lung fibroblasts ... of patients with the fibrotic disease Systemic Sclerosis (SSc) showed ... modulated at multiple levels: transcription, translation, and post-.

  12. Design and characteristics of cytotoxic fibroblast growth factor 1 conjugate for fibroblast growth factor receptor-targeted cancer therapy

    Directory of Open Access Journals (Sweden)

    Szlachcic A

    2016-08-01

    Full Text Available Anna Szlachcic, Malgorzata Zakrzewska, Michal Lobocki, Piotr Jakimowicz, Jacek Otlewski Department of Protein Engineering, Faculty of Biotechnology, University of Wroclaw, Wroclaw, Poland Abstract: Fibroblast growth factor receptors (FGFRs are attractive candidate cancer therapy targets as they are overexpressed in multiple types of tumors, such as breast, prostate, bladder, and lung cancer. In this study, a natural ligand of FGFR, an engineered variant of fibroblast growth factor 1 (FGF1V, was conjugated to a potent cytotoxic drug, monomethyl auristatin E (MMAE, and used as a targeting agent for cancer cells overexpressing FGFRs, similar to antibodies in antibody–drug conjugates. The FGF1V–valine–citrulline–MMAE conjugate showed a favorable stability profile, bound FGFRs on the cell surface specifically, and efficiently released the drug (MMAE upon cleavage by the lysosomal protease cathepsin B. Importantly, the conjugate showed a prominent cytotoxic effect toward cell lines expressing FGFR. FGF1V–vcMMAE was highly cytotoxic at concentrations even an order of magnitude lower than those found for free MMAE. This effect was FGFR-specific as cells lacking FGFR did not show any increased mortality. Keywords: fibroblast growth factor 1, FGF receptor, targeted cancer therapy, cytotoxic conjugates, FGFR-dependent cancer, MMAE, auristatin

  13. Optimization of the agar-gel method for isolation of migrating Ascaris suum larvae from the liver and lungs of pigs

    DEFF Research Database (Denmark)

    Saeed, I.; Roepstorff, A.; Rasmussen, T.

    2001-01-01

    Experiments on use of an agar-gel method for recovery of migrating Ascaris suum larvae from the liver and lungs of pigs were conducted to obtain fast standardized methods. Subsamples of blended tissues of pig liver and lungs were mixed with agar to a final concentration of 1% agar and the larvae...... clean suspension which reduced the sample counting time. Blending the liver for 60 sec in a commercial blender showed significantly higher larvae recovery than blending for 30 sec. Addition of gentamycin to reduce bacterial growth during incubation, glucose to increase larval motility during migration...

  14. Ex vivo expansion of alveolar macrophages with Mycobacterium tuberculosis from the resected lungs of patients with pulmonary tuberculosis

    Science.gov (United States)

    Petrunina, Ekaterina; Umpeleva, Tatiana; Karskanova, Svetlana; Bayborodin, Sergey; Vakhrusheva, Diana; Kravchenko, Marionella; Skornyakov, Sergey

    2018-01-01

    Tuberculosis (TB), with the Mycobacterium tuberculosis (Mtb) as the causative agent, remains to be a serious world health problem. Traditional methods used for the study of Mtb in the lungs of TB patients do not provide information about the number and functional status of Mtb, especially if Mtb are located in alveolar macrophages. We have developed a technique to produce ex vivo cultures of cells from different parts of lung tissues surgically removed from patients with pulmonary TB and compared data on the number of cells with Mtb inferred by the proposed technique to the results of bacteriological and histological analyses used for examination of the resected lungs. The ex vivo cultures of cells obtained from the resected lungs of all patients were largely composed of CD14-positive alveolar macrophages, foamy or not, with or without Mtb. Lymphocytes, fibroblasts, neutrophils, and multinucleate Langhans giant cells were also observed. We found alveolar macrophages with Mtb in the ex vivo cultures of cells from the resected lungs of even those TB patients, whose sputum smears and lung tissues did not contain acid-fast Mtb or reveal growing Mtb colonies on dense medium. The detection of alveolar macrophages with Mtb in ex vivo culture as soon as 16–18 h after isolation of cells from the resected lungs of all TB patients suggests that the technique proposed for assessing the level of infection in alveolar macrophages of TB patients has higher sensitivity than do prolonged bacteriological or pathomorphological methods. The proposed technique allowed us to rapidly (in two days after surgery) determine the level of infection with Mtb in the cells of the resected lungs of TB patients and, by the presence or absence of Mtb colonies, including those with cording morphology, the functional status of the TB agent at the time of surgery. PMID:29401466

  15. Bio-artificial pleura using an autologous dermal fibroblast sheet

    Science.gov (United States)

    Kanzaki, Masato; Takagi, Ryo; Washio, Kaoru; Kokubo, Mami; Yamato, Masayuki

    2017-10-01

    Air leaks (ALs) are observed after pulmonary resections, and without proper treatment, can produce severe complications. AL prevention is a critical objective for managing patients after pulmonary resection. This study applied autologous dermal fibroblast sheets (DFS) to close ALs. For sealing ALs in a 44-year-old male human patient with multiple bullae, a 5 × 15-mm section of skin was surgically excised. From this skin specimen, primary dermal fibroblasts were isolated and cultured for 4 weeks to produce DFSs that were harvested after a 10-day culture. ALs were completely sealed using surgical placement of these autologous DFSs. DFS were found to be a durable long-term AL sealant, exhibiting requisite flexibility, elasticity, durability, biocompatibility, and usability, resulting reliable AL closure. DFS should prove to be an extremely useful tissue-engineered pleura substitute.

  16. Coxsackievirus B3 induces the formation of autophagosomes in cardiac fibroblasts both in vitro and in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Zhai, Xia, E-mail: zhai_xia_cool@126.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Qin, Ying, E-mail: qinyinggaofeng@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Chen, Yang, E-mail: cy_hmu@126.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Lin, Lexun, E-mail: linlexun@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Wang, Tianying, E-mail: wangty0929@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Zhong, Xiaoyan, E-mail: littlerock712@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Wu, Xiaoyu, E-mail: xiaoyu_wu2006@163.com [Department of Cardiology, The First Hospital of Harbin Medical University, 23 Youzheng Street, Harbin 150001 (China); Chen, Sijia, E-mail: chensj0802@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Li, Jing, E-mail: jing070822@163.com [Center of Electron Microscopy, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Wang, Yan, E-mail: wangyan@hrbmu.edu.cn [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Zhang, Fengmin, E-mail: fengminzhang@ems.hrbmu.edu.cn [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Zhao, Wenran, E-mail: zhaowenran2002@aliyun.com [Department of Cell Biology, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); and others

    2016-12-10

    Coxsackievirus group B (CVB) is one of the common pathogens that cause myocarditis and cardiomyopathy. Evidence has shown that CVB replication in cardiomyocytes is responsible for the damage and loss of cardiac muscle and the dysfunction of the heart. However, it remains largely undefined how CVB would directly impact cardiac fibroblasts, the most abundant cells in human heart. In this study, cardiac fibroblasts were isolated from Balb/c mice and infected with CVB type 3 (CVB3). Increased double-membraned, autophagosome-like vesicles in the CVB3-infected cardiac fibroblasts were observed with electron microscope. Punctate distribution of LC3 and increased level of LC3-II were also detected in the infected cardiac fibroblasts. Furthermore, we observed that the expression of pro-inflammatory cytokines, IL-6 and TNF-α, was increased in the CVB3-infected cardiac fibroblasts, while suppressed autophagy by 3-MA and Atg7-siRNA inhibited cytokine expression. Consistent with the in vitro findings, increased formation of autophagosomes was observed in the cardiac fibroblasts of Balb/c mice infected with CVB3. In conclusion, our data demonstrated that cardiac fibroblasts respond to CVB3 infection with the formation of autophagosomes and the release of the pro-inflammatory cytokines. These results suggest that the autophagic response of cardiac fibroblasts may play a role in the pathogenesis of myocarditis caused by CVB3 infection. - Highlights: • CVB3 replication induced autophagosome assembly in primary cardiac fibroblasts. • Both IL-6 and TNF-α in cardiac fibroblasts infected by CVB3 were increased. • IL-6 and TNF-α were reduced in cardiac fibroblasts when autophagy was inhibited. • Autophagosome assembly in cardiac fibroblasts of CVB-infected mice was increased.

  17. Coxsackievirus B3 induces the formation of autophagosomes in cardiac fibroblasts both in vitro and in vivo

    International Nuclear Information System (INIS)

    Zhai, Xia; Qin, Ying; Chen, Yang; Lin, Lexun; Wang, Tianying; Zhong, Xiaoyan; Wu, Xiaoyu; Chen, Sijia; Li, Jing; Wang, Yan; Zhang, Fengmin; Zhao, Wenran

    2016-01-01

    Coxsackievirus group B (CVB) is one of the common pathogens that cause myocarditis and cardiomyopathy. Evidence has shown that CVB replication in cardiomyocytes is responsible for the damage and loss of cardiac muscle and the dysfunction of the heart. However, it remains largely undefined how CVB would directly impact cardiac fibroblasts, the most abundant cells in human heart. In this study, cardiac fibroblasts were isolated from Balb/c mice and infected with CVB type 3 (CVB3). Increased double-membraned, autophagosome-like vesicles in the CVB3-infected cardiac fibroblasts were observed with electron microscope. Punctate distribution of LC3 and increased level of LC3-II were also detected in the infected cardiac fibroblasts. Furthermore, we observed that the expression of pro-inflammatory cytokines, IL-6 and TNF-α, was increased in the CVB3-infected cardiac fibroblasts, while suppressed autophagy by 3-MA and Atg7-siRNA inhibited cytokine expression. Consistent with the in vitro findings, increased formation of autophagosomes was observed in the cardiac fibroblasts of Balb/c mice infected with CVB3. In conclusion, our data demonstrated that cardiac fibroblasts respond to CVB3 infection with the formation of autophagosomes and the release of the pro-inflammatory cytokines. These results suggest that the autophagic response of cardiac fibroblasts may play a role in the pathogenesis of myocarditis caused by CVB3 infection. - Highlights: • CVB3 replication induced autophagosome assembly in primary cardiac fibroblasts. • Both IL-6 and TNF-α in cardiac fibroblasts infected by CVB3 were increased. • IL-6 and TNF-α were reduced in cardiac fibroblasts when autophagy was inhibited. • Autophagosome assembly in cardiac fibroblasts of CVB-infected mice was increased.

  18. Redox Modulation by Amaranth Oil in Human Lung Fibroblasts

    NARCIS (Netherlands)

    Semen, K.O.; den Hartog, G.J.M.; Kaminsky, D.V.; Sirota, T.V.; Maij, N.G.A.A.; Yelisyeyeva, O.P.; Bast, A.

    2013-01-01

    Amaranth oil has several health benefits. It has lipid lowering, anti-diabetic, immune modulatory and cytoprotective properties, activates the function of mitochondria and improves heart rate variability. It has been suggested that the effect of amaranth oil on redox status is involved in this

  19. 25-Hydroxycholesterol promotes fibroblast-mediated tissue remodeling through NF-κB dependent pathway

    International Nuclear Information System (INIS)

    Ichikawa, Tomohiro; Sugiura, Hisatoshi; Koarai, Akira; Kikuchi, Takashi; Hiramatsu, Masataka; Kawabata, Hiroki; Akamatsu, Keiichiro; Hirano, Tsunahiko; Nakanishi, Masanori; Matsunaga, Kazuto; Minakata, Yoshiaki; Ichinose, Masakazu

    2013-01-01

    Abnormal structural alterations termed remodeling, including fibrosis and alveolar wall destruction, are important features of the pathophysiology of chronic airway diseases such as chronic obstructive pulmonary disease (COPD) and asthma. 25-hydroxycholesterol (25-HC) is enzymatically produced by cholesterol 25-hydorxylase (CH25H) in macrophages and is reported to be involved in the formation of arteriosclerosis. We previously demonstrated that the expression of CH25H and production of 25HC were increased in the lungs of COPD. However, the role of 25-HC in lung tissue remodeling is unknown. In this study, we investigated the effect of 25-HC on fibroblast-mediated tissue remodeling using human fetal lung fibroblasts (HFL-1) in vitro. 25-HC significantly augmented α-smooth muscle actin (SMA) (P 1 production (P 1 release. These results suggest that 25-HC could contribute to fibroblast-mediated lung tissue remodeling by promoting myofibroblast differentiation and the excessive release of extracellular matrix protein and MMPs via an NF-κB-TGF-β dependent pathway

  20. Human skeletal muscle fibroblasts stimulate in vitro myogenesis and in vivo muscle regeneration.

    Science.gov (United States)

    Mackey, Abigail L; Magnan, Mélanie; Chazaud, Bénédicte; Kjaer, Michael

    2017-08-01

    Accumulation of skeletal muscle extracellular matrix is an unfavourable characteristic of many muscle diseases, muscle injury and sarcopenia. The extent of cross-talk between fibroblasts, as the source of matrix protein, and satellite cells in humans is unknown. We studied this in human muscle biopsies and cell-culture studies. We observed a strong stimulation of myogenesis by human fibroblasts in cell culture. In biopsies collected 30 days after a muscle injury protocol, fibroblast number increased to four times control levels, where fibroblasts were found to be preferentially located immediately surrounding regenerating muscle fibres. These novel findings indicate an important role for fibroblasts in supporting the regeneration of muscle fibres, potentially through direct stimulation of satellite cell differentiation and fusion, and contribute to understanding of cell-cell cross-talk during physiological and pathological muscle remodelling. Accumulation of skeletal muscle extracellular matrix is an unfavourable characteristic of many muscle diseases, muscle injury and sarcopenia. In addition to the indispensable role satellite cells play in muscle regeneration, there is emerging evidence in rodents for a regulatory influence on fibroblast activity. However, the influence of fibroblasts on satellite cells and muscle regeneration in humans is unknown. The purpose of this study was to investigate this in vitro and during in vivo regeneration in humans. Following a muscle injury protocol in young healthy men (n = 7), the number of fibroblasts (TCF7L2+), satellite cells (Pax7+), differentiating myogenic cells (myogenin+) and regenerating fibres (neonatal/embryonic myosin+) was determined from biopsy cross-sections. Fibroblasts and myogenic precursor cells (MPCs) were also isolated from human skeletal muscle (n = 4) and co-cultured using different cell ratios, with the two cell populations either in direct contact with each other or separated by a permeable

  1. Casticin, an active compound isolated from Vitex Fructus, ameliorates the cigarette smoke-induced acute lung inflammatory response in a murine model.

    Science.gov (United States)

    Lee, Hyeonhoon; Jung, Kyung-Hwa; Lee, Hangyul; Park, Soojin; Choi, Woosung; Bae, Hyunsu

    2015-10-01

    The aim of this study was to determine of the effect of casticin, as an anti-inflammatory agent, on an acute lung inflammation in vivo model established through exposure to cigarette smoke (CS). Casticin is a phytochemical from Vitex species such as Vitex rotundifolia and Vitex agnus-castus that was recently shown to exert an anti-inflammatory effect in vivo. To demonstrate the effects of casticin, C57BL/6 mice were whole-body exposed to mainstream CS or fresh air for two weeks and treated with 1, 2, and 10mg/kg casticin via an i.p. injection. Immune cell infiltrations and cytokine productions were assessed from bronchoalveolar lavage Fluid (BALF), and lung histological analysis was performed. Treatment with casticin was observed to significantly inhibit the numbers of total cells, neutrophils, macrophages, and lymphocytes and reduce the levels of proinflammatory cytokines and chemokines in the BALF. In addition, casticin significantly decreased the infiltration of peribronchial and perivascular inflammatory cells and the epithelium thickness. The results of this study indicate that casticin has significant effects on the lung inflammation induced by CS in a mouse model. According to these outcomes, casticin may have therapeutic potential in inflammatory lung diseases, such as chronic obstructive pulmonary disease (COPD). Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Differentiation of human umbilical cord mesenchymal stem cells into dermal fibroblasts in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Han, Yanfu [Department of Burn and Plastic Surgery, Burns Institute, First Hospital Affiliated to General Hospital of PLA, Beijing (China); Chai, Jiake, E-mail: cjk304@126.com [Department of Burn and Plastic Surgery, Burns Institute, First Hospital Affiliated to General Hospital of PLA, Beijing (China); Sun, Tianjun; Li, Dongjie; Tao, Ran [Department of Burn and Plastic Surgery, Burns Institute, First Hospital Affiliated to General Hospital of PLA, Beijing (China)

    2011-10-07

    Highlights: {yields} Mesenchymal stem cells (MSCs) are potential seed cells for tissue-engineered skin. {yields} Tissue-derived umbilical cord MSCs (UCMSCs) can readily be isolated in vitro. {yields} We induce UCMSCs to differentiate into dermal fibroblasts via conditioned medium. {yields} Collagen type I and collagen type III mRNA level was higher in differentiated cells. {yields} UCMSCs-derived fibroblast-like cells strongly express fibroblast-specific protein. -- Abstract: Tissue-derived umbilical cord mesenchymal stem cells (UCMSCs) can be readily obtained, avoid ethical or moral constraints, and show excellent pluripotency and proliferation potential. UCMSCs are considered to be a promising source of stem cells in regenerative medicine. In this study, we collected newborn umbilical cord tissue under sterile conditions and isolated UCMSCs through a tissue attachment method. UCMSC cell surface markers were examined using flow cytometry. On the third passage, UCMSCs were induced to differentiate into dermal fibroblasts in conditioned induction media. The induction results were detected using immunofluorescence with a fibroblast-specific monoclonal antibody and real time PCR for type I and type III collagen. UCMSCs exhibited a fibroblast-like morphology and reached 90% confluency 14 to 18 days after primary culture. Cultured UCMSCs showed strong positive staining for CD73, CD29, CD44, CD105, and HLA-I, but not CD34, CD45, CD31, or HLA-DR. After differentiation, immunostaining for collagen type I, type III, fibroblast-specific protein, vimentin, and desmin were all strongly positive in induced cells, and staining was weak or negative in non-induced cells; total transcript production of collagen type I and collagen type III mRNA was higher in induced cells than in non-induced cells. These results demonstrate that UCMSCs can be induced to differentiate into fibroblasts with conditioned induction media and, in turn, could be used as seed cells for tissue

  3. Differentiation of human umbilical cord mesenchymal stem cells into dermal fibroblasts in vitro

    International Nuclear Information System (INIS)

    Han, Yanfu; Chai, Jiake; Sun, Tianjun; Li, Dongjie; Tao, Ran

    2011-01-01

    Highlights: → Mesenchymal stem cells (MSCs) are potential seed cells for tissue-engineered skin. → Tissue-derived umbilical cord MSCs (UCMSCs) can readily be isolated in vitro. → We induce UCMSCs to differentiate into dermal fibroblasts via conditioned medium. → Collagen type I and collagen type III mRNA level was higher in differentiated cells. → UCMSCs-derived fibroblast-like cells strongly express fibroblast-specific protein. -- Abstract: Tissue-derived umbilical cord mesenchymal stem cells (UCMSCs) can be readily obtained, avoid ethical or moral constraints, and show excellent pluripotency and proliferation potential. UCMSCs are considered to be a promising source of stem cells in regenerative medicine. In this study, we collected newborn umbilical cord tissue under sterile conditions and isolated UCMSCs through a tissue attachment method. UCMSC cell surface markers were examined using flow cytometry. On the third passage, UCMSCs were induced to differentiate into dermal fibroblasts in conditioned induction media. The induction results were detected using immunofluorescence with a fibroblast-specific monoclonal antibody and real time PCR for type I and type III collagen. UCMSCs exhibited a fibroblast-like morphology and reached 90% confluency 14 to 18 days after primary culture. Cultured UCMSCs showed strong positive staining for CD73, CD29, CD44, CD105, and HLA-I, but not CD34, CD45, CD31, or HLA-DR. After differentiation, immunostaining for collagen type I, type III, fibroblast-specific protein, vimentin, and desmin were all strongly positive in induced cells, and staining was weak or negative in non-induced cells; total transcript production of collagen type I and collagen type III mRNA was higher in induced cells than in non-induced cells. These results demonstrate that UCMSCs can be induced to differentiate into fibroblasts with conditioned induction media and, in turn, could be used as seed cells for tissue-engineered dermis.

  4. Hypoxia and loss of PHD2 inactivate stromal fibroblasts to decrease tumour stiffness and metastasis

    DEFF Research Database (Denmark)

    Madsen, Chris D; Pedersen, Jesper Thorhauge; Venning, Freja A

    2015-01-01

    , which can be prevented by simultaneous depletion of HIF-1α. Treatment with the PHD inhibitor DMOG in an orthotopic breast cancer model significantly decreases spontaneous metastases to the lungs and liver, associated with decreased tumour stiffness and fibroblast activation. PHD2 depletion in CAFs co......-injected with tumour cells similarly prevents CAF-induced metastasis to lungs and liver. Our data argue that reversion of CAFs towards a less active state is possible and could have important clinical implications....

  5. Analysis of plasma membrane Ca(2+)-ATPase expression in control and SV40-transformed human fibroblasts.

    Science.gov (United States)

    Reisner, P D; Brandt, P C; Vanaman, T C

    1997-01-01

    It has been long known that neoplastic transformation is accompanied by a lowered requirement for extracellular Ca2+ for growth. The studies presented here demonstrate that human fibroblastic cell lines produce the two commonly found 'housekeeping' isoforms of the plasma membrane Ca(2+)-ATPase (PMCA), PMCA1b and 4b, and at the expression of both is demonstrably lower in cell lines neoplastically transformed by SV40 than in the corresponding parental cell lines. Western blot analyses of lysates from control (GM00037) and SV40-transformed (GM00637) skin fibroblasts revealed a 138 kDa PMCA whose level was significantly lower in the SV40-transformed cells relative to either total cellular protein or alpha-tubulin. Similar analyses of plasma membrane preparations from control WI-38) and SV40-transformed (WI-38VA13) lung fibroblasts revealed 3-4-fold lower levels of PMCA in the SV40-transformed cells. Competitive ELISAs performed on detergent solubilized plasma membrane preparations indicated at least 3-4-fold lower levels of PMCA in the SV40-transformed cell lines compared to controls. Reverse transcriptase coupled-PCR analyses showed that PMCA1b and PMCA4b were the only isoforms expressed in all four cell lines. The PMCA4b mRNA level detected by Northern analysis also was substantially lower in SV40 transformed skin fibroblasts than in non-transformed fibroblasts. Quantitative RT-PCR analyses showed levels of PMCA1b and 4b mRNAs to be 5 and 10-fold lower, respectively, in GM00637 than in GM00037 when the levels of PCR products were normalized to glyceraldehyde-3-phosphate dehydrogenase (G3PDH) mRNA. These results demonstrate that the expression of these distinct PMCA genes is substantially lower in SV40 transformed human skin and lung fibroblasts and may be coordinately regulated in these cells.

  6. Antifibrotic effects of curcumin are associated with overexpression of cathepsins K and L in bleomycin treated mice and human fibroblasts

    Directory of Open Access Journals (Sweden)

    Zhang Dongwei

    2011-11-01

    Full Text Available Abstract Background Lung fibrosis is characterized by fibroblast proliferation and the deposition of collagens. Curcumin, a polyphenol antioxidant from the spice tumeric, has been shown to effectively counteract fibroblast proliferation and reducing inflammation and fibrotic progression in animal models of bleomycin-induced lung injury. However, there is little mechanistic insight in the biological activity of curcumin. Here, we study the effects of curcumin on the expression and activity of cathepsins which have been implicated in the development of fibrotic lung diseases. Methods We investigated the effects of curcumin administration to bleomycin stimulated C57BL/6 mice and human fetal lung fibroblasts (HFL-1 on the expression of cathepsins K and L which have been implicated in matrix degradation, TGF-β1 modulation, and apoptosis. Lung tissues were evaluated for their contents of cathepsins K and L, collagen, and TGF-β1. HFL-1 cells were used to investigate the effects of curcumin and cathepsin inhibition on cell proliferation, migration, apoptosis, and the expression of cathepsins K and L and TGF-β1. Results Collagen deposition in lungs was decreased by 17-28% after curcumin treatment which was accompanied by increased expression levels of cathepsins L (25%-39% and K (41%-76% and a 30% decrease in TGF-β1 expression. Moreover, Tunel staining of lung tissue revealed a 33-41% increase in apoptotic cells after curcumin treatment. These in vivo data correlated well with data obtained from the human fibroblast line, HFL-1. Here, cathepsin K and L expression increased 190% and 240%, respectively, in the presence of curcumin and the expression of TGF-β1 decreased by 34%. Furthermore, curcumin significantly decreased cell proliferation and migration and increased the expression of surrogate markers of apoptosis. In contrast, these curcumin effects were partly reversed by a potent cathepsin inhibitor. Conclusion This study demonstrates that

  7. Fibroblast spheroids as a model to study sustained fibroblast quiescence and their crosstalk with tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Salmenperä, Pertteli, E-mail: pertteli.salmenpera@helsinki.fi [Department of Virology, Medicum, Faculty of Medicine, University of Helsinki, P.O. Box 21, FIN-00014 (Finland); Karhemo, Piia-Riitta [Research Programs Unit, Translational Cancer Biology, and Institute of Biomedicine, University of Helsinki, P.O. Box 63, FIN-00014 (Finland); Räsänen, Kati [Department of Virology, Medicum, Faculty of Medicine, University of Helsinki, P.O. Box 21, FIN-00014 (Finland); Laakkonen, Pirjo [Research Programs Unit, Translational Cancer Biology, and Institute of Biomedicine, University of Helsinki, P.O. Box 63, FIN-00014 (Finland); Vaheri, Antti [Department of Virology, Medicum, Faculty of Medicine, University of Helsinki, P.O. Box 21, FIN-00014 (Finland)

    2016-07-01

    Stromal fibroblasts have an important role in regulating tumor progression. Normal and quiescent fibroblasts have been shown to restrict and control cancer cell growth, while cancer-associated, i. e. activated fibroblasts have been shown to enhance proliferation and metastasis of cancer cells. In this study we describe generation of quiescent fibroblasts in multicellular spheroids and their effects on squamous cell carcinoma (SCC) growth in soft-agarose and xenograft models. Quiescent phenotype of fibroblasts was determined by global down-regulation of expression of genes related to cell cycle and increased expression of p27. Interestingly, microarray analysis showed that fibroblast quiescence was associated with similar secretory phenotype as seen in senescence and they expressed senescence-associated-β-galactosidase. Quiescent fibroblasts spheroids also restricted the growth of RT3 SCC cells both in soft-agarose and xenograft models unlike proliferating fibroblasts. Restricted tumor growth was associated with marginally increased tumor cell senescence and cellular differentiation, showed with senescence-associated-β-galactosidase and cytokeratin 7 staining. Our results show that the fibroblasts spheroids can be used as a model to study cellular quiescence and their effects on cancer cell progression. - Highlights: • Fibroblasts acquire a sustained quiescence when grown as multicellular spheroids. • This quiescence is associated with drastic change in gene expression. • Fibroblasts spheroids secrete various inflammation-linked cytokines and chemokines. • Fibroblasts spheroids reduced growth of RT3 SCC cells in xenograft model.

  8. A dual-color luciferase assay system reveals circadian resetting of cultured fibroblasts by co-cultured adrenal glands.

    Directory of Open Access Journals (Sweden)

    Takako Noguchi

    Full Text Available In mammals, circadian rhythms of various organs and tissues are synchronized by pacemaker neurons in the suprachiasmatic nucleus (SCN of the hypothalamus. Glucocorticoids released from the adrenal glands can synchronize circadian rhythms in other tissues. Many hormones show circadian rhythms in their plasma concentrations; however, whether organs outside the SCN can serve as master synchronizers to entrain circadian rhythms in target tissues is not well understood. To further delineate the function of the adrenal glands and the interactions of circadian rhythms in putative master synchronizing organs and their target tissues, here we report a simple co-culture system using a dual-color luciferase assay to monitor circadian rhythms separately in various explanted tissues and fibroblasts. In this system, circadian rhythms of organs and target cells were simultaneously tracked by the green-emitting beetle luciferase from Pyrearinus termitilluminans (ELuc and the red-emitting beetle luciferase from Phrixothrix hirtus (SLR, respectively. We obtained tissues from the adrenal glands, thyroid glands, and lungs of transgenic mice that expressed ELuc under control of the promoter from a canonical clock gene, mBmal1. The tissues were co-cultured with Rat-1 fibroblasts as representative target cells expressing SLR under control of the mBmal1 promoter. Amplitudes of the circadian rhythms of Rat-1 fibroblasts were potentiated when the fibroblasts were co-cultured with adrenal gland tissue, but not when co-cultured with thyroid gland or lung tissue. The phases of Rat-1 fibroblasts were reset by application of adrenal gland tissue, whereas the phases of adrenal gland tissue were not influenced by Rat-1 fibroblasts. Furthermore, the effect of the adrenal gland tissue on the fibroblasts was blocked by application of a glucocorticoid receptor (GR antagonist. These results demonstrate that glucocorticoids are strong circadian synchronizers for fibroblasts and that

  9. Cellular Mechanics of Primary Human Cervical Fibroblasts: Influence of Progesterone and a Pro-inflammatory Cytokine.

    Science.gov (United States)

    Shukla, Vasudha; Barnhouse, Victoria; Ackerman, William E; Summerfield, Taryn L; Powell, Heather M; Leight, Jennifer L; Kniss, Douglas A; Ghadiali, Samir N

    2018-01-01

    The leading cause of neonatal mortality, pre-term birth, is often caused by pre-mature ripening/opening of the uterine cervix. Although cervical fibroblasts play an important role in modulating the cervix's extracellular matrix (ECM) and mechanical properties, it is not known how hormones, i.e., progesterone, and pro-inflammatory insults alter fibroblast mechanics, fibroblast-ECM interactions and the resulting changes in tissue mechanics. Here we investigate how progesterone and a pro-inflammatory cytokine, IL-1β, alter the biomechanical properties of human cervical fibroblasts and the fibroblast-ECM interactions that govern tissue-scale mechanics. Primary human fibroblasts were isolated from non-pregnant cervix and treated with estrogen/progesterone, IL-1β or both. The resulting changes in ECM gene expression, matrix remodeling, traction force generation, cell-ECM adhesion and tissue contractility were monitored. Results indicate that IL-1β induces a significant reduction in traction force and ECM adhesion independent of pre-treatment with progesterone. These cell level effects altered tissue-scale mechanics where IL-1β inhibited the contraction of a collagen gel over 6 days. Interestingly, progesterone treatment alone did not modulate traction forces or gel contraction but did result in a dramatic increase in cell-ECM adhesion. Therefore, the protective effect of progesterone may be due to altered adhesion dynamics as opposed to altered ECM remodeling.

  10. Pentadecapeptide BPC 157 Enhances the Growth Hormone Receptor Expression in Tendon Fibroblasts

    Directory of Open Access Journals (Sweden)

    Chung-Hsun Chang

    2014-11-01

    Full Text Available BPC 157, a pentadecapeptide derived from human gastric juice, has been demonstrated to promote the healing of different tissues, including skin, muscle, bone, ligament and tendon in many animal studies. However, the underlying mechanism has not been fully clarified. The present study aimed to explore the effect of BPC 157 on tendon fibroblasts isolated from Achilles tendon of male Sprague-Dawley rat. From the result of cDNA microarray analysis, growth hormone receptor was revealed as one of the most abundantly up-regulated genes in tendon fibroblasts by BPC 157. BPC 157 dose- and time-dependently increased the expression of growth hormone receptor in tendon fibroblasts at both the mRNA and protein levels as measured by RT/real-time PCR and Western blot, respectively. The addition of growth hormone to BPC 157-treated tendon fibroblasts dose- and time-dependently increased the cell proliferation as determined by MTT assay and PCNA expression by RT/real-time PCR. Janus kinase 2, the downstream signal pathway of growth hormone receptor, was activated time-dependently by stimulating the BPC 157-treated tendon fibroblasts with growth hormone. In conclusion, the BPC 157-induced increase of growth hormone receptor in tendon fibroblasts may potentiate the proliferation-promoting effect of growth hormone and contribute to the healing of tendon.

  11. Fibrosis of Two: Epithelial Cell-Fibroblast Interactions in Pulmonary Fibrosis

    Science.gov (United States)

    Sakai, Norihiko; Tager, Andrew M.

    2013-01-01

    Idiopathic pulmonary fibrosis (IPF) is characterized by the progressive and ultimately fatal accumulation of fibroblasts and extracellular matrix in the lung that distorts its architecture and compromises its function. IPF is now thought to result from wound-healing processes that, although initiated to protect the host from injurious environmental stimuli, lead to pathological fibrosis due to these processes becoming aberrant or over-exuberant. Although the environmental stimuli that trigger IPF remain to be identified, recent evidence suggests that they initially injure the alveolar epithelium. Repetitive cycles of epithelial injury and resultant alveolar epithelial cell death provoke the migration, proliferation, activation and myofibroblast differentiation of fibroblasts, causing the accumulation of these cells and the extracellular matrix that they synthesize. In turn, these activated fibroblasts induce further alveolar epithelial cell injury and death, thereby creating a vicious cycle of pro-fibrotic epithelial cell-fibroblast interactions. Though other cell types certainly make important contributions, we focus here on the “pas de deux” (steps of two), or perhaps more appropriate to IPF pathogenesis, the “folie à deux” (madness of two) of epithelial cells and fibroblasts that drives the progression of pulmonary fibrosis. We describe the signaling molecules that mediate the interactions of these cell types in their “fibrosis of two”, including transforming growth factor-β, connective tissue growth factor, sonic hedgehog, prostaglandin E2, angiotensin II and reactive oxygen species. PMID:23499992

  12. p53/PUMA expression in human pulmonary fibroblasts mediates cell activation and migration in silicosis.

    Science.gov (United States)

    Wang, Wei; Liu, Haijun; Dai, Xiaoniu; Fang, Shencun; Wang, Xingang; Zhang, Yingming; Yao, Honghong; Zhang, Xilong; Chao, Jie

    2015-11-18

    Phagocytosis of SiO2 into the lung causes an inflammatory cascade that results in fibroblast proliferation and migration, followed by fibrosis. Clinical evidence has indicated that the activation of alveolar macrophages by SiO2 produces rapid and sustained inflammation characterized by the generation of monocyte chemotactic protein 1, which, in turn, induces fibrosis. However, the details of events downstream of monocyte chemotactic protein 1 activity in pulmonary fibroblasts remain unclear. Here, to elucidate the role of p53 in fibrosis induced by silica, both the upstream molecular mechanisms and the functional effects on cell proliferation and migration were investigated. Experiments using primary cultured adult human pulmonary fibroblasts led to the following results: 1) SiO2 treatment resulted in a rapid and sustained increase in p53 and PUMA protein levels; 2) the MAPK and PI3K pathways were involved in the SiO2-induced alteration of p53 and PUMA expression; and 3) RNA interference targeting p53 and PUMA prevented the SiO2-induced increases in fibroblast activation and migration. Our study elucidated a link between SiO2-induced p53/PUMA expression in fibroblasts and cell migration, thereby providing novel insight into the potential use of p53/PUMA in the development of novel therapeutic strategies for silicosis treatment.

  13. Cell Surface Glycoprotein of Reactive Stromal Fibroblasts as a Potential Antibody Target in Human Epithelial Cancers

    Science.gov (United States)

    Garin-Chesa, Pilar; Old, Lloyd J.; Rettig, Wolfgang J.

    1990-09-01

    The F19 antigen is a cell surface glycoprotein (M_r, 95,000) of human sarcomas and proliferating, cultured fibroblasts that is absent from resting fibroblasts in normal adult tissues. Normal and malignant epithelial cells are also F19^-. The present immunohistochemical study describes induction of F19 in the reactive mesenchyme of epithelial tumors. F19^+ fibroblasts were found in primary and metastatic carcinomas, including colorectal (18 of 18 cases studied), breast (14/14), ovarian (21/21), bladder (9/10), and lung carcinomas (13/13). In contrast, the stroma of benign colorectal adenomas, fibrocystic disease and fibroadenomas of breast, benign prostate hyperplasia, in situ bladder carcinomas, and benign ovarian tumors showed no or only moderate numbers of F19^+ fibroblasts. Analysis of dermal incision wounds revealed that F19 is strongly induced during scar formation. Comparison of F19 with the extracellular matrix protein tenascin, a putative marker of tumor mesenchyme, showed a cellular staining pattern for F19 vs. the extracellular matrix pattern for tenascin and widespread expression of tenascin in F19^- normal tissues and benign tumors. Our results suggest that the F19^+ phenotype correlates with specialized fibroblast functions in wound healing and malignant tumor growth. Because of its abundance in tumor mesenchyme, F19 may serve as a target for antibodies labeled with radioisotopes or toxic agents, or inflammatogenic antibodies, in carcinoma patients.

  14. Effect of botulinum toxin type A on transforming growth factor beta1 in fibroblasts derived from hypertrophic scar: a preliminary report.

    Science.gov (United States)

    Xiao, Zhibo; Zhang, Fengmin; Lin, Weibin; Zhang, Miaobo; Liu, Ying

    2010-08-01

    Hypertrophic scar is a common dermal disease. Numerous treatments are currently available but they do not always yield excellent therapeutic results. Hence, alternatives are needed. Recent basic and clinical research has shown that botulinum toxin type A (BTXA) has antihypertrophic scar properties but the molecular mechanism for this action is unknown. The aim of this study was to explore the effect of BTXA on transforming growth factor beta1 (TGF-beta1) in fibroblasts derived from hypertrophic scar and further elucidate its actual mechanism. Fibroblasts were isolated from tissue specimens of hypertrophic scar. Fibroblasts were treated with BTXA and the difference in proliferation between treated and nontreated cells was analyzed through the MTT method from the first to the fifth day after treatment. Proteins of TGF-beta1 were checked using ELISA in fibroblasts with BTXA and without BTXA from the first to the fifth day. The growth of the fibroblast treated with BTXA was obviously slower than that of the fibroblast without BTXA treatment (p < 0.01), which showed that BTXA effectively inhibited the growth of fibroblasts. Proteins of TGF-beta1 between fibroblasts with BTXA and fibroblasts without BTXA are statistically significant (p < 0.01). These results suggest that BTXA effectively inhibited the growth of fibroblasts derived from hypertrophic scar and in turn caused a decrease in TGF-beta1 protein, indicating that BTXA-based therapies for hypertrophic scar are promising and worth investigating further.

  15. Lung scintigraphy

    International Nuclear Information System (INIS)

    Dalenz, Roberto.

    1994-01-01

    A review of lung scintigraphy, perfusion scintigraphy with SPECT, lung ventilation SPECT, blood pool SPECT. The procedure of lung perfusion studies, radiopharmaceutical, administration and clinical applications, imaging processing .Results encountered and evaluation criteria after Biello and Pioped. Recommendations and general considerations have been studied about relation of this radiopharmaceutical with other pathologies

  16. Granulocyte macrophage colony stimulating factor (GM-CSF biological actions on human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    S Montagnani

    2009-12-01

    Full Text Available Fibroblasts are involved in all pathologies characterized by increased ExtraCellularMatrix synthesis, from wound healing to fibrosis. Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF is a cytokine isolated as an hemopoietic growth factor but recently indicated as a differentiative agent on endothelial cells. In this work we demonstrated the expression of the receptor for GM-CSF (GMCSFR on human normal skin fibroblasts from healthy subjects (NFPC and on a human normal fibroblast cell line (NHDF and we try to investigate the biological effects of this cytokine. Human normal fibroblasts were cultured with different doses of GM-CSF to study the effects of this factor on GMCSFR expression, on cell proliferation and adhesion structures. In addition we studied the production of some Extra-Cellular Matrix (ECM components such as Fibronectin, Tenascin and Collagen I. The growth rate of fibroblasts from healthy donors (NFPC is not augmented by GM-CSF stimulation in spite of increased expression of the GM-CSFR. On the contrary, the proliferation of normal human dermal fibroblasts (NHDF cell line seems more influenced by high concentration of GM-CSF in the culture medium. The adhesion structures and the ECM components appear variously influenced by GM-CSF treatment as compared to fibroblasts cultured in basal condition, but newly only NHDF cells are really induced to increase their synthesis activity. We suggest that the in vitro treatment with GM-CSF can shift human normal fibroblasts towards a more differentiated state, due or accompanied by an increased expression of GM-CSFR and that such “differentiation” is an important event induced by such cytokine.

  17. Exosomes isolation and characterization in serum is feasible in non-small cell lung cancer patients: critical analysis of evidence and potential role in clinical practice.

    Science.gov (United States)

    Taverna, Simona; Giallombardo, Marco; Gil-Bazo, Ignacio; Carreca, Anna Paola; Castiglia, Marta; Chacártegui, Jorge; Araujo, Antonio; Alessandro, Riccardo; Pauwels, Patrick; Peeters, Marc; Rolfo, Christian

    2016-05-10

    Exosomes are nano-sized vesicles of endolysosomal origin, released by several cytotypes in physiological and pathological conditions. Tumor derived exosomes, interacting with other cells of the tumor microenvironment, modulate tumor progression, angiogenic switch, metastasis, and immune escape. Recently, extracellular vesicles were proposed as excellent biomarkers for disease monitoring and prognosis in cancer patients. Non-small cell lung cancer (NSCLC) has a poor 5-year survival rate due to the delay in the detection of the disease. The majority of patients are diagnosed in an advanced disease stage. Exosomes might be promising beneficial tools as biomarker candidates in the scenario of NSCLC, since they contain both, proteins and miRNAs. The clinical case reported in this manuscript is a proof of concept revealing that NSCLC exosomes and sorted miRNAs might constitute, in a near future, novel biomarkers. This review summarizes the role of exosomes in NSCLC, focusing on the importance of exosomal microRNAs in lung cancer diagnosis and prognosis.

  18. Expression of coding (mRNA) and non-coding (microRNA) RNA in lung tissue and blood isolated from pigs suffering from bacterial pleuropneumonia

    DEFF Research Database (Denmark)

    Skovgaard, Kerstin; Schou, Kirstine Klitgaard; Wendt, Karin Tarp

    2010-01-01

    MicroRNAs are small non-coding RNA molecules (18-23 nt), that regulate the activity of other genes at the post-transcriptional level. Recently it has become evident that microRNA plays an important role in modulating and fine tuning innate and adaptive immune responses. Still, little is known about...... the impact of microRNAs in the development and pathogenesis of lung infections. Expression of microRNA known to be induced by bacterial (i.e., LPS) ligands and thus supposed to play a role in the regulation of antimicrobial defence, were studied in lung tissue and in blood from pigs experimentally infected...... with Actinobacillus pleuropneumoniae (AP). Expression differences of mRNA and microRNA were quantified at different time points (6h, 12h, 24h, 48h PI) using reverse transcription quantitative real-time PCR (Rotor-Gene and Fluidigm). Expression profiles of miRNA in blood of seven animals were further studied using mi...

  19. Lung density

    DEFF Research Database (Denmark)

    Garnett, E S; Webber, C E; Coates, G

    1977-01-01

    The density of a defined volume of the human lung can be measured in vivo by a new noninvasive technique. A beam of gamma-rays is directed at the lung and, by measuring the scattered gamma-rays, lung density is calculated. The density in the lower lobe of the right lung in normal man during quiet...... breathing in the sitting position ranged from 0.25 to 0.37 g.cm-3. Subnormal values were found in patients with emphsema. In patients with pulmonary congestion and edema, lung density values ranged from 0.33 to 0.93 g.cm-3. The lung density measurement correlated well with the findings in chest radiographs...... but the lung density values were more sensitive indices. This was particularly evident in serial observations of individual patients....

  20. What Is Lung Cancer?

    Science.gov (United States)

    ... Shareable Graphics Infographics “African-American Men and Lung Cancer” “Lung Cancer Is the Biggest Cancer Killer in Both ... starts in the lungs, it is called lung cancer. Lung cancer begins in the lungs and may spread ...

  1. Abscess in the Lungs

    Science.gov (United States)

    ... Home Lung and Airway Disorders Abscess in the Lungs Abscess in the Lungs Causes Symptoms Diagnosis Treatment Resources ... here for the Professional Version Abscess in the Lungs Abscess in the Lungs A lung abscess is a ...

  2. Heterogeneity in Fibroblast Proliferation and Survival in Idiopathic Pulmonary Fibrosis

    Directory of Open Access Journals (Sweden)

    David Michael Habiel

    2014-01-01

    Full Text Available Idiopathic pulmonary fibrosis (IPF is the most common form of interstitial lung disease characterized by the persistence of activated myofibroblasts resulting in excessive deposition of extracellular matrix proteins and profound tissue remodeling. Myofibroblasts have been shown to arise from interstitial fibroblasts, epithelial to mesenchymal transition of type II alveolar epithelial cells, and the differentiation of recruited fibrocytes. There are many mechanisms that are utilized by these cells for survival, proliferation and persistent activation including up-regulation of cytokines (i.e. Interlukin 6 (IL-6, cytokine receptors (i.e. Interlukin 6 Receptor 1 (IL-6R1, Glycoprotein 130 (gp130 and C-C Chemokine Receptor type 7 (CCR7 and innate pattern recognition receptors (PRRs; i.e. Toll Like Receptor 9 (TLR9. In this review, we will discuss the role of the cytokines IL-6 and CCL21, their receptors and the pattern recognition receptor (PRR, TLR9, in fibroblast recruitment, activation, survival and differentiation into myofibroblasts in IPF.

  3. Botulinum Toxin Type A Inhibits α-Smooth Muscle Actin and Myosin II Expression in Fibroblasts Derived From Scar Contracture.

    Science.gov (United States)

    Chen, Minliang; Yan, Tongtong; Ma, Kui; Lai, Linying; Liu, Chang; Liang, Liming; Fu, Xiaobing

    2016-09-01

    Scar contracture (SC) is one of the most common complications resulting from major burn injuries. Numerous treatments are currently available but they do not always yield excellent therapeutic results. Recent reports suggest that botulinum toxin type A (BTXA) is effective at reducing SC clinically, but the molecular mechanism for this action is unknown. α-Smooth muscle actin (α-SMA) and myosin II are the main components of stress fibers, which are the contractile structures of fibroblasts. The effects of BTXA on α-SMA and myosin II in SC are still unknown. This study aimed to explore the effect of BTXA on α-SMA and myosin II expression in fibroblasts derived from SC and to elucidate its actual mechanism further. Fibroblasts were isolated from tissue specimens of SC. Fibroblasts were cultured in Dulbecco modified Eagle medium with different concentrations of BTXA and their proliferation was analyzed through the tetrazolium-based colorimetric method at 1, 4, and 7 days. Proteins of α-SMA and myosin II were checked using Western blot in fibroblasts treated with different concentrations of BTXA at 1, 4, and 7 days. Fibroblasts without BTXA treatment had a higher proliferation than that in other groups, which indicated that the proliferation of fibroblasts was significantly inhibited by BTXA (P < 0.05). Proteins of α-SMA and myosin II between fibroblasts with BTXA and fibroblasts without BTXA are statistically significant (P < 0.05). These results suggest that BTXA effectively inhibited the growth of fibroblasts derived from SC and reduced the expression of α-SMA and myosin II, which provided theoretical support for the application of BTXA to control SC.

  4. Microporous dermal-mimetic electrospun scaffolds pre-seeded with fibroblasts promote tissue regeneration in full-thickness skin wounds.

    Directory of Open Access Journals (Sweden)

    Paul P Bonvallet

    Full Text Available Electrospun scaffolds serve as promising substrates for tissue repair due to their nanofibrous architecture and amenability to tailoring of chemical composition. In this study, the regenerative potential of a microporous electrospun scaffold pre-seeded with dermal fibroblasts was evaluated. Previously we reported that a 70% collagen I and 30% poly(Ɛ-caprolactone electrospun scaffold (70:30 col/PCL containing 160 μm diameter pores had favorable mechanical properties, supported fibroblast infiltration and subsequent cell-mediated deposition of extracellular matrix (ECM, and promoted more rapid and effective in vivo skin regeneration when compared to scaffolds lacking micropores. In the current study we tested the hypothesis that the efficacy of the 70:30 col/PCL microporous scaffolds could be further enhanced by seeding scaffolds with dermal fibroblasts prior to implantation into skin wounds. To address this hypothesis, a Fischer 344 (F344 rat syngeneic model was employed. In vitro studies showed that dermal fibroblasts isolated from F344 rat skin were able to adhere and proliferate on 70:30 col/PCL microporous scaffolds, and the cells also filled the 160 μm pores with native ECM proteins such as collagen I and fibronectin. Additionally, scaffolds seeded with F344 fibroblasts exhibited a low rate of contraction (~14% over a 21 day time frame. To assess regenerative potential, scaffolds with or without seeded F344 dermal fibroblasts were implanted into full thickness, critical size defects created in F344 hosts. Specifically, we compared: microporous scaffolds containing fibroblasts seeded for 4 days; scaffolds containing fibroblasts seeded for only 1 day; acellular microporous scaffolds; and a sham wound (no scaffold. Scaffolds containing fibroblasts seeded for 4 days had the best response of all treatment groups with respect to accelerated wound healing, a more normal-appearing dermal matrix structure, and hair follicle regeneration

  5. Excision of gamma-ray induced thymine lesions by preparations from ataxia telangiectasia fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Remsen, J F; Cerutti, P A [Florida Univ., Gainesville (USA). Inst. of Food and Agricultural Sciences; Florida Univ., Gainesville (USA). Dept. of Biochemistry)

    1977-04-01

    The capacity of whole cell sonicates of skin fibroblasts of normal individuals and patients with the autosomal recessive disease Ataxia telangiectasia (AT) to remove aerobic gamma-ray products of the 5,6-dihydroxydihydrothymine type (tsub(O/sub 2/)sup(..gamma..)) from exogenous DNA substrates was investigated. All four AT strains (AT CRL 1312, AT CRL 1343, AT GM 367, AT 4BI) possessed normal capabilities to excise tsub(O/sub 2/)sup(..gamma..) from irradiated bacteriophage DNA and irradiated chromatin isolated from normal and AT-skin fibroblasts.

  6. Targeting tissue factor as a novel therapeutic oncotarget for eradication of cancer stem cells isolated from tumor cell lines, tumor xenografts and patients of breast, lung and ovarian cancer.

    Science.gov (United States)

    Hu, Zhiwei; Xu, Jie; Cheng, Jijun; McMichael, Elizabeth; Yu, Lianbo; Carson, William E

    2017-01-03

    Targeting cancer stem cell (CSC) represents a promising therapeutic approach as it can potentially fight cancer at its root. The challenge is to identify a surface therapeutic oncotarget on CSC. Tissue factor (TF) is known as a common yet specific surface target for cancer cells and tumor neovasculature in several solid cancers. However, it is unknown if TF is expressed by CSCs. Here we demonstrate that TF is constitutively expressed on CD133 positive (CD133+) or CD24-CD44+ CSCs isolated from human cancer cell lines, tumor xenografts from mice and breast tumor tissues from patients. TF-targeted agents, i.e., a factor VII (fVII)-conjugated photosensitizer (fVII-PS for targeted photodynamic therapy) and fVII-IgG1Fc (Immunoconjugate or ICON for immunotherapy), can eradicate CSC via the induction of apoptosis and necrosis and via antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity, respectively. In conclusion, these results demonstrate that TF is a novel surface therapeutic oncotarget for CSC, in addition to cancer cell TF and tumor angiogenic vascular endothelial TF. Moreover, this research highlights that TF-targeting therapeutics can effectively eradicate CSCs, without drug resistance, isolated from breast, lung and ovarian cancer with potential to translate into other most commonly diagnosed solid cancer, in which TF is also highly expressed.

  7. Impact of matrix stiffness on fibroblast function

    Energy Technology Data Exchange (ETDEWEB)

    El-Mohri, Hichem; Wu, Yang; Mohanty, Swetaparna; Ghosh, Gargi, E-mail: gargi@umich.edu

    2017-05-01

    Chronic non-healing wounds, caused by impaired production of growth factors and reduced vascularization, represent a significant burden to patients, health care professionals, and health care system. While several wound dressing biomaterials have been developed, the impact of the mechanical properties of the dressings on the residing cells and consequently on the healing of the wounds is largely overlooked. The primary focus of this study is to explore whether manipulation of the substrate mechanics can regulate the function of fibroblasts, particularly in the context of their angiogenic activity. A photocrosslinkable hydrogel platform with orthogonal control over gel modulus and cell adhesive sites was developed to explore the quantitative relationship between ECM compliance and fibroblast function. Increase in matrix stiffness resulted in enhanced fibroblast proliferation and stress fiber formation. However, the angiogenic activity of fibroblasts was found to be optimum when the cells were seeded on compliant matrices. Thus, the observations suggest that the stiffness of the wound dressing material may play an important role in the progression of wound healing. - Highlights: • Proliferation and stress fiber formation of fibroblasts increase with increasing matrix mechanics. • Cell area correlates with the growth of fibroblasts. • Angiogenic activity of fibroblasts optimum when cells seeded on compliant gels.

  8. Silencing Histone Deacetylase 7 Alleviates Transforming Growth Factor-β1-Induced Profibrotic Responses in Fibroblasts Derived from Peyronie’s Plaque

    Directory of Open Access Journals (Sweden)

    Dong Hyuk Kang

    2018-05-01

    Full Text Available Purpose: Epigenetic modifications, such as histone acetylation/deacetylation and DNA methylation, play a crucial role in the pathogenesis of inflammatory disorders and fibrotic diseases. The aim of this study was to study the differential gene expression of histone deacetylases (HDACs in fibroblasts isolated from plaque tissue of Peyronie’s disease (PD or normal tunica albuginea (TA and to examine the anti-fibrotic effect of small interfering RNA (siRNA-mediated silencing of HDAC7 in fibroblasts derived from human PD plaque. Materials and Methods: For differential gene expression study, we performed reverse-transcriptase polymerase chain reaction for HDAC isoforms (1–11 in fibroblasts isolated from PD plaque or normal TA. Fibroblasts isolated from PD plaque were pretreated with HDAC7 siRNA (100 pmol and then stimulated with transforming growth factor-β1 (TGF-β1, 10 ng/mL. Protein was extracted from treated fibroblasts for Western blotting. We also performed immunocytochemistry to detect the expression of extracellular matrix proteins and to examine the effect of HDAC2 siRNA on the TGF-β1-induced nuclear translocation of Smad2/3 and myofibroblastic differentiation. Results: The mRNA expression of HDAC2, 3, 4, 5, 7, 8, 10, and 11 was higher in fibroblasts isolated from PD plaque than in fibroblasts isolated from normal TA tissue. Knockdown of HDAC7 in PD fibroblasts inhibited TGF-β1-induced nuclear shuttle of Smad2 and Smad3, transdifferentiation of fibroblasts into myofibroblasts, and abrogated TGF-β1-induced production of extracellular matrix protein. Conclusions: These findings suggest that specific inhibition of HDAC7 with RNA interference may represent a promising epigenetic therapy for PD.

  9. Re-evaluation of lung to thorax transverse area ratio immediately before birth in predicting postnatal short-term outcomes of fetuses with isolated left-sided congenital diaphragmatic hernia: A single center analysis.

    Science.gov (United States)

    Kido, Saki; Hidaka, Nobuhiro; Sato, Yuka; Fujita, Yasuyuki; Miyoshi, Kina; Nagata, Kouji; Taguchi, Tomoaki; Kato, Kiyoko

    2018-05-01

    We aimed to investigate whether the lung-to-thorax transverse area ratio (LTR) immediately before birth is of diagnostic value for the prediction of postnatal short-term outcomes in cases of isolated left-sided congenital diaphragmatic hernia (CDH). We retrospectively reviewed the cases of fetal isolated left-sided CDH managed at our institution between April 2008 and July 2016. We divided the patients into two groups based on LTR immediately before birth, using a cut-off value of 0.08. We compared the proportions of subjects within the two groups who survived until discharge using Fisher's exact test. Further, using Spearman's rank correlation, we assessed whether LTR was correlated with length of stay, duration of mechanical ventilation, and supplemental oxygen. Twenty-nine subjects were included (five with LTR < 0.08, and 24 with LTR ≥ 0.08). The proportion of subjects surviving until discharge was 40% (2/5) for patients with LTR < 0.08, as compared with 96% (23/24) for those with LTR ≥ 0.08. LTR measured immediately before birth was negatively correlated with the postnatal length of stay (Spearman's rank correlation coefficient, rs = -0.486), and the duration of supplemental oxygen (rs = -0.537). Further, the duration of mechanical ventilation was longer in patients with a lower LTR value. LTR immediately before birth is useful for the prediction of postnatal short-term outcomes in fetuses with isolated left-sided CDH. In particular, patients with prenatal LTR value less than 0.08 are at increased risk of postnatal death. © 2017 Japanese Teratology Society.

  10. The mitochondrial genome of Paragonimus westermani (Kerbert, 1878, the Indian isolate of the lung fluke representative of the family Paragonimidae (Trematoda

    Directory of Open Access Journals (Sweden)

    Devendra K. Biswal

    2014-08-01

    Full Text Available Among helminth parasites, Paragonimus (zoonotic lung fluke gains considerable importance from veterinary and medical points of view because of its diversified effect on its host. Nearly fifty species of Paragonimus have been described across the globe. It is estimated that more than 20 million people are infected worldwide and the best known species is Paragonimus westermani, whose type locality is probably India and which infects millions of people in Asia causing disease symptoms that mimic tuberculosis. Human infections occur through eating raw crustaceans containing metacercarie or ingestion of uncooked meat of paratenic hosts such as pigs. Though the fluke is known to parasitize a wide range of mammalian hosts representing as many as eleven families, the status of its prevalence, host range, pathogenic manifestations and its possible survivors in nature from where the human beings contract the infection is not well documented in India. We took advantage of the whole genome sequence data for P. westermani, generated by Next Generation Sequencing, and its comparison with the existing data for the P. westermani for comparative mt DNA phylogenomic analyses. Specific primers were designed for the 12 protein coding genes with the aid of existing P. westermani mtDNA as the reference. The Ion torrent next generation sequencing platform was harnessed to completely sequence the mitochondrial genome, and applied innovative approaches to bioinformatically assemble and annotate it. A strategic PCR primer design utilizing the whole genome sequence data from P. westermani enabled us to design specific primers capable of amplifying all regions of the mitochondrial genome from P. westermani. Assembly of NGS data from libraries enriched in mtDNA sequence by PCR gave rise to a total of 11 contigs spanning the entire 14.7 kb mt DNA sequence of P. westermani available at NCBI. We conducted gap-filling by traditional Sanger sequencing to fill in the gaps

  11. Increased alveolar soluble Annexin V promotes lung inflammation and fibrosis

    OpenAIRE

    Buckley, S.; Shi, W.; Xu, W.; Frey, M.R.; Moats, R.; Pardo, A.; Selman, M.; Warburton, D.

    2015-01-01

    The causes underlying the self-perpetuating nature of idiopathic pulmonary fibrosis (IPF), a progressive and usually lethal disease, remain unknown. We hypothesized that alveolar soluble Annexin V contributes to lung fibrosis, based on the observation that human IPF BALF containing high Annexin V levels promoted fibroblast involvement in alveolar epithelial wound healing that was reduced when Annexin V was depleted from the BALF.

  12. Extracellular matrix proteins: a positive feedback loop in lung fibrosis?

    NARCIS (Netherlands)

    Blaauboer, M.E.; van Boeijen, F.R.; Emson, C.L.; Turner, S.M.; Zandieh-Doulabi, B.; Hanemaaijer, R.; Smit, T.H.; Stoop, R.; Everts, V.

    2014-01-01

    Lung fibrosis is characterized by excessive deposition of extracellular matrix. This not only affects tissue architecture and function, but it also influences fibroblast behavior and thus disease progression. Here we describe the expression of elastin, type V collagen and tenascin C during the

  13. Extracellular matrix proteins: A positive feedback loop in lung fibrosis?

    NARCIS (Netherlands)

    Blaauboer, M.E.; Boeijen, F.R.; Emson, C.L.; Turner, S.M.; Zandieh-Doulabi, B.; Hanemaaijer, R.; Smit, T.H.; Stoop, R.; Everts, V.

    2014-01-01

    Lung fibrosis is characterized by excessive deposition of extracellular matrix. This not only affects tissue architecture and function, but it also influences fibroblast behavior and thus disease progression. Here we describe the expression of elastin, type V collagen and tenascin C during the

  14. Lung Cancer

    International Nuclear Information System (INIS)

    Maghfoor, Irfan; Perry, M.C.

    2005-01-01

    Lung cancer is the leading cause of cancer-related mortality. Since tobacco smoking is the cause in vast majority of cases, the incidence of lung cancer is expected to rise in those countries with high or rising incidence of tobacco smoking. Even though population at a risk of developing lung cancer are easily identified, mass screening for lung cancer is not supported by currently available evidence. In case of non-small cell lung cancer, a cure may be possible with surgical resection followed by post-operative chemotherapy in those diagnosed at an early stage. A small minority of patients who present with locally advanced disease may also benefit from preoperative chemotherapy and/or radiation therapy to down stage the tumor to render it potentially operable. In a vast majority of patients, however, lung cancer presents at an advanced stage and a cure is not possible with currently available therapeutic strategies. Similarly small cell lung cancer confined to one hemi-thorax may be curable with a combination of chemotherapy and thoracic irradiation followed by prophylactic cranial irradiation, if complete remission is achieved at the primary site. Small cell lung cancer that is spread beyond the confines of one hemi-thorax is however, considered incurable. In this era of molecular targeted therapies, new agents are constantly undergoing pre-clinical and clinical testing with the aim of targeting the molecular pathways thought to involved in etiology and pathogenesis of lung cancer. (author)

  15. Fibroblast cultures in duchenne muscular dystrophy

    International Nuclear Information System (INIS)

    Ionasescu, V.; Lara-Braud, C.; Zellweger, H.; Ionasescu, R.; Burmeister, L.

    1977-01-01

    Primary skin fibroblast cultures were grown from forearm pinch skin biopsies obtained from 24 patients with Duchenne muscular dystrophy (DMD) and ten normal controls matched for sex and age. The first subcultures were grown for 7 days and incubated with L-( 3 H)-proline for 24 hours. Intracellular collagen incoption was significantly decreased (2.2 X) and extracellular collagen incorporation significantly increased (1.8 X) in fibroblast cultures from patients with DMD by both collagenase assay and polyacrylamide gel electrophoresis. The synthesis of noncollagen proteins showed low values from the DMD fibroblast cultures. The alterations in synthesis and secretion of collagen and noncollagen proteins were characteristic only for the log phase of DMD fibroblasts. (author)

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

    Science.gov (United States)

    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.

  17. Capilliposide Isolated from Lysimachia capillipes Hemsl. Induces ROS Generation, Cell Cycle Arrest, and Apoptosis in Human Nonsmall Cell Lung Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Zheng-hua Fei

    2014-01-01

    Full Text Available Several data has reported that capilliposide, extracted from a traditional Chinese medicine, Lysimachia capillipes Hemsl. (LC could exhibit inhibitory effect on cell proliferation in various cancers. The current study investigated the antitumor efficacy of Capilliposide and elucidated its potential molecular mechanism involved in vivo and vitro. Our results indicated that LC capilliposide inhibited proliferation of lung cancer cells in a dose-dependent manner. LC capilliposide induced cell cycle arrest at the S stage and enhanced apoptosis in NSCLC cells. Treatment with LC capilliposide increased the intracellular level of ROS, which activated the mitochondrial apoptotic pathway. Blockage of ROS by NAC highly reversed the effect of LC capilliposide on apoptosis. Xenograft tumor growth was significantly lower in the LC-treated group compared with the untreated control group (P<0.05. The results also show that LC treatment does not produce any overt signs of acute toxicity in vivo. These findings demonstrate that LC capilliposide could exert an anti-tumor effect on NSCLC through mitochondrial-mediated apoptotic pathway and the activation of ROS is involved.

  18. Colony size distributions according to in vitro aging in human skin fibroblasts

    International Nuclear Information System (INIS)

    Kim, Jun Sang; Kim, Jae Sung; Cho, Moon June; Park, Jeong Kyu; Paik, Tae Hyun

    1999-01-01

    To investigate the percentage of colonies with 16 or more cells distribution of human skin fibroblast according to in vitro aging, and to evaluate the relationship between percentage of colonies with 16 or more cells and in vivo donor age in human skin fibroblast culture. C1, C2, C3a, and C3b human skin fibroblast samples from three breast cancer patients were used as subjects. The C1, C2, and C3a donor were 44, 54, and 55 years old, respectively. C3a and C3b cells were isolated from the same person. Single cell suspension of skin fibroblasts was prepared with primary explant technique. One hundred cells are plated into 100ml tissue culture flask and cultured for two weeks. The colony size was defined as colonies with 16 or more cells. The cultured cell was stained with crystal violet, and number of cells in each colony was determined with stereo microscope at x 10 magnification. Passage number of C1, C2, C3a and C3b skin fibroblast were 12th, 17th, and 14th, respectively. Percentage of colonies with 16 or more cells of skin fibroblast samples decreased with increasing in vitro passage number. In contrast, cumulative population doublings of skin fibroblast sample increased with increasing in vitro passage number. Percentage of colonies with 16 or more cells also decreased with increasing population doublings in human skin fibroblast culture. There was strong correlation with percentage of colonised with 16 or more cells and population doublings in C3a skin fibroblast sample. At the same point of population doublings, the percentage of colonies with 16 or more cells of the young C1 donor was higher level than the old C3a donor. The population doublings increased with increasing in vitro passage number but percentage of colonies with 16 or more cells decreased. The results of this study imply that percentage of colonies with 16 or more cells is useful as a indicator of in vitro human skin fibroblast aging and may estimate the in vivo donor age

  19. Identification of colonic fibroblast secretomes reveals secretory factors regulating colon cancer cell proliferation.

    Science.gov (United States)

    Chen, Sun-Xia; Xu, Xiao-En; Wang, Xiao-Qing; Cui, Shu-Jian; Xu, Lei-Lei; Jiang, Ying-Hua; Zhang, Yang; Yan, Hai-Bo; Zhang, Qian; Qiao, Jie; Yang, Peng-Yuan; Liu, Feng

    2014-10-14

    Stromal microenvironment influences tumor cell proliferation and migration. Fibroblasts represent the most abundant stromal constituents. Here, we established two pairs of normal fibroblast (NF) and cancer-associated fibroblast (CAF) cultures from colorectal adenocarcinoma tissues and the normal counterparts. The NFs and CAFs were stained positive for typical fibroblast markers and inhibited colon cancer (CC) cell proliferation in in vitro cocultures and in xenograft mouse models. The fibroblast conditioned media were analyzed using LC-MS and 227 proteins were identified at a false discovery rate of 1.3%, including 131 putative secretory and 20 plasma membrane proteins. These proteins were enriched for functional categories of extracellular matrix, adhesion, cell motion, inflammatory response, redox homeostasis and peptidase inhibitor. Secreted protein acidic and rich in cysteine, transgelin, follistatin-related protein 1 (FSTL1) and decorin was abundant in the fibroblast secretome as confirmed by Western blot. Silencing of FSTL1 and transgelin in colonic fibroblast cell line CCD-18Co induced an accelerated proliferation of CC cells in cocultures. Exogenous FSTL1 attenuates CC cell proliferation in a negative fashion. FSTL1 was upregulated in CC patient plasma and cancerous tissues but had no implication in prognosis. Our results provided novel insights into the molecular signatures and modulatory role of CC associated fibroblasts. In this study, a label-free LC-MS was performed to analyze the secretomes of two paired primary fibroblasts, which were isolated from fresh surgical specimen of colorectal adenocarcinoma and adjacent normal colonic tissues and exhibited negative modulatory activity for colon cancer cell growth in in vitro cocultures and in vivo xenograph mouse models. Follistatin-related protein 1 was further revealed to be one of the stroma-derived factors of potential suppression role for colon cancer cell proliferation. Our results provide novel

  20. Aspirin suppresses cardiac fibroblast proliferation and collagen formation through downregulation of angiotensin type 1 receptor transcription

    International Nuclear Information System (INIS)

    Wang, Xianwei; Lu, Jingjun; Khaidakov, Magomed; Mitra, Sona; Ding, Zufeng; Raina, Sameer; Goyal, Tanu; Mehta, Jawahar L.

    2012-01-01

    Aspirin (acetyl salicylic acid, ASA) is a common drug used for its analgesic and antipyretic effects. Recent studies show that ASA not only blocks cyclooxygenase, but also inhibits NADPH oxidase and resultant reactive oxygen species (ROS) generation, a pathway that underlies pathogenesis of several ailments, including hypertension and tissue remodeling after injury. In these disease states, angiotensin II (Ang II) activates NADPH oxidase via its type 1 receptor (AT1R) and leads to fibroblast growth and collagen synthesis. In this study, we examined if ASA would inhibit NADPH oxidase activation, upregulation of AT1R transcription, and subsequent collagen generation in mouse cardiac fibroblasts challenged with Ang II. Mouse heart fibroblasts were isolated and treated with Ang II with or without ASA. As expected, Ang II induced AT1R expression, and stimulated cardiac fibroblast growth and collagen synthesis. The AT1R blocker losartan attenuated these effects of Ang II. Similarly to losartan, ASA, and its SA moiety suppressed Ang II-mediated AT1R transcription and fibroblast proliferation as well as expression of collagens and MMPs. ASA also suppressed the expression of NADPH oxidase subunits (p22 phox , p47 phox , p67 phox , NOX2 and NOX4) and ROS generation. ASA did not affect total NF-κB p65, but inhibited its phosphorylation and activation. These observations suggest that ASA inhibits Ang II-induced NADPH oxidase expression, NF-κB activation and AT1R transcription in cardiac fibroblasts, and fibroblast proliferation and collagen expression. The critical role of NADPH oxidase activity in stimulation of AT1R transcription became apparent in experiments where ASA also inhibited AT1R transcription in cardiac fibroblasts challenged with H 2 O 2 . Since SA had similar effect as ASA on AT1R expression, we suggest that ASA's effect is mediated by its SA moiety. -- Highlights: ► Aspirin in therapeutic concentrations decreases mouse cardiac fibroblast growth and collagen

  1. Aspirin suppresses cardiac fibroblast proliferation and collagen formation through downregulation of angiotensin type 1 receptor transcription

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Xianwei, E-mail: XWang2@UAMS.edu; Lu, Jingjun; Khaidakov, Magomed; Mitra, Sona; Ding, Zufeng; Raina, Sameer; Goyal, Tanu; Mehta, Jawahar L., E-mail: MehtaJL@UAMS.edu

    2012-03-15

    Aspirin (acetyl salicylic acid, ASA) is a common drug used for its analgesic and antipyretic effects. Recent studies show that ASA not only blocks cyclooxygenase, but also inhibits NADPH oxidase and resultant reactive oxygen species (ROS) generation, a pathway that underlies pathogenesis of several ailments, including hypertension and tissue remodeling after injury. In these disease states, angiotensin II (Ang II) activates NADPH oxidase via its type 1 receptor (AT1R) and leads to fibroblast growth and collagen synthesis. In this study, we examined if ASA would inhibit NADPH oxidase activation, upregulation of AT1R transcription, and subsequent collagen generation in mouse cardiac fibroblasts challenged with Ang II. Mouse heart fibroblasts were isolated and treated with Ang II with or without ASA. As expected, Ang II induced AT1R expression, and stimulated cardiac fibroblast growth and collagen synthesis. The AT1R blocker losartan attenuated these effects of Ang II. Similarly to losartan, ASA, and its SA moiety suppressed Ang II-mediated AT1R transcription and fibroblast proliferation as well as expression of collagens and MMPs. ASA also suppressed the expression of NADPH oxidase subunits (p22{sup phox}, p47{sup phox}, p67{sup phox}, NOX2 and NOX4) and ROS generation. ASA did not affect total NF-κB p65, but inhibited its phosphorylation and activation. These observations suggest that ASA inhibits Ang II-induced NADPH oxidase expression, NF-κB activation and AT1R transcription in cardiac fibroblasts, and fibroblast proliferation and collagen expression. The critical role of NADPH oxidase activity in stimulation of AT1R transcription became apparent in experiments where ASA also inhibited AT1R transcription in cardiac fibroblasts challenged with H{sub 2}O{sub 2}. Since SA had similar effect as ASA on AT1R expression, we suggest that ASA's effect is mediated by its SA moiety. -- Highlights: ► Aspirin in therapeutic concentrations decreases mouse cardiac

  2. Establishment of a pig fibroblast-derived cell line for locus-directed transgene expression in cell cultures and blastocysts

    DEFF Research Database (Denmark)

    Jakobsen, Jannik E; Li, Juan; Moldt, Brian

    2011-01-01

    We report the establishment of a spontaneously immortalized pig cell line designated Pig Flip-in Visualize (PFV) for locus-directed transgene expression in pig cells and blastocysts. The PFV cell line was isolated from pig ear fibroblasts transfected with a Sleeping Beauty DNA transposon-based do......We report the establishment of a spontaneously immortalized pig cell line designated Pig Flip-in Visualize (PFV) for locus-directed transgene expression in pig cells and blastocysts. The PFV cell line was isolated from pig ear fibroblasts transfected with a Sleeping Beauty DNA transposon...

  3. Stereotactic Ablative Radiation Therapy for Centrally Located Early Stage or Isolated Parenchymal Recurrences of Non-Small Cell Lung Cancer: How to Fly in a “No Fly Zone”

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Joe Y., E-mail: jychang@mdanderson.org [Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Li, Qiao-Qiao; Xu, Qing-Yong; Allen, Pamela K.; Rebueno, Neal; Gomez, Daniel R. [Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Balter, Peter [Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Komaki, Ritsuko [Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Mehran, Reza; Swisher, Stephen G.; Roth, Jack A. [Department of Thoracic and Cardiovascular Surgery, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States)

    2014-04-01

    Purpose: We extended our previous experience with stereotactic ablative radiation therapy (SABR; 50 Gy in 4 fractions) for centrally located non-small cell lung cancer (NSCLC); explored the use of 70 Gy in 10 fractions for cases in which dose-volume constraints could not be met with the previous regimen; and suggested modified dose-volume constraints. Methods and Materials: Four-dimensional computed tomography (4DCT)-based volumetric image-guided SABR was used for 100 patients with biopsy-proven, central T1-T2N0M0 (n=81) or isolated parenchymal recurrence of NSCLC (n=19). All disease was staged with positron emission tomography/CT; all tumors were within 2 cm of the bronchial tree, trachea, major vessels, esophagus, heart, pericardium, brachial plexus, or vertebral body. Endpoints were toxicity, overall survival (OS), local and regional control, and distant metastasis. Results: At a median follow-up time of 30.6 months, median OS time was 55.6 months, and the 3-year OS rate was 70.5%. Three-year cumulative actuarial local, regional, and distant control rates were 96.5%, 87.9%, and 77.2%, respectively. The most common toxicities were chest-wall pain (18% grade 1, 13% grade 2) and radiation pneumonitis (11% grade 2 and 1% grade 3). No patient experienced grade 4 or 5 toxicity. Among the 82 patients receiving 50 Gy in 4 fractions, multivariate analyses showed mean total lung dose >6 Gy, V{sub 20} >12%, or ipsilateral lung V{sub 30} >15% to independently predict radiation pneumonitis; and 3 of 9 patients with brachial plexus D{sub max} >35 Gy experienced brachial neuropathy versus none of 73 patients with brachial D{sub max} <35 Gy (P=.001). Other toxicities were analyzed and new dose-volume constraints are proposed. Conclusions: SABR for centrally located lesions produces clinical outcomes similar to those for peripheral lesions when normal tissue constraints are respected.

  4. Fibroblasts in fibrosis: novel roles and mediators

    Directory of Open Access Journals (Sweden)

    Ryan Thomas Kendall

    2014-05-01

    Full Text Available Fibroblasts are the most common cell type of the connective tissues found throughout the body and the principal source of the extensive extracellular matrix (ECM characteristic of these tissues. They are also the central mediators of the pathological fibrotic accumulation of ECM and the cellular proliferation and differentiation that occurs in response to prolonged tissue injury and chronic inflammation. The transformation of the fibroblast cell lineage involves classical developmental signaling programs and includes a surprisingly diverse range of precursor cell types—most notably, myofibroblasts that are the apex of the fibrotic phenotype. Myofibroblasts display exaggerated ECM production; constitutively secrete and are hypersensitive to chemical signals such as cytokines, chemokines, and growth factors; and are endowed with a contractile apparatus allowing them to manipulate the ECM fibers physically to close open wounds. In addition to ECM production, fibroblasts have multiple concomitant biological roles, such as in wound healing, inflammation, and angiogenesis, which are each interwoven with the process of fibrosis. We now recognize many common fibroblast-related features across various physiological and pathological protracted processes. Indeed, a new appreciation has emerged for the role of noncancerous fibroblast interactions with tumors in cancer progression. Although the predominant current clinical treatments of fibrosis involve nonspecific immunosuppressive and anti-proliferative drugs, a variety of potential therapies under investigation specifically target fibroblast biology.

  5. Mucosal stromal fibroblasts markedly enhance HIV infection of CD4+ T cells.

    Directory of Open Access Journals (Sweden)

    Jason A Neidleman

    2017-02-01

    Full Text Available Understanding early events of HIV transmission within mucosal tissues is vital for developing effective prevention strategies. Here, we report that primary stromal fibroblasts isolated from endometrium, cervix, foreskin, male urethra, and intestines significantly increase HIV infection of CD4+ T cells-by up to 37-fold for R5-tropic HIV and 100-fold for X4-tropic HIV-without themselves becoming infected. Fibroblasts were more efficient than dendritic cells at trans-infection and mediate this response in the absence of the DC-SIGN and Siglec-1 receptors. In comparison, mucosal epithelial cells secrete antivirals and inhibit HIV infection. These data suggest that breaches in the epithelium allow external or luminal HIV to escape an antiviral environment to access the infection-favorable environment of the stromal fibroblasts, and suggest that resident fibroblasts have a central, but previously unrecognized, role in HIV acquisition at mucosal sites. Inhibiting fibroblast-mediated enhancement of HIV infection should be considered as a novel prevention strategy.

  6. Differential Gene Expression in Primary Human Skin Keratinocytes and Fibroblasts in Response to Ionizing Radiation

    Science.gov (United States)

    Warters, Raymond L.; Packard, Ann T.; Kramer, Gwen F.; Gaffney, David K.; Moos, Philip J.

    2009-01-01

    Although skin is usually exposed during human exposures to ionizing radiation, there have been no thorough examinations of the transcriptional response of skin fibroblasts and keratinocytes to radiation. The transcriptional response of quiescent primary fibroblasts and keratinocytes exposed to from 10 cGy to 5 Gy and collected 4 h after treatment was examined. RNA was isolated and examined by microarray analysis for changes in the levels of gene expression. Exposure to ionizing radiation altered the expression of 279 genes across both cell types. Changes in RNA expression could be arranged into three main categories: (1) changes in keratinocytes but not in fibroblasts, (2) changes in fibroblasts but not in keratinocytes, and (3) changes in both. All of these changes were primarily of p53 target genes. Similar radiation-induced changes were induced in immortalized fibroblasts or keratinocytes. In separate experiments, protein was collected and analyzed by Western blotting for expression of proteins observed in microarray experiments to be overexpressed at the mRNA level. Both Q-PCR and Western blot analysis experiments validated these transcription changes. Our results are consistent with changes in the expression of p53 target genes as indicating the magnitude of cell responses to ionizing radiation. PMID:19580510

  7. Fibroblast-Derived Extracellular Matrices: An Alternative Cell Culture System That Increases Metastatic Cellular Properties.

    Directory of Open Access Journals (Sweden)

    Michael T Scherzer

    Full Text Available Poor survival rates from lung cancer can largely be attributed to metastatic cells that invade and spread throughout the body. The tumor microenvironment (TME is composed of multiple cell types, as well as non-cellular components. The TME plays a critical role in the development of metastatic cancers by providing migratory cues and changing the properties of the tumor cells. The Extracellular Matrix (ECM, a main component of the TME, has been shown to change composition during tumor progression, contributing to cancer cell invasion and survival away from the primary cancer site. Although the ECM is well-known to influence the fate of tumor progression, little is known about the molecular mechanisms that are affected by the cancer cell-ECM interactions. It is imperative that these mechanisms are elucidated in order to properly understand and prevent lung cancer dissemination. However, common in vitro studies do not incorporate these interactions into everyday cell culture assays. We have adopted a model that examines decellularized human fibroblast-derived ECM as a 3-dimensional substrate for growth of lung adenocarcinoma cell lines. Here, we have characterized the effect of fibroblast-derived matrices on the properties of various lung-derived epithelial cell lines, including cancerous and non-transformed cells. This work highlights the significance of the cell-ECM interaction and its requirement for incorporation into in vitro experiments. Implementation of a fibroblast-derived ECM as an in vitro technique will provide researchers with an important factor to manipulate to better recreate and study the TME.

  8. 25-Hydroxycholesterol promotes fibroblast-mediated tissue remodeling through NF-κB dependent pathway

    Energy Technology Data Exchange (ETDEWEB)

    Ichikawa, Tomohiro [Third Department of Internal Medicine, Wakayama Medical University, School of Medicine, 811-1 Kimiidera, Wakayama 641-8509 (Japan); Sugiura, Hisatoshi, E-mail: sugiura@rm.med.tohoku.ac.jp [Department of Respiratory Medicine, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574 (Japan); Koarai, Akira; Kikuchi, Takashi; Hiramatsu, Masataka; Kawabata, Hiroki; Akamatsu, Keiichiro; Hirano, Tsunahiko; Nakanishi, Masanori; Matsunaga, Kazuto; Minakata, Yoshiaki [Third Department of Internal Medicine, Wakayama Medical University, School of Medicine, 811-1 Kimiidera, Wakayama 641-8509 (Japan); Ichinose, Masakazu [Department of Respiratory Medicine, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574 (Japan)

    2013-05-01

    Abnormal structural alterations termed remodeling, including fibrosis and alveolar wall destruction, are important features of the pathophysiology of chronic airway diseases such as chronic obstructive pulmonary disease (COPD) and asthma. 25-hydroxycholesterol (25-HC) is enzymatically produced by cholesterol 25-hydorxylase (CH25H) in macrophages and is reported to be involved in the formation of arteriosclerosis. We previously demonstrated that the expression of CH25H and production of 25HC were increased in the lungs of COPD. However, the role of 25-HC in lung tissue remodeling is unknown. In this study, we investigated the effect of 25-HC on fibroblast-mediated tissue remodeling using human fetal lung fibroblasts (HFL-1) in vitro. 25-HC significantly augmented α-smooth muscle actin (SMA) (P<0.001) and collagen I (P<0.001) expression in HFL-1. 25-HC also significantly enhanced the release and activation of matrix metallaoproteinase (MMP)-2 (P<0.001) and MMP-9 (P<0.001) without any significant effect on the production of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2. 25-HC stimulated transforming growth factor (TGF)-β{sub 1} production (P<0.01) and a neutralizing anti-TGF-β antibody restored these 25-HC-augmented pro-fibrotic responses. 25-HC significantly promoted the translocation of nuclear factor (NF)-κB p65 into the nuclei (P<0.01), but not phospholylated-c-jun, a complex of activator protein-1. Pharmacological inhibition of NF-κB restored the 25-HC-augmented pro-fibrotic responses and TGF-β{sub 1} release. These results suggest that 25-HC could contribute to fibroblast-mediated lung tissue remodeling by promoting myofibroblast differentiation and the excessive release of extracellular matrix protein and MMPs via an NF-κB-TGF-β dependent pathway.

  9. Lung Cancer

    Science.gov (United States)

    Lung cancer is one of the most common cancers in the world. It is a leading cause of cancer death in men and women in the United States. Cigarette smoking causes most lung cancers. The more cigarettes you smoke per day and ...

  10. Lung Cancer Screening

    Science.gov (United States)

    ... factors increase or decrease the risk of lung cancer. Lung cancer is a disease in which malignant (cancer) ... following PDQ summaries for more information about lung cancer: Lung Cancer Prevention Non-Small Cell Lung Cancer Treatment ...

  11. Concentration-dependent effect of platelet-rich plasma on keratinocyte and fibroblast wound healing.

    Science.gov (United States)

    Xian, Law Jia; Chowdhury, Shiplu Roy; Bin Saim, Aminuddin; Idrus, Ruszymah Bt Hj

    2015-03-01

    Platelet-rich plasma (PRP) has been found to contain a high concentration of growth factors that are present during the process of healing. Studies conducted found that application of PRP accelerates wound healing. In this study, we characterized the skin cell suspension harvested using the co-isolation technique and evaluated the effects of PRP (10% and 20%, v/v) on co-cultured keratinocytes and fibroblasts in terms of wound healing. Human keratinocytes and fibroblasts were harvested via co-isolation technique and separated via differential trypsinization. These cells were then indirectly co-cultured in medium supplemented with 10% or 20% PRP for 3 days without medium change for analysis of wound-healing potential. The wound-healing potential of keratinocytes and fibroblasts was evaluated in terms of growth property, migratory property, extracellular matrix gene expression and soluble factor secretion. The co-isolation technique yielded a skin cell population dominated by fibroblasts and keratinocytes, with a small amount of melanocytes. Comparison between the 10% and 20% PRP cultures showed that the 10% PRP culture exhibited higher keratinocyte apparent specific growth rate, and secretion of hepatocyte growth factor, monocyte chemoattractant protein-1, epithelial-derived neutrophil-activating protein 78 and vascular endothelial growth factor A, whereas the 20% PRP culture has significantly higher collagen type 1 and collagen type 3 expressions and produced more granulocyte-macrophage colony-stimulating factor. PRP concentration modulates keratinocyte and fibroblast wound healing potential, whereby the 10% PRP promotes wound remodeling, whereas the 20% PRP enhances inflammation and collagen deposition. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  12. Biological effects of plasma rich in growth factors (PRGF) on human endometrial fibroblasts.

    Science.gov (United States)

    Anitua, Eduardo; de la Fuente, María; Ferrando, Marcos; Quintana, Fernando; Larreategui, Zaloa; Matorras, Roberto; Orive, Gorka

    2016-11-01

    To evaluate the biological outcomes of plasma rich in growth factors (PRGF) on human endometrial fibroblasts in culture. PRGF was obtained from three healthy donors and human endometrial fibroblasts (HEF) were isolated from endometrial specimens from five healthy women. The effects of PRGF on cell proliferation and migration, secretion of vascular endothelial growth factor (VEGF), procollagen type I and hyaluronic acid (HA) and contractility of isolated and cultured human endometrial fibroblasts (HEF) were analyzed. Statistical analysis was performed in order to compare the effects of PRGF with respect to control situation (T-test or Mann-Whitney U-test). We report a significantly elevated human endometrial fibroblast proliferation and migration after treatment with PRGF. In addition, stimulation of HEF with PRGF induced an increased expression of the angiogenic factor VEGF and favored the endometrial matrix remodeling by the secretion of procollagen type I and HA and endometrial regeneration by elevating the contractility of HEF. These results were obtained for all PRGF donors and each endometrial cell line. The myriad of growth factors contained in PRGF promoted HEF proliferation, migration and synthesis of paracrine molecules apart from increasing their contractility potential. These preliminary results suggest that PRGF improves the biological activity of HEF in vitro, enhancing the regulation of several cellular processes implied in endometrial regeneration. This innovative treatment deserves further investigation for its potential in "in vivo" endometrial development and especially in human embryo implantation. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  13. [Management of Lung Abscess].

    Science.gov (United States)

    Marra, A; Hillejan, L; Ukena, D

    2015-10-01

    A lung abscess is an infectious pulmonary disease characterised by the presence of a pus-filled cavity within the lung parenchyma. The content of an abscess often drains into the airways spontaneously, leading to an air-fluid level visible on chest X-rays and CT scans. Primary lung abscesses occur in patients who are prone to aspiration or in otherwise healthy individuals; secondary lung abscesses typically develop in association with a stenosing lung neoplasm or a systemic disease that compromises immune defences, such as AIDS, or after organ transplantation. The organisms found in abscesses caused by aspiration pneumonia reflect the resident flora of the oropharynx. The most commonly isolated organisms are anaerobic bacteria (Prevotella, Bacteroides, Fusobacterium, Peptostreptococcus) or streptococci; in alcoholics with poor oral hygiene, the spectrum of pathogens includes Staphylococcus aureus, Streptococcus pyogenes and Actinomyces. Chest radiography and computed tomography (CT) are mandatory procedures in the diagnostic algorithm. Standard treatment for a lung abscess consists of systemic antibiotic therapy, which is based on the anticipated or proven bacterial spectrum of the abscess. In most cases, primary abscesses are successfully treated by calculated empiric antibiotic therapy, with an estimated lethality rate of less than 10 %. Secondary abscesses, despite targeted antimicrobial therapy, are associated with a poor prognosis, which depends on the patient's general condition and underlying disease; lethality is as high as 75 %. Negative prognostic factors are old age, severe comorbidities, immunosuppression, bronchial obstruction, and neoplasms. Surgical intervention due to failure of conservative treatment is required in only 10 % of patients, with a success rate of up to 90 % and postoperative mortality rates ranging between 0 and 33 %. Treatment success after endoscopic or percutaneous drainage is achieved in 73 to 100 % of cases, with an

  14. Incidence of isolated nodal failure in non-small cell lung cancer patients included in a prospective study of the value of PET–CT

    International Nuclear Information System (INIS)

    Kolodziejczyk, Milena; Bujko, Krzysztof; Michalski, Wojciech; Kepka, Lucyna

    2012-01-01

    Purpose: Elective nodal irradiation (ENI) is not recommended in PET–CT-based radiotherapy for NSCLC despite a low level of evidence to support such guidelines. The aim of this investigation is to find out whether omitting ENI is safe. Materials and methods: Sixty-seven patients treated within a frame of a previously published prospective trial of the value of PET–CT were included in the analysis. Seventeen (25%) patients received ENI due to higher initial nodal involvement and in the remaining 50 patients (75%) with N0-N1 or single N2 disease ENI was omitted. Isolated nodal failure (INF) was recorded if relapse occurred in the initially uninvolved regional lymph node without previous or simultaneous local recurrence regardless of the status of distant metastases. Results: With a median follow-up of 32 months, the estimated 3-year overall survival was 42%, local progression-free interval was 55%, and distant metastases-free interval was 62%. Three patients developed INF; all had ENI omitted from treatment, giving a final result of three INFs in 50 (6%) patients treated without ENI. In this group of patients, the 3-year cause-specific cumulative incidence of INF was 6.4% (95% confidence interval: 0–17%). Conclusions: The omission of ENI appears to be not as safe as suggested by current recommendations.

  15. Incidence of isolated nodal failure in non-small cell lung cancer patients included in a prospective study of the value of PET-CT.

    Science.gov (United States)

    Kolodziejczyk, Milena; Bujko, Krzysztof; Michalski, Wojciech; Kepka, Lucyna

    2012-07-01

    Elective nodal irradiation (ENI) is not recommended in PET-CT-based radiotherapy for NSCLC despite a low level of evidence to support such guidelines. The aim of this investigation is to find out whether omitting ENI is safe. Sixty-seven patients treated within a frame of a previously published prospective trial of the value of PET-CT were included in the analysis. Seventeen (25%) patients received ENI due to higher initial nodal involvement and in the remaining 50 patients (75%) with N0-N1 or single N2 disease ENI was omitted. Isolated nodal failure (INF) was recorded if relapse occurred in the initially uninvolved regional lymph node without previous or simultaneous local recurrence regardless of the status of distant metastases. With a median follow-up of 32 months, the estimated 3-year overall survival was 42%, local progression-free interval was 55%, and distant metastases-free interval was 62%. Three patients developed INF; all had ENI omitted from treatment, giving a final result of three INFs in 50 (6%) patients treated without ENI. In this group of patients, the 3-year cause-specific cumulative incidence of INF was 6.4% (95% confidence interval: 0-17%). The omission of ENI appears to be not as safe as suggested by current recommendations. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  16. Thy-1 attenuates TNF-alpha-activated gene expression in mouse embryonic fibroblasts via Src family kinase.

    Directory of Open Access Journals (Sweden)

    Bin Shan

    Full Text Available Heterogeneous surface expression of Thy-1 in fibroblasts modulates inflammation and may thereby modulate injury and repair. As a paradigm, patients with idiopathic pulmonary fibrosis, a disease with pathologic features of chronic inflammation, demonstrate an absence of Thy-1 immunoreactivity within areas of fibrotic activity (fibroblast foci in contrast to the predominant Thy-1 expressing fibroblasts in the normal lung. Likewise, Thy-1 deficient mice display more severe lung fibrosis in response to an inflammatory injury than wildtype littermates. We investigated the role of Thy-1 in the response of fibroblasts to the pro-inflammatory cytokine TNF-alpha. Our study demonstrates distinct profiles of TNF-alpha-activated gene expression in Thy-1 positive (Thy-1+ and negative (Thy-1- subsets of mouse embryonic fibroblasts (MEF. TNF-alpha induced a robust activation of MMP-9, ICAM-1, and the IL-8 promoter driven reporter in Thy-1- MEFs, in contrast to only a modest increase in Thy-1+ counterparts. Consistently, ectopic expression of Thy-1 in Thy-1- MEFs significantly attenuated TNF-alpha-activated gene expression. Mechanistically, TNF-alpha activated Src family kinase (SFK only in Thy-1- MEFs. Blockade of SFK activation abrogated TNF-alpha-activated gene expression in Thy-1- MEFs, whereas restoration of SFK activation rescued the TNF-alpha response in Thy-1+ MEFs. Our findings suggest that Thy-1 down-regulates TNF-alpha-activated gene expression via interfering with SFK- and NF-kappaB-mediated transactivation. The current study provides a novel mechanistic insight to the distinct roles of fibroblast Thy-1 subsets in inflammation.

  17. Isolated Amoebic Abscess of Spleen

    Directory of Open Access Journals (Sweden)

    Kaushik M

    2013-04-01

    Full Text Available Amoebic liver abscess is the most common extraintestinal manifestation of amoebiasis. Extrahepatic amoebic abscesses have occasionally been described in the lung, brain, and skin and presumably result from hematogenous spread. Isolated amoebic abscess of spleen has been reported scarcely in literature. We report here a case of isolated amoebic abscess of spleen.

  18. L-Carnosine reduces telomere damage and shortening rate in cultured normal fibroblasts

    International Nuclear Information System (INIS)

    Shao Lan; Li Qinghuan; Tan Zheng

    2004-01-01

    Telomere is the repetitive DNA sequence at the end of chromosomes, which shortens progressively with cell division and limits the replicative potential of normal human somatic cells. L-Carnosine, a naturally occurring dipeptide, has been reported to delay the replicative senescence, and extend the lifespan of cultured human diploid fibroblasts. In this work, we studied the effect of carnosine on the telomeric DNA of cultured human fetal lung fibroblast cells. Cells continuously grown in 20 mM carnosine exhibited a slower telomere shortening rate and extended lifespan in population doublings. When kept in a long-term nonproliferating state, they accumulated much less damages in the telomeric DNA when cultured in the presence of carnosine. We suggest that the reduction in telomere shortening rate and damages in telomeric DNA made an important contribution to the life-extension effect of carnosine

  19. N-cadherin is overexpressed in Crohn's stricture fibroblasts and promotes intestinal fibroblast migration.

    LENUS (Irish Health Repository)

    Burke, John P

    2012-02-01

    BACKGROUND: Intestinal fibroblasts mediate stricture formation in Crohn\\'s disease (CD). Transforming growth factor-beta (TGF-beta) is important in fibroblast activation, while cell attachment and migration is regulated by the adhesion molecule N-cadherin. The aim of this study was to investigate the expression and function of N-cadherin in intestinal fibroblasts in patients with fibrostenosing CD. METHODS: Intestinal fibroblasts were cultured from seromuscular biopsies from patients undergoing resection for terminal ileal fibrostenosing CD (n = 14) or controls patients (n = 8). N-cadherin expression was assessed using Western blot and quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Fibroblasts were stimulated with TGF-beta and selective pathway inhibitors Y27632, PD98050, and LY294002 were used to examine the Rho\\/ROCK, ERK-1\\/2, and Akt signaling pathways, respectively. Cell migration was assessed using a scratch wound assay. N-cadherin was selectively overexpressed using a plasmid. RESULTS: Fibroblasts from fibrostenosing CD express increased constitutive N-cadherin mRNA and protein and exhibit enhanced basal cell migration relative to those from directly adjacent normal bowel. Control fibroblasts treated with TGF-beta induced N-cadherin in a dose-dependent manner which was inhibited by Rho\\/ROCK and Akt pathway modulation. Control fibroblasts exhibited enhanced cell migration in response to treatment with TGF-beta or transfection with an N-cadherin plasmid. CONCLUSIONS: Fibroblasts from strictures in CD express increased constitutive N-cadherin and exhibit enhanced basal cell migration. TGF-beta is a potent inducer of N-cadherin in intestinal fibroblasts resulting in enhanced cell migration. The TGF-beta-mediated induction of N-cadherin may potentiate Crohn\\'s stricture formation.

  20. Mitochondrial DNA levels in Huntington disease leukocytes and dermal fibroblasts.

    Science.gov (United States)

    Jędrak, Paulina; Krygier, Magdalena; Tońska, Katarzyna; Drozd, Małgorzata; Kaliszewska, Magdalena; Bartnik, Ewa; Sołtan, Witold; Sitek, Emilia J; Stanisławska-Sachadyn, Anna; Limon, Janusz; Sławek, Jarosław; Węgrzyn, Grzegorz; Barańska, Sylwia

    2017-08-01

    Huntington disease (HD) is an inherited neurodegenerative disorder caused by mutations in the huntingtin gene. Involvement of mitochondrial dysfunctions in, and especially influence of the level of mitochondrial DNA (mtDNA) on, development of this disease is unclear. Here, samples of blood from 84 HD patients and 79 controls, and dermal fibroblasts from 10 HD patients and 9 controls were analysed for mtDNA levels. Although the type of mitochondrial haplogroup had no influence on the mtDNA level, and there was no correlation between mtDNA level in leukocytes in HD patients and various parameters of HD severity, some considerable differences between HD patients and controls were identified. The average mtDNA/nDNA relative copy number was significantly higher in leukocytes, but lower in fibroblasts, of symptomatic HD patients relative to the control group. Moreover, HD women displayed higher mtDNA levels in leukocytes than HD men. Because this is the largest population analysed to date, these results might contribute to explanation of discrepancies between previously published studies concerning levels of mtDNA in cells of HD patients. We suggest that the size of the investigated population and type of cells from which DNA is isolated could significantly affect results of mtDNA copy number estimation in HD. Hence, these parameters should be taken into consideration in studies on mtDNA in HD, and perhaps also in other diseases where mitochondrial dysfunction occurs.

  1. Lung abscess

    International Nuclear Information System (INIS)

    Ha, H.K.; Kang, M.W.; Park, J.M.; Yang, W.J.; Shinn, K.S.; Bahk, Y.W.

    1993-01-01

    Lung abscess was successfully treated with percutaneous drainage in 5 of 6 patients. Complete abscess resolution occurred in 4 patients, partial resolution in one, and no response in one. The duration of drainage ranged from 7 to 18 days (mean 15.5 days) in successful cases. The failure of drainage in one neurologicall impaired patient was attributed to persistent aspiration. In 2 patients, concurrent pleural empyema was also cured. CT provided the anatomic details necessary for choosing the puncture site and avoiding puncture of the lung parenchyma. Percutaneous catheter drainage is a safe and effective method for treating lung abscess. (orig.)

  2. Glutamine deprivation induces interleukin-8 expression in ataxia telangiectasia fibroblasts.

    Science.gov (United States)

    Kim, Min-Hyun; Kim, Aryung; Yu, Ji Hoon; Lim, Joo Weon; Kim, Hyeyoung

    2014-05-01

    To investigate whether glutamine deprivation induces expression of inflammatory cytokine interleukin-8 (IL-8) by determining NF-κB activity and levels of oxidative indices (ROS, reactive oxygen species; hydrogen peroxide; GSH, glutathione) in fibroblasts isolated from patients with ataxia telangiectasia (A-T). We used A-T fibroblasts stably transfected with empty vector (Mock) or with human full-length ataxia telangiectasia mutated (ATM) cDNA (YZ5) and mouse embryonic fibroblasts (MEFs) transiently transfected with ATM small interfering RNA (siRNA) or with non-specific control siRNA. The cells were cultured with or without glutamine or GSH. ROS levels were determined using a fluorescence reader and confocal microscopy. IL-8 or murine IL-8 homolog, keratinocyte chemoattractant (KC), and hydrogen peroxide levels in the medium were determined by enzyme-linked immunosorbent assay and colorimetric assay. GSH level was assessed by enzymatic assay, while IL-8 (KC) mRNA level was measured by reverse transcription-polymerase chain reaction (RT-PCR) and/or quantitative real-time PCR. NF-κB DNA-binding activity was determined by electrophoretic mobility shift assay. Catalase activity and ATM protein levels were determined by O2 generation and Western blotting. While glutamine deprivation induced IL-8 expression and increased NF-κB DNA-binding activity in Mock cells, both processes were decreased by treatment of cells with glutamine or GSH or both glutamine and GSH. Glutamine deprivation had no effect on IL-8 expression or NF-κB DNA-binding activity in YZ5 cells. Glutamine-deprived Mock cells had higher oxidative stress indices (increases in ROS and hydrogen peroxide, reduction in GSH) than glutamine-deprived YZ5 cells. In Mock cells, glutamine deprivation-induced oxidative stress indices were suppressed by treatment with glutamine or GSH or both glutamine and GSH. GSH levels and catalase activity were lower in Mock cells than YZ5 cells. MEFs transfected with ATM siRNA and

  3. Culture conditions for bovine embryonic stem cell-like cells isolated from blastocysts after external fertilization

    OpenAIRE

    Jin, Muzi; Wu, Asga; Dorzhin, Sergei; Yue, Qunhua; Ma, Yuzhen; Liu, Dongjun

    2012-01-01

    Although isolation and characterization of embryonic stem cells have been successful in cattle, maintenance of bovine embryonic stem cells in culture remains difficult. In this study, we compared different methods of cell passaging, feeder cell layers and medium conditions for bovine embryonic stem cell-like cells. We found that a murine embryonic fibroblast feeder layer is more suitable for embryonic stem cell-like cells than bovine embryonic fibroblasts. When murine embryonic fibroblasts we...

  4. Fibroblast growth factor-mediated proliferation of central nervous system precursors depends on endogenous production of insulin-like growth factor I

    International Nuclear Information System (INIS)

    Drago, J.; Murphy, M.; Carroll, S.M.; Harvey, R.P.; Bartlett, P.F.

    1991-01-01

    Fibroblast growth factor stimulates proliferation and subsequent differentiation of precursor cells isolated from the neuroepithelium of embryonic day 10 mice in vitro. Here we show that fibroblast growth factor-induced proliferation is dependent on the presence of insulin-like growth factors (IGFs) and that IGF-I is endogenously produced by the neuroepithelial cells. Blocking of endogenous IGF-I activity with anti-IGF-I antibodies results in complete inhibition of fibroblast growth factor-mediated proliferation and in cell death. IGF-I alone acts as a survival agent. These observations correlate with the detection of transcripts for IGF-I and basic fibroblast growth factor in freshly isolated neuroepithelium and are consistent with an autocrine action of these factors in early brain development in vivo

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

    Science.gov (United States)

    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.

  6. Lung cancer

    Science.gov (United States)

    ... causing chemicals such as uranium, beryllium, vinyl chloride, nickel chromates, coal products, mustard gas, chloromethyl ethers, gasoline, and diesel exhaust Exposure to radon gas Family history of lung cancer ...

  7. Lung surgery

    Science.gov (United States)

    ... are thoracotomy and video-assisted thoracoscopic surgery (VATS). Robotic surgery may also be used. Lung surgery using ... Center-Shreveport, Shreveport, LA. Review provided by VeriMed Healthcare Network. Also reviewed by David Zieve, MD, MHA, ...

  8. Inhibition of EGF processing in responsive and nonresponsive human fibroblasts

    International Nuclear Information System (INIS)

    Schaudies, R.P.; Wray, H.L.

    1988-01-01

    We have examined the proteolytic processing of radiolabeled epidermal growth factor (EGF) in EGF growth-responsive human foreskin fibroblasts (HFF) versus EGF nonresponsive human fetal lung fibroblasts (HFL). Previous studies have shown that both cell lines demonstrate similar binding affinities and numbers of binding sites, as well as similar rates of internalization and degradation of the bound, radiolabeled hormone. We have used nondenaturing electrophoresis to compare how these two cell lines process EGF at its carboxy terminus. EGF lacking either one [des-(53)-EGF] or six [des (48-53)-EGF] carboxy terminal amino acids could be distinguished by this method. Chloroquine or leupeptin were added to the incubation system in an attempt to accentuate potential differences in hormonal processing between the responsive and nonresponsive cell lines. In the absence of inhibitors, the responsive and nonresponsive cells generated similar distributions of processed forms of EGF after 30-minutes incubation. However, after 4-hours incubation in the constant presence of 125I-EGF, the electrophoretic profiles of extracted hormone were substantially different. The radiolabel within the responsive cells, as well as that released from them, migrated predominantly at the dye front, indicating complete degradation of EGF. In contrast, the majority of the radiolabel within the nonresponsive cells migrated as partially processed forms of hormone, while the released radiolabel migrated at the dye front. Addition of chloroquine to either cell line inhibited processing of EGF beyond removal of the carboxyl terminal arginine residue. Both intact 125I-EGF, and 125I-EGF lacking the carboxyl terminal arginine were released from chloroquine-treated cells in a ratio equal to that present in the intact cells

  9. Unexpandable lung.

    Science.gov (United States)

    Pereyra, Marco F; Ferreiro, Lucía; Valdés, Luis

    2013-02-01

    Unexpandable lung is a mechanical complication by which the lung does not expand to the chest wall, impeding a normal apposition between the two pleural layers. The main mechanism involved is the restriction of the visceral pleura due to the formation of a fibrous layer along this pleural membrane. This happens because of the presence of an active pleural disease (lung entrapment), which can be resolved if proper therapeutic measures are taken, or a remote disease (trapped lung), in which an irreversible fibrous pleural layer has been formed. The clinical suspicion arises with the presence of post-thoracocentesis hydropneumothorax or a pleural effusion that cannot be drained due to the appearance of thoracic pain. The diagnosis is based on the analysis of the pleural liquid, the determination of pleural pressures as we drain the effusion and on air-contrast chest CT. As both represent the continuity of one same process, the results will depend on the time at which these procedures are done. If, when given a lung that is becoming entrapped, the necessary therapeutic measures are not taken, the final result will be a trapped lung. In this instance, most patients are asymptomatic or have mild exertional dyspnea and therefore they do not require treatment. Nevertheless, in cases of incapacitating dyspnea, it may be necessary to use pleural decortication in order to resolve the symptoms. Copyright © 2012 SEPAR. Published by Elsevier Espana. All rights reserved.

  10. Age-associated intracellular superoxide dismutase deficiency potentiates dermal fibroblast dysfunction during wound healing.

    Science.gov (United States)

    Fujiwara, Toshihiro; Dohi, Teruyuki; Maan, Zeshaan N; Rustad, Kristine C; Kwon, Sun Hyung; Padmanabhan, Jagannath; Whittam, Alexander J; Suga, Hirotaka; Duscher, Dominik; Rodrigues, Melanie; Gurtner, Geoffrey C

    2017-07-04

    Reactive oxygen species (ROS) impair wound healing through destructive oxidation of intracellular proteins, lipids and nucleic acids. Intracellular superoxide dismutase (SOD1) regulates ROS levels and plays a critical role in tissue homoeostasis. Recent evidence suggests that age-associated wound healing impairments may partially result from decreased SOD1 expression. We investigated the mechanistic basis by which increased oxidative stress links to age-associated impaired wound healing. Fibroblasts were isolated from unwounded skin of young and aged mice, and myofibroblast differentiation was assessed by measuring α-smooth muscle actin and collagen gel contraction. Excisional wounds were created on young and aged mice to study the healing rate, ROS levels and SOD1 expression. A mechanistic link between oxidative stress and fibroblast function was explored by assessing the TGF-β1 signalling pathway components in young and aged mice. Age-related wounds displayed reduced myofibroblast differentiation and delayed wound healing, consistent with a decrease in the in vitro capacity for fibroblast-myofibroblast transition following oxidative stress. Young fibroblasts with normal SOD1 expression exhibited increased phosphorylation of ERK in response to elevated ROS. In contrast, aged fibroblasts with reduced SOD1 expression displayed a reduced capacity to modulate intracellular ROS. Collectively, age-associated wound healing impairments are associated with fibroblast dysfunction that is likely the result of decreased SOD1 expression and subsequent dysregulation of intracellular ROS. Strategies targeting these mechanisms may suggest a new therapeutic approach in the treatment of chronic non-healing wounds in the aged population. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. Scleroderma keratinocytes promote fibroblast activation independent of transforming growth factor beta.

    Science.gov (United States)

    McCoy, Sara S; Reed, Tamra J; Berthier, Celine C; Tsou, Pei-Suen; Liu, Jianhua; Gudjonsson, Johann E; Khanna, Dinesh; Kahlenberg, J Michelle

    2017-11-01

    SSc is a devastating disease that results in fibrosis of the skin and other organs. Fibroblasts are a key driver of the fibrotic process through deposition of extracellular matrix. The mechanisms by which fibroblasts are induced to become pro-fibrotic remain unclear. Thus, we examined the ability of SSc keratinocytes to promote fibroblast activation and the source of this effect. Keratinocytes were isolated from skin biopsies of 9 lcSSc, 10 dcSSc and 13 control patients. Conditioned media was saved from the cultures. Normal fresh primary fibroblasts were exposed to healthy control and SSc keratinocyte conditioned media in the presence or absence of neutralizing antibodies for TGF-β. Gene expression was assessed by microarrays and real-time PCR. Immunocytochemistry was performed for α-smooth muscle actin (α-SMA), collagen type 1 (COL1A1) and CCL5 expression. SSc keratinocyte conditioned media promoted fibroblast activation, characterized by increased α-SMA and COL1A1 mRNA and protein expression. This effect was independent of TGF-β. Microarray analysis identified upregulation of nuclear factor κB (NF-κB) and downregulation of peroxisome proliferator-activated receptor γ (PPAR-γ) pathways in both SSc subtypes. Scleroderma keratinocytes exhibited increased expression of NF-κB-regulated cytokines and chemokines and lesional skin staining confirmed upregulation of CCL5 in basal keratinocytes. Scleroderma keratinocytes promote the activation of fibroblasts in a TGF-β-independent manner and demonstrate an imbalance in NF-κB1 and PPAR-γ expression leading to increased cytokine and CCL5 production. Further study of keratinocyte mediators of fibrosis, including CCL5, may provide novel targets for skin fibrosis therapy. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  12. New isomalabaricane triterpenes from the marine sponge Stelletta globostellata that induce morphological changes in rat fibroblasts.

    Science.gov (United States)

    Oku, N; Matsunaga, S; Wada, S i; Watabe, S; Fusetani, N

    2000-02-01

    Three new isomalabaricane triterpenes, 29-hydroxystelliferin D (2), 3-epi-29-hydroxystelliferin E (3), and 3-epi-29-hydroxystelliferin A (4), were isolated from the marine sponge Stelletta globostellata. Their structures, including absolute stereochemistry, were determined on the basis of spectral data and chemical methods. Rat fibroblasts treated with 0.2 microM of 2-4 exhibited unusual morphological characteristics, followed by death in 5 days.

  13. Testosterone metabolism of fibroblasts grown from prostatic carcinoma, benign prostatic hyperplasia and skin fibroblasts

    International Nuclear Information System (INIS)

    Schweikert, H.U.; Hein, H.J.; Romijn, J.C.; Schroeder, F.H.

    1982-01-01

    The metabolism of [1,2,6,7-3H]testosterone was assessed in fibroblast monolayers derived from tissue of 5 prostates with benign hyperplasia (BPH), 4 prostates with carcinoma (PC), and 3 biopsy samples of skin, 2 nongenital skin (NG) and 1 genital skin. The following metabolites could be identified: androstanedione androstenedione, dihydrotestosterone, androsterone, epiandrosterone, androstane-3 alpha, 17 beta-diol and androstane-3 beta, 17 beta-diol. Testosterone was metabolized much more rapidly in fibroblasts originating from prostatic tissue than in fibroblasts derived from NG. A significantly higher formation of 5 alpha-androstanes and 3 alpha-hydroxysteroids could be observed in fibroblasts from BPH as compared to PC. 17-ketosteroid formation exceeded 5 alpha-androstane formation in BPH, whereas 5 alpha-reduction was the predominant pathway in fibroblasts grown from PC and NG. Since testosterone metabolism in fibroblasts of prostatic origin therefore resembles in many aspects that in whole prostatic tissue, fibroblasts grown from prostatic tissues might be a valuable tool for further investigation of the pathogenesis of human BPH and PC

  14. Hedgehog signaling contributes to basic fibroblast growth factor-regulated fibroblast migration

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Zhong Xin [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Sun, Cong Cong [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Wenzhou People' s Hospital, Wenzhou, Zhejiang (China); Ting Zhu, Yu; Wang, Ying; Wang, Tao; Chi, Li Sha; Cai, Wan Hui [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Zheng, Jia Yong [Wenzhou People' s Hospital, Wenzhou, Zhejiang (China); Zhou, Xuan [Ningbo First Hospital, Ningbo, Zhejiang (China); Cong, Wei Tao [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Li, Xiao Kun, E-mail: proflxk@163.com [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Jin, Li Tai, E-mail: jin_litai@126.com [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China)

    2017-06-15

    Fibroblast migration is a central process in skin wound healing, which requires the coordination of several types of growth factors. bFGF, a well-known fibroblast growth factor (FGF), is able to accelerate fibroblast migration; however, the underlying mechanism of bFGF regulation fibroblast migration remains unclear. Through the RNA-seq analysis, we had identified that the hedgehog (Hh) canonical pathway genes including Smoothened (Smo) and Gli1, were regulated by bFGF. Further analysis revealed that activation of the Hh pathway via up-regulation of Smo promoted fibroblast migration, invasion, and skin wound healing, but which significantly reduced by GANT61, a selective antagonist of Gli1/Gli2. Western blot analyses and siRNA transfection assays demonstrated that Smo acted upstream of phosphoinositide 3-kinase (PI3K)-c-Jun N-terminal kinase (JNK)-β-catenin to promote cell migration. Moreover, RNA-seq and qRT-PCR analyses revealed that Hh pathway genes including Smo and Gli1 were under control of β-catenin, suggesting that β-catenin turn feedback activates Hh signaling. Taken together, our analyses identified a new bFGF-regulating mechanism by which Hh signaling regulates human fibroblast migration, and the data presented here opens a new avenue for the wound healing therapy. - Highlights: • bFGF regulates Hedgehog (Hh) signaling in fibroblasts. • The Smo and Gli two master regulators of Hh signaling positively regulate fibroblast migration. • Smo facilitates β-catenin nuclear translocation via activation PI3K/JNK/GSK3β. • β-catenin positively regulates fibroblast cell migration and the expression of Hh signaling genes including Smo and Gli.

  15. Hedgehog signaling contributes to basic fibroblast growth factor-regulated fibroblast migration

    International Nuclear Information System (INIS)

    Zhu, Zhong Xin; Sun, Cong Cong; Ting Zhu, Yu; Wang, Ying; Wang, Tao; Chi, Li Sha; Cai, Wan Hui; Zheng, Jia Yong; Zhou, Xuan; Cong, Wei Tao; Li, Xiao Kun; Jin, Li Tai

    2017-01-01

    Fibroblast migration is a central process in skin wound healing, which requires the coordination of several types of growth factors. bFGF, a well-known fibroblast growth factor (FGF), is able to accelerate fibroblast migration; however, the underlying mechanism of bFGF regulation fibroblast migration remains unclear. Through the RNA-seq analysis, we had identified that the hedgehog (Hh) canonical pathway genes including Smoothened (Smo) and Gli1, were regulated by bFGF. Further analysis revealed that activation of the Hh pathway via up-regulation of Smo promoted fibroblast migration, invasion, and skin wound healing, but which significantly reduced by GANT61, a selective antagonist of Gli1/Gli2. Western blot analyses and siRNA transfection assays demonstrated that Smo acted upstream of phosphoinositide 3-kinase (PI3K)-c-Jun N-terminal kinase (JNK)-β-catenin to promote cell migration. Moreover, RNA-seq and qRT-PCR analyses revealed that Hh pathway genes including Smo and Gli1 were under control of β-catenin, suggesting that β-catenin turn feedback activates Hh signaling. Taken together, our analyses identified a new bFGF-regulating mechanism by which Hh signaling regulates human fibroblast migration, and the data presented here opens a new avenue for the wound healing therapy. - Highlights: • bFGF regulates Hedgehog (Hh) signaling in fibroblasts. • The Smo and Gli two master regulators of Hh signaling positively regulate fibroblast migration. • Smo facilitates β-catenin nuclear translocation via activation PI3K/JNK/GSK3β. • β-catenin positively regulates fibroblast cell migration and the expression of Hh signaling genes including Smo and Gli.

  16. Deposition of insoluble elastin by pulmonary fibroblasts from patients with COPD is increased by treatment with versican siRNA.

    Science.gov (United States)

    Wu, Lian; Zhang, Jing; Qu, Jie Ming; Bai, Chun-Xue; Merrilees, Mervyn J

    2017-01-01

    A reduced content of alveolar elastic fibers is a key feature of COPD lung. Despite continued elastogenic potential by alveolar fibroblasts in the lung affected by COPD, repair of elastic fibers does not take place, which is due to increased levels of the chondroitin sulfate proteoglycan versican that inhibits the assembly of tropoelastin into fibers. In this study, primary pulmonary fibroblast cell lines from COPD and non-COPD patients were treated with a small interfering RNA (siRNA) against versican to determine if knockdown of versican could restore the deposition of insoluble elastin. Versican siRNA treatment reduced versican expression and secretion by pulmonary fibroblasts from both COPD and non-COPD patients ( P elastin in the COPD cell cultures ( P elastin (tropoelastin) in either the COPD or non-COPD cell cultures, supporting a role for versican in inhibiting assembly but not synthesis of tropoelastin. These results suggest that removal or knockdown of versican may be a possible therapeutic strategy for increasing deposition of insoluble elastin and stimulating repair of elastic fibers in COPD lung.

  17. A fibroblast-associated antigen: Characterization in fibroblasts and immunoreactivity in smooth muscle differentiated stromal cells

    DEFF Research Database (Denmark)

    Rønnov-Jessen, Lone; Celis, Julio E.; van Deurs, Bo

    1992-01-01

    major brands migrating at apparent Mr of 38,000, 45,000, and 80,000, in addition to many minor bands between Mr 45,000 and 97,000, including Mr 52,000. The Mr 45,000 and 38,000 were associated with the cell membrane and Mr 52,000 as well as Mr 38,000 were associated with the lysosomes. The 1B10......Fibroblasts with smooth muscle differentiation are frequently derived from human breast tissue. Immunofluorescence cytochemistry of a fibroblast-associated antigen recognized by a monoclonal antibody (MAb), 1B10, was analyzed with a view to discriminating smooth muscle differentiated fibroblasts...

  18. Umbilical cord fibroblasts: Could they be considered as mesenchymal stem cells?

    Science.gov (United States)

    Zeddou, Mustapha; Relic, Biserka; Malaise, Michel G

    2014-01-01

    In cell therapy protocols, many tissues were proposed as a source of mesenchymal stem cells (MSC) isolation. So far, bone marrow (BM) has been presented as the main source of MSC despite the invasive isolation procedure related to this source. During the last years, the umbilical cord (UC) matrix was cited in different studies as a reliable source from which long term ex vivo proliferating fibroblasts were isolated but with contradictory data about their immunophenotype, gene expression profile, and differentiation potential. Hence, an interesting question emerged: Are cells isolated from cord matrix (UC-MSC) different from other MSCs? In this review, we will summarize different studies that isolated and characterized UC-MSC. Considering BM-MSC as gold standard, we will discuss if UC-MSC fulfill different criteria that define MSC, and what remain to be done in this issue. PMID:25126385

  19. Role of human pulmonary fibroblast-derived MCP-1 in cell activation and migration in experimental silicosis

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xueting [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Fang, Shencun [Nine Department of Respiratory Medicine, Nanjing Chest Hospital, Nanjing, Jiangsu 210029 (China); Liu, Haijun [Neurobiology Laboratory, New Drug Screening Centre, China Pharmaceutical University, Nanjing, Jiangsu 210009 (China); Wang, Xingang; Dai, Xiaoniu; Yin, Qing; Yun, Tianwei [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Wang, Wei; Zhang, Yingming [Nine Department of Respiratory Medicine, Nanjing Chest Hospital, Nanjing, Jiangsu 210029 (China); Liao, Hong [Neurobiology Laboratory, New Drug Screening Centre, China Pharmaceutical University, Nanjing, Jiangsu 210009 (China); Zhang, Wei [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Yao, Honghong [Department of Pharmacology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China); Chao, Jie, E-mail: chaojie@seu.edu.cn [Department of Physiology, Medical School of Southeast University, Nanjing, Jiangsu 210009 (China)

    2015-10-15

    Background: Silicosis is a systemic disease caused by inhaling silicon dioxide (SiO{sub 2}). Phagocytosis of SiO{sub 2} in the lung initiates an inflammatory cascade that results in fibroblast proliferation and migration and subsequent fibrosis. Clinical evidence indicates that the activation of alveolar macrophages by SiO{sub 2} produces rapid and sustained inflammation that is characterized by the generation of monocyte chemotactic protein 1 (MCP-1), which induces fibrosis. Pulmonary fibroblast-derived MCP-1 may play a critical role in fibroblast proliferation and migration. Methods and results: Experiments using primary cultured adult human pulmonary fibroblasts (HPF-a) demonstrated the following results: 1) SiO{sub 2} treatment resulted in the rapid and sustained induction of MCP-1 as well as the elevation of the CC chemokine receptor type 2 (CCR2) protein levels; 2) pretreatment of HPF-a with RS-102895, a specific CCR2 inhibitor, abolished the SiO{sub 2}-induced increase in cell activation and migration in both 2D and 3D culture systems; and 3) RNA interference targeting CCR2 prevented the SiO{sub 2}-induced increase in cell migration. Conclusion: These data demonstrated that the up-regulation of pulmonary fibroblast-derived MCP-1 is involved in pulmonary fibroblast migration induced by SiO{sub 2}. CCR2 was also up-regulated in response to SiO{sub 2}, and this up-regulation facilitated the effect of MCP-1 on fibroblasts. Our study deciphered the link between fibroblast-derived MCP-1 and SiO{sub 2}-induced cell migration. This finding provides novel insight into the potential of MCP-1 in the development of novel therapeutic strategies for silicosis. - Highlights: • Role of pulmonary fibroblast-derived MCP-1 in experimental silicosis was studied. • SiO{sub 2} induced MCP-1 release from cultured human pulmonary fibroblast (HPF-a). • SiO{sub 2} directly activated HPF-a via the MCP-1/CCR2 pathway. • SiO{sub 2} increased HPF-a migration in both 2D and 3D

  20. Role of human pulmonary fibroblast-derived MCP-1 in cell activation and migration in experimental silicosis

    International Nuclear Information System (INIS)

    Liu, Xueting; Fang, Shencun; Liu, Haijun; Wang, Xingang; Dai, Xiaoniu; Yin, Qing; Yun, Tianwei; Wang, Wei; Zhang, Yingming; Liao, Hong; Zhang, Wei; Yao, Honghong; Chao, Jie

    2015-01-01

    Background: Silicosis is a systemic disease caused by inhaling silicon dioxide (SiO 2 ). Phagocytosis of SiO 2 in the lung initiates an inflammatory cascade that results in fibroblast proliferation and migration and subsequent fibrosis. Clinical evidence indicates that the activation of alveolar macrophages by SiO 2 produces rapid and sustained inflammation that is characterized by the generation of monocyte chemotactic protein 1 (MCP-1), which induces fibrosis. Pulmonary fibroblast-derived MCP-1 may play a critical role in fibroblast proliferation and migration. Methods and results: Experiments using primary cultured adult human pulmonary fibroblasts (HPF-a) demonstrated the following results: 1) SiO 2 treatment resulted in the rapid and sustained induction of MCP-1 as well as the elevation of the CC chemokine receptor type 2 (CCR2) protein levels; 2) pretreatment of HPF-a with RS-102895, a specific CCR2 inhibitor, abolished the SiO 2 -induced increase in cell activation and migration in both 2D and 3D culture systems; and 3) RNA interference targeting CCR2 prevented the SiO 2 -induced increase in cell migration. Conclusion: These data demonstrated that the up-regulation of pulmonary fibroblast-derived MCP-1 is involved in pulmonary fibroblast migration induced by SiO 2 . CCR2 was also up-regulated in response to SiO 2 , and this up-regulation facilitated the effect of MCP-1 on fibroblasts. Our study deciphered the link between fibroblast-derived MCP-1 and SiO 2 -induced cell migration. This finding provides novel insight into the potential of MCP-1 in the development of novel therapeutic strategies for silicosis. - Highlights: • Role of pulmonary fibroblast-derived MCP-1 in experimental silicosis was studied. • SiO 2 induced MCP-1 release from cultured human pulmonary fibroblast (HPF-a). • SiO 2 directly activated HPF-a via the MCP-1/CCR2 pathway. • SiO 2 increased HPF-a migration in both 2D and 3D model via the MCP-1/CCR2 pathway. • RNA-i of MCP-1/CCR2

  1. Role of periodontal ligament fibroblasts in osteoclastogenesis: a review

    NARCIS (Netherlands)

    Sokos, D.; Everts, V.; de Vries, T.J.

    2015-01-01

    During the last decade it has become clear that periodontal ligament fibroblasts may contribute to the in vitro differentiation of osteoclasts. We surveyed the current findings regarding their osteoclastogenesis potential. Periodontal ligament fibroblasts have the capacity to select and attract

  2. Lung function

    International Nuclear Information System (INIS)

    Sorichter, S.

    2009-01-01

    The term lung function is often restricted to the assessment of volume time curves measured at the mouth. Spirometry includes the assessment of lung volumes which can be mobilised with the corresponding flow-volume curves. In addition, lung volumes that can not be mobilised, such as the residual volume, or only partially as FRC and TLC can be measured by body plethysmography combined with the determination of the airway resistance. Body plethysmography allows the correct positioning of forced breathing manoeuvres on the volume-axis, e.g. before and after pharmacotherapy. Adding the CO single breath transfer factor (T LCO ), which includes the measurement of the ventilated lung volume using He, enables a clear diagnosis of different obstructive, restrictive or mixed ventilatory defects with and without trapped air. Tests of reversibility and provocation, as well as the assessment of inspiratory mouth pressures (PI max , P 0.1 ) help to classify the underlying disorder and to clarify treatment strategies. For further information and to complete the diagnostic of disturbances of the ventilation, diffusion and/or perfusion (capillar-)arterial bloodgases at rest and under physical strain sometimes amended by ergospirometry are recommended. Ideally, lung function measurements are amended by radiological and nuclear medicine techniques. (orig.) [de

  3. Transformation and scattering activities of the receptor tyrosine kinase RON/Stk in rodent fibroblasts and lack of regulation by the jaagsiekte sheep retrovirus receptor, Hyal2

    International Nuclear Information System (INIS)

    Miller, A Dusty; Van Hoeven, Neal S; Liu, Shan-Lu

    2004-01-01

    The envelope (Env) protein of jaagsiekte sheep retrovirus (JSRV) can transform cells in culture and is likely to be the main factor responsible for lung cancer induction by JSRV in animals. A recent report indicates that the epithelial-cell transforming activity of JSRV Env depends on activation of the cell-surface receptor tyrosine kinase Mst1r (called RON for the human and Stk for the rodent orthologs). In the immortalized line of human epithelial cells used (BEAS-2B cells), the virus receptor Hyal2 was found to bind to and suppress the activity of RON. When Env was expressed it bound to Hyal2 causing its degradation, release of RON activity from Hyal2 suppression, and activation of pathways resulting in cell transformation. Due to difficulty with reproducibility of the transformation assay in BEAS-2B cells, we have used more tractable rodent fibroblast models to further study Hyal2 modulation of RON/Stk transforming activity and potential effects of Hyal2 on RON/Stk activation by its natural ligand, macrophage stimulating protein (MSP). We did not detect transformation of NIH 3T3 cells by plasmids expressing RON or Stk, but did detect transformation of 208F rat fibroblasts by these plasmids at a very low rate. We were able to isolate 208F cell clones that expressed RON or Stk and that showed changes in morphology indicative of transformation. The parental 208F cells did not respond to MSP but 208F cells expressing RON or Stk showed obvious increases in scattering/transformation in response to MSP. Human Hyal2 had no effect on the basal or MSP-induced phenotypes of RON-expressing 208F cells, and human, mouse or rat Hyal2 had no effect on the basal or MSP-induced phenotypes of Stk-expressing 208F cells. We have shown that RON or Stk expression in 208F rat fibroblasts results in a transformed phenotype that is enhanced by addition of the natural ligand for these proteins, MSP. Hyal2 does not directly modulate the basal or MSP-induced RON/Stk activity, although it

  4. Blockade of lysophosphatidic acid receptors LPAR1/3 ameliorates lung fibrosis induced by irradiation

    International Nuclear Information System (INIS)

    Gan, Lu; Xue, Jian-Xin; Li, Xin; Liu, De-Song; Ge, Yan; Ni, Pei-Yan; Deng, Lin; Lu, You; Jiang, Wei

    2011-01-01

    Highlights: → Lysophosphatidic acid (LPA) levels and its receptors LPAR1/3 transcripts were elevated during the development of radiation-induced lung fibrosis. → Lung fibrosis was obviously alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. → VPC12249 administration effectively inhibited radiation-induced fibroblast accumulation in vivo, and suppressed LPA-induced fibroblast proliferation in vitro. → LPA-LPAR1/3 signaling regulated TGFβ1 and CTGF expressions in radiation-challenged lungs, but only influenced CTGF expression in cultured fibroblasts. → LPA-LPAR1/3 signaling induced fibroblast proliferation through a CTGF-dependent pathway, rather than through TGFβ1 activation. -- Abstract: Lung fibrosis is a common and serious complication of radiation therapy for lung cancer, for which there are no efficient treatments. Emerging evidence indicates that lysophosphatidic acid (LPA) and its receptors (LPARs) are involved in the pathogenesis of fibrosis. Here, we reported that thoracic radiation with 16 Gy in mice induced development of radiation lung fibrosis (RLF) accompanied by obvious increases in LPA release and LPAR1 and LPAR3 (LPAR1/3) transcripts. RLF was significantly alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. VPC12249 administration effectively prolonged animal survival, restored lung structure, inhibited fibroblast accumulation and reduced collagen deposition. Moreover, profibrotic cytokines in radiation-challenged lungs obviously decreased following administration of VPC12249, including transforming growth factor β1 (TGFβ1) and connective tissue growth factor (CTGF). In vitro, LPA induced both fibroblast proliferation and CTGF expression in a dose-dependent manner, and both were suppressed by blockade of LPAR1/3. The pro-proliferative activity of LPA on fibroblasts was inhibited by siRNA directed against CTGF. Together, our data suggest that the LPA-LPAR1/3 signaling system is involved in the

  5. Blockade of lysophosphatidic acid receptors LPAR1/3 ameliorates lung fibrosis induced by irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Gan, Lu [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Xue, Jian-Xin [Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Laboratory of Stem Cell Biology, West China Hospital, Sichuan University, Chengdu (China); Li, Xin [Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Liu, De-Song [Department of Pediatrics, Sichuan Provincial Hospital of Women and Children, Chengdu (China); Ge, Yan; Ni, Pei-Yan; Deng, Lin [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Lu, You, E-mail: radyoulu@hotmail.com [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Jiang, Wei, E-mail: wcumsjw72@hotmail.com [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Molecular Medicine Research Center, West China Hospital, Sichuan University, Chengdu (China)

    2011-05-27

    Highlights: {yields} Lysophosphatidic acid (LPA) levels and its receptors LPAR1/3 transcripts were elevated during the development of radiation-induced lung fibrosis. {yields} Lung fibrosis was obviously alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. {yields} VPC12249 administration effectively inhibited radiation-induced fibroblast accumulation in vivo, and suppressed LPA-induced fibroblast proliferation in vitro. {yields} LPA-LPAR1/3 signaling regulated TGF{beta}1 and CTGF expressions in radiation-challenged lungs, but only influenced CTGF expression in cultured fibroblasts. {yields} LPA-LPAR1/3 signaling induced fibroblast proliferation through a CTGF-dependent pathway, rather than through TGF{beta}1 activation. -- Abstract: Lung fibrosis is a common and serious complication of radiation therapy for lung cancer, for which there are no efficient treatments. Emerging evidence indicates that lysophosphatidic acid (LPA) and its receptors (LPARs) are involved in the pathogenesis of fibrosis. Here, we reported that thoracic radiation with 16 Gy in mice induced development of radiation lung fibrosis (RLF) accompanied by obvious increases in LPA release and LPAR1 and LPAR3 (LPAR1/3) transcripts. RLF was significantly alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. VPC12249 administration effectively prolonged animal survival, restored lung structure, inhibited fibroblast accumulation and reduced collagen deposition. Moreover, profibrotic cytokines in radiation-challenged lungs obviously decreased following administration of VPC12249, including transforming growth factor {beta}1 (TGF{beta}1) and connective tissue growth factor (CTGF). In vitro, LPA induced both fibroblast proliferation and CTGF expression in a dose-dependent manner, and both were suppressed by blockade of LPAR1/3. The pro-proliferative activity of LPA on fibroblasts was inhibited by siRNA directed against CTGF. Together, our data suggest that the LPA-LPAR1

  6. Fibroblast growth factor 23 - et fosfatregulerende hormon

    DEFF Research Database (Denmark)

    Beck-Nielsen, Signe; Pedersen, Susanne Møller; Kassem, Moustapha

    2010-01-01

    Fibroblast growth factor 23 (FGF23) er et nyligt identificeret fosfatonin. FGF23's fysiologiske hovedfunktion er at opretholde normalt serumfosfat og at virke som et D-vitaminmodregulatorisk hormon. Sygdomme, der er koblet til forhøjet serum FGF23, er hypofosfatæmisk rakitis, fibrøs dysplasi og t...

  7. Open lung biopsy

    Science.gov (United States)

    Biopsy - open lung ... An open lung biopsy is done in the hospital using general anesthesia . This means you will be asleep and ... The open lung biopsy is done to evaluate lung problems seen on x-ray or CT scan .

  8. Lung Cancer: Glossary

    Science.gov (United States)

    ... professional support team today. Learn More . Find more lung cancer resources. Learn More Donate Today! What is Lung ... to Give How Your Support Helps Events Lung Cancer Awareness © Lung Cancer Alliance. The information presented in this website ...

  9. Lung Cancer Prevention

    Science.gov (United States)

    ... Colorectal Cancer Kidney (Renal Cell) Cancer Leukemia Liver Cancer Lung Cancer Lymphoma Pancreatic Cancer Prostate Cancer Skin Cancer ... following PDQ summaries for more information about lung cancer: Lung Cancer Screening Non-Small Cell Lung Cancer Treatment ...

  10. Lung cancer - small cell

    Science.gov (United States)

    Cancer - lung - small cell; Small cell lung cancer; SCLC ... About 15% of all lung cancer cases are SCLC. Small cell lung cancer is slightly more common in men than women. Almost all cases of SCLC are ...

  11. [Lung scintigraphy].

    Science.gov (United States)

    Schümichen, Carl; Schmidt, Matthias; Krause, Thomas

    2018-06-01

    The S1 guideline for lung scintigraphy has been updated and extended in order to emphasize the advantages oft the method in detecting acute pulmonary embolism (PE) in the periphery oft the lung (subsegmental PE), in underlying subacute and chronic pulmonary disorders, as well as in detecting chronic LE (CTEPH). Method of choice is ventilation / perfusion (V/P) SPECT or V/P SPECT/CT with even higher specificity. Because of its high sensitivity, a threshold (V/P mismatch in at least one segment or two subsegments) is introduced to avoid overtreatment. In case of a change in the therapeutic approach (observation only instead of anticoaculation) the threshold can be omitted. New data concerning the clinical and therapeutical impact of subsegmental PE are included, the chapters open questions have been extented. Other indications for V/P SPECT (secondary diagnoses, abnormalities in pulmonary perfusion, prediction of postoperative lung function) are presented with new data. Schattauer GmbH.

  12. Lung radiopharmaceuticals

    International Nuclear Information System (INIS)

    Gonzalez, B.M.

    1994-01-01

    Indication or main clinical use of Lung radiopharmaceuticals is presented and clasification of radiopharmaceuticals as ventilation and perfusion studies. Perfusion radiopharmaceuticals, main controls for administration quality acceptance. Clearence after blood administration and main clinical applications. Ventilation radiopharmaceuticals, gases and aerosols, characteristics of a ideal radioaerosol, techniques of good inhalation procedure, clinical applications. Comparison of several radiopharmaceuticals reflering to retention time as 50% administered dose, percent administered dose at 6 hours post inhalation, blood activity at 30 and 60 minutes post inhalation, initial lung absorbed dose, cumulated activity.Kinetic description of two radiopharmaceuticals, 99mTcDTPA and 99mTc-PYP

  13. Lung Transplant

    Science.gov (United States)

    ... Severity of the recipient's lung disease Recipient's overall health Likelihood that the transplant will be successful Immediately before ... will begin within days of your surgery. Your health care team will likely work with you to design an exercise program that's right for you. Your doctor may ...

  14. Lung cancer

    DEFF Research Database (Denmark)

    Hansen, H H; Rørth, M

    1999-01-01

    The results of the many clinical trials published in 1997 had only modest impact on the treatment results using either cytostatic agents alone or combined with radiotherapy in lung cancer. In SCLC, combination chemotherapy including platin-compounds (cisplatin, carboplatin) and the podophyllotoxins...

  15. The effect of tranilast on fibroblast activation protein α (FAP-α expression in normal and keloid fibroblasts in vitro

    Directory of Open Access Journals (Sweden)

    Paweł P. Antończak

    2017-07-01

    Full Text Available Introduction . Tranilast (N-(3’,4’-demethoxycinnamoyl-anthranilic acid is an anti-allergic drug. Its mechanism of action is based on the inhibition of antigen-induced release of chemical mediators from mast cells and basophils. It also reveals antifibroproliferative activities. These properties of tranilast are used in the treatment of hypertrophic scars and keloids. Keloids are characterized by incorrect extracellular matrix components turnover. Fibroblasts derived from keloids reveal overproduction of collagen type I and decreased degradation of extracellular matrix in comparison with normal fibroblasts. Fibroblast activation protein α (FAP-α may play an important role in remodeling of extracellular matrix and the invasive properties of keloids. Objective . In the present study, the effect of tranilast on expression of FAP-α gene and its protein was evaluated in normal human dermal fibroblasts and fibroblasts derived from keloids cultured in vitro . Materials and methods. In the first stage of the study, the influence of tranilast on cell viability was estimated. The second stage of the study included the quantitative evaluation of FAP-α mRNA expression in normal and keloid fibroblasts treated with tranilast. The third stage of the study comprised fibroblast activation protein α expression analysis in the examined cells treated with tranilast. Results and conclusions . The expression of FAP-α gene and fibroblast activation protein α is higher in keloid fibroblasts. Tranilast at concentrations of 3 μM and 30 μM up-regulated mRNA FAP-α expression in normal fibroblasts but did not influence keloid fibroblasts. The drug, at concentrations of 30 μM and 300 μM up-regulated fibroblast activation protein α expression in normal fibroblasts and did not influence keloid fibroblasts. Tranilast antiproliferative effect is not associated with FAP-α expression in keloid fibroblasts.

  16. Deposition of insoluble elastin by pulmonary fibroblasts from patients with COPD is increased by treatment with versican siRNA

    Directory of Open Access Journals (Sweden)

    Wu L

    2017-01-01

    Full Text Available Lian Wu,1,2 Jing Zhang,3 Jie Ming Qu,4 Chun-xue Bai,3 Mervyn J Merrilees5 1Department of Community and Health Services, Unitec, 2Department of Pharmacology & Clinical Pharmacology, University of Auckland, Auckland, New Zealand; 3Department of Pulmonary Medicine, Zhongshan Hospital, Fudan University, 4Department of Pulmonary Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China; 5Department of Anatomy and Medical Imaging, University of Auckland, Auckland, New Zealand Abstract: A reduced content of alveolar elastic fibers is a key feature of COPD lung. Despite continued elastogenic potential by alveolar fibroblasts in the lung affected by COPD, repair of elastic fibers does not take place, which is due to increased levels of the chondroitin sulfate proteoglycan versican that inhibits the assembly of tropoelastin into fibers. In this study, primary pulmonary fibroblast cell lines from COPD and non-COPD patients were treated with a small interfering RNA (siRNA against versican to determine if knockdown of versican could restore the deposition of insoluble elastin. Versican siRNA treatment reduced versican expression and secretion by pulmonary fibroblasts from both COPD and non-COPD patients (P<0.01 and significantly increased deposition of insoluble elastin in the COPD cell cultures (P<0.05. The treatment, however, did not significantly affect production of soluble elastin (tropoelastin in either the COPD or non-COPD cell cultures, supporting a role for versican in inhibiting assembly but not synthesis of tropoelastin. These results suggest that removal or knockdown of versican may be a possible therapeutic strategy for increasing deposition of insoluble elastin and stimulating repair of elastic fibers in COPD lung. Keywords: pulmonary fibroblasts, COPD, elastin, versican

  17. miR-130b-3p Modulates Epithelial-Mesenchymal Crosstalk in Lung Fibrosis by Targeting IGF-1.

    Science.gov (United States)

    Li, Shuhong; Geng, Jing; Xu, Xuefeng; Huang, Xiaoxi; Leng, Dong; Jiang, Dingyuan; Liang, Jiurong; Wang, Chen; Jiang, Dianhua; Dai, Huaping

    2016-01-01

    Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and usually lethal fibrotic lung disease with largely unknown etiology and pathogenesis. Evidence suggests microRNAs (miRNA) contribute to pathogenesis of IPF. In this study, we sought to identify miRNA expression signatures and determine the role of miR-130b-3p in lung fibrosis. The miRNA expression profile of the lungs from patients with IPF and normal donors was determined by Affymetrix microarray, and transcriptome with Affymetrix array. The functions and signal pathways as well as miRNA-mRNA networks were established by bioinformatics analysis. Luciferase assays and ELISA were used to confirm the miRNA target gene. The effect of miRNA-transfected epithelium on fibroblast activities was assessed using a co-culture system. The fibroblast activities were determined by qRT-PCR, western blotting, Transwell and BrdU assays. Seven miRNAs were significantly decreased in IPF lungs, with miR-130b-3p being the highest in the miRNA-mRNA network. Insulin-like growth factor (IGF-1) was a target gene of miR-130b-3p in the epithelium. miR-130b-3p inhibition in the epithelium induced collagen I expression and enhanced the proliferation and migration ability of fibroblast in co-culture systems, which mimicked the functions of exogenous IGF-1 on fibroblasts. Neutralizing IGF-1 with an antibody significantly reduced the modulatory effects of miR-130b-3p inhibitor-transfected epithelium on the activation of fibroblasts. Our results show that miR-130b-3p was downregulated in IPF lungs. miR-130b-3p downregulation contributed to the activation of fibroblasts and the dysregulated epithelial-mesenchymal crosstalk by promoting IGF-1 secretion from lung epithelium, suggesting a key regulatory role for this miRNA in preventing lung fibrosis.

  18. Isolation of pulmonary artery smooth muscle cells from neonatal mice.

    Science.gov (United States)

    Lee, Keng Jin; Czech, Lyubov; Waypa, Gregory B; Farrow, Kathryn N

    2013-10-19

    Pulmonary hypertension is a significant cause of morbidity and mortality in infants. Historically, there has been significant study of the signaling pathways involved in vascular smooth muscle contraction in PASMC from fetal sheep. While sheep make an excellent model of term pulmonary hypertension, they are very expensive and lack the advantage of genetic manipulation found in mice. Conversely, the inability to isolate PASMC from mice was a significant limitation of that system. Here we described the isolation of primary cultures of mouse PASMC from P7, P14, and P21 mice using a variation of the previously described technique of Marshall et al. that was previously used to isolate rat PASMC. These murine PASMC represent a novel tool for the study of signaling pathways in the neonatal period. Briefly, a slurry of 0.5% (w/v) agarose + 0.5% iron particles in M199 media is infused into the pulmonary vascular bed via the right ventricle (RV). The iron particles are 0.2 μM in diameter and cannot pass through the pulmonary capillary bed. Thus, the iron lodges in the small pulmonary arteries (PA). The lungs are inflated with agarose, removed and dissociated. The iron-containing vessels are pulled down with a magnet. After collagenase (80 U/ml) treatment and further dissociation, the vessels are put into a tissue culture dish in M199 media containing 20% fetal bovine serum (FBS), and antibiotics (M199 complete media) to allow cell migration onto the culture dish. This initial plate of cells is a 50-50 mixture of fibroblasts and PASMC. Thus, the pull down procedure is repeated multiple times to achieve a more pure PASMC population and remove any residual iron. Smooth muscle cell identity is confirmed by immunostaining for smooth muscle myosin and desmin.

  19. Mechanisms of vitamin K transport and metabolism in Swiss 3T3 mouse fibroblasts

    International Nuclear Information System (INIS)

    Canfield, L.M.; Townsend, A.F.; Hibbs, D.B.

    1986-01-01

    Transport of vitamin K into isolated fibroblasts was followed using 3 H vitamin K 1 . The initial rate is saturable by 5 min. at 25μM vitamin K with a Km(app) of 10μM and V/sub max/ of 50 pmols/min/10 6 cells. Kinetics of uptake are biphasic with a second slower rate ensuing after 10 minutes. Insensitivity of the initial rate of uptake to FCCP or ouabain indicates an ATP-independent transport mechanism. Specificity of transport is shown by competition of uptake of 3 H vitamin K by unlabelled vitamin and strong (>90%) inhibition of the initial rate by equimolar concentrations of the vitamin K analog, Chloro-K. In addition, following uptake, both vitamins K 1 and K 2 are metabolized to their respective epoxides. Vitamin K 1 epoxide is also transported into fibroblasts and metabolized to the parent quinone in a Warfarin-sensitive reaction. Following alkaline hydrolysis of isolated intracellular protein, the vitamin K-dependent amino acid, gamma carboxyglutamic acid (gla) was detected. It is concluded that vitamin K is specifically transported into fibroblasts and metabolized via the classical pathway described in liver with the concomitant production of vitamin K-dependent proteins

  20. Effects of plant lectins on in vitro fibroblast proliferation

    Directory of Open Access Journals (Sweden)

    Ana Maria Sell

    2003-06-01

    Full Text Available Lectins are carbohydrate-binding proteins that have been isolated from various sources and presented a wide spectrum of biological activities. The effects of four lectins, namely, Phaseolus vulgaris phytohemagglutinin, PHA, wheat germ agglutinin, WGA, Artocarpus integrifolia seed lectins, jacalin and artocarpin, on in vitro fibroblasts proliferation were investigated. The lectins did not influence the initial cell adhesion to the plate. PHA and WGA at 10-20 µg/mL concentrations significantly decreased fibroblasts proliferation. At these concentrations, they caused morphological alterations on cells and over 80 µg/mL, promoted cell death. Neither jacalin nor artocarpin significantly affected cell proliferation.O objetivo deste trabalho foi avaliar a influência das lectinas PHA, WGA, jacalina e artocarpina sobre a proliferação de fibroblastos in vitro. Para tanto, fibroblastos gengivais de voluntários saudáveis foram cultivados, por cinco dias, em DMEM suplementado com soro bovino fetal (10% v/v e na presença das lectinas nas concentrações finais de 0.1 a 300 µg/mL. A adesão, o crescimento e a morfologia celular foram acompanhados por microscopia de inversão e contraste de fase. O índice de proliferação foi avaliado pelo método calorimétrico usando MTT. As lectinas não alteraram a adesão inicial dos fibroblastos à placa de poliestireno. PHA e WGA, nas concentrações de 10 a 20 µg/mL, diminuíram significativamente a proliferação celular. Nestas concentrações a morfologia celular é alterada e acima de 80 µg/mL, houve100% de morte celular. As lectinas jacalina e artocarpina não influenciaram a proliferação celular.

  1. circHIPK2-mediated σ-1R promotes endoplasmic reticulum stress in human pulmonary fibroblasts exposed to silica.

    Science.gov (United States)

    Cao, Zhouli; Xiao, Qingling; Dai, Xiaoniu; Zhou, Zewei; Jiang, Rong; Cheng, Yusi; Yang, Xiyue; Guo, Huifang; Wang, Jing; Xi, Zhaoqing; Yao, Honghong; Chao, Jie

    2017-12-13

    Silicosis is characterized by fibroblast accumulation and excessive deposition of extracellular matrix. Although the roles of SiO 2 -induced chemokines and cytokines released from alveolar macrophages have received significant attention, the direct effects of SiO 2 on protein production and functional changes in pulmonary fibroblasts have been less extensively studied. Sigma-1 receptor, which has been associated with cell proliferation and migration in the central nervous system, is expressed in the lung, but its role in silicosis remains unknown. To elucidate the role of sigma-1 receptor in fibrosis induced by silica, both the upstream molecular mechanisms and the functional effects on cell proliferation and migration were investigated. Both molecular biological assays and pharmacological techniques, combined with functional experiments, such as migration and proliferation, were applied in human pulmonary fibroblasts from adults to analyze the molecular and functional changes induced by SiO 2 . SiO 2 induced endoplasmic reticulum stress in association with enhanced expression of sigma-1 receptor. Endoplasmic reticulum stress promoted migration and proliferation of human pulmonary fibroblasts-adult exposed to SiO 2 , inducing the development of silicosis. Inhibition of sigma-1 receptor ameliorated endoplasmic reticulum stress and fibroblast functional changes induced by SiO 2 . circHIPK2 is involved in the regulation of sigma-1 receptor in human pulmonary fibroblasts-adult exposed to SiO 2 . Our study elucidated a link between SiO 2 -induced fibrosis and sigma-1 receptor signaling, thereby providing novel insight into the potential use of sigma-1 receptor/endoplasmic reticulum stress in the development of novel therapeutic strategies for silicosis treatment.

  2. Nicotine signals through muscle-type and neuronal nicotinic acetylcholine receptors in both human bronchial epithelial cells and airway fibroblasts

    Directory of Open Access Journals (Sweden)

    Luketich James D

    2004-12-01

    Full Text Available Abstract Background Non-neuronal cells, including those derived from lung, are reported to express nicotinic acetylcholine receptors (nAChR. We examined nAChR subunit expression in short-term cultures of human airway cells derived from a series of never smokers, ex-smokers, and active smokers. Methods and Results At the mRNA level, human bronchial epithelial (HBE cells and airway fibroblasts expressed a range of nAChR subunits. In multiple cultures of both cell types, mRNA was detected for subunits that constitute functional muscle-type and neuronal-type pentomeric receptors. Two immortalized cell lines derived from HBE cells also expressed muscle-type and neuronal-type nAChR subunits. Airway fibroblasts expressed mRNA for three muscle-type subunits (α1, δ, and ε significantly more often than HBE cells. Immunoblotting of HBE cell and airway fibroblast extracts confirmed that mRNA for many nAChR subunits is translated into detectable levels of protein, and evidence of glycosylation of nAChRs was observed. Some minor differences in nAChR expression were found based on smoking status in fibroblasts or HBE cells. Nicotine triggered calcium influx in the immortalized HBE cell line BEAS2B, which was blocked by α-bungarotoxin and to a lesser extent by hexamethonium. Activation of PKC and MAPK p38, but not MAPK p42/44, was observed in BEAS2B cells exposed to nicotine. In contrast, nicotine could activate p42/44 in airway fibroblasts within five minutes of exposure. Conclusions These results suggest that muscle-type and neuronal-type nAChRs are functional in airway fibroblasts and HBE cells, that prior tobacco exposure does not appear to be an important variable in nAChR expression, and that distinct signaling pathways are observed in response to nicotine.

  3. Aryl hydrocarbon receptor–ligand axis mediates pulmonary fibroblast migration and differentiation through increased arachidonic acid metabolism

    International Nuclear Information System (INIS)

    Su, Hsiang-Han; Lin, Hsin-Ting; Suen, Jau-Ling; Sheu, Chau Chyun; Yokoyama, Kazunari K.; Huang, Shau-Ku; Cheng, Chih Mei

    2016-01-01

    Pulmonary fibroblast migration and differentiation are critical events in fibrogenesis; meanwhile, fibrosis characterizes the pathology of many respiratory diseases. The role of aryl hydrocarbon receptor (AhR), a unique cellular chemical sensor, has been suggested in tissue fibrosis, but the mechanisms through which the AhR-ligand axis influences the fibrotic process remain undefined. In this study, the potential impact of the AhR-ligand axis on pulmonary fibroblast migration and differentiation was analyzed using human primary lung fibroblasts HFL-1 and CCL-202 cells. Boyden chamber-based cell migration assay showed that activated AhR in HFL-1cells significantly enhanced cell migration in response to 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD), and a known AhR antagonist, CH223191, inhibited its migratory activity. Furthermore, the calcium mobilization and subsequent upregulated expression of arachidonic acid metabolizing enzymes, including cyclooxygenase2 (COX-2) and 5-lipoxygenase (5-LOX), were observed in TCDD-treated HFL-1 cells, concomitant with elevated levels of prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) secretion. Also, significantly increased expression of α-smooth muscle actin α-SMA), a fibroblast differentiation marker, was also noted in TCDD-treated HFL-1 cells (p < 0.05), resulting in a dynamic change in cytoskeleton protein levels and an increase in the nuclear translocation of the myocardin-related transcription factor. Moreover, the enhanced levels of α-SMA expression and fibroblast migration induced by TCDD, PGE2 and LTB4 were abrogated by selective inhibitors for COX-2 and 5-LOX. Knockdown of AhR by siRNA Completely diminished intracellular calcium uptake and reduced α-SMA protein verified by promoter-reporter assays and chromatin immunoprecipitation. Taken together, our results suggested the importance of the AhR-ligand axis in fibroblast migration and differentiation through its capacity in enhancing arachidonic acid metabolism.

  4. Differentiation state of skin fibroblast cultures versus risk of subcutaneous fibrosis after radiotherapy

    International Nuclear Information System (INIS)

    Herskind, C.; Bamberg, M.; Rodemann, H.P.; Bentzen, S.M.; Overgaard, J.; Overgaard, M.

    1998-01-01

    Background and purpose: There is increasing evidence for patient-to-patient variation in the response of normal tissue to radiotherapy. Recently, it has been suggested that accumulation of functional fibrocytes may be a key step in the development of radiation-induced fibrosis. Therefore, we have examined a possible relationship between the differentiation state of untreated fibroblasts and the risk of radiation-induced subcutaneous fibrosis in individual patients. Materials and methods: We used skin fibroblast cultures isolated from eight postmastectomy radiotherapy patients whose individual clinical radiosensitivity was assessed by the mean excess risk of fibrosis. Different types of potentially mitotic progenitor fibroblasts (MF) and postmitotic functional fibrocytes (PMF) in the terminal differentiation lineage (MFI approaches MFII approaches MFIII approaches PMF) were scored morphologically in clonal culture. Progression of differentiation was quantified by the ratio L/E of colony-forming late (MFIII and late MFII) and early (MFI and early MFII) progenitors. Results: We observed a correlation between the ratio L/E and the mean risk of fibrosis (r S =0.743, P=0.03), indicating an approximately 10-fold increase in L/E with an increasing risk of fibrosis. This was paralleled by a decreasing trend in the absolute numbers of early progenitor types. By contrast, there was no significant correlation between the plating efficiency and the risk of fibrosis. Conclusions: The data suggest that the risk of fibrosis increases with the progression of the differentiation of untreated progenitor fibroblasts, indicating that the progression of fibroblast differentiation may be a co-factor in the development of radiation-induced fibrosis. If this hypothesis is validated, it provides a rationale for a novel predictive test to identify patients with an increased risk of subcutaneous fibrosis. (Copyright (c) 1998 Elsevier Science B.V., Amsterdam. All rights reserved.)

  5. Fibroblast activation protein is dispensable in the anti-influenza immune response in mice.

    Directory of Open Access Journals (Sweden)

    Sioh-Yang Tan

    Full Text Available Fibroblast activation protein alpha (FAP is a unique dual peptidase of the S9B serine protease family, being capable of both dipeptidyl peptidase and endopeptidase activities. FAP is expressed at low level in healthy adult organs including the pancreas, cervix, uterus, submaxillary gland and the skin, and highly upregulated in embryogenesis, chronic inflammation and tissue remodelling. It is also expressed by cancer-associated stromal fibroblasts in more than 90% of epithelial tumours. FAP has enzymatic and non-enzymatic functions in the growth, immunosuppression, invasion and cell signalling of tumour cells. FAP deficient mice are fertile and viable with no gross abnormality, but little data exist on the role of FAP in the immune system. FAP is upregulated in association with microbial stimulation and chronic inflammation, but its function in infection remains unknown. We showed that major populations of immune cells including CD4+ and CD8+ T cells, B cells, dendritic cells and neutrophils are generated and maintained normally in FAP knockout mice. Upon intranasal challenge with influenza virus, FAP mRNA was increased in the lungs and lung-draining lymph nodes. Nonetheless, FAP deficient mice showed similar pathologic kinetics to wildtype controls, and were capable of supporting normal anti-influenza T and B cell responses. There was no evidence of compensatory upregulation of other DPP4 family members in influenza-infected FAP-deficient mice. FAP appears to be dispensable in anti-influenza adaptive immunity.

  6. Understanding the mechanisms of lung mechanical stress

    Directory of Open Access Journals (Sweden)

    C.S.N.B. Garcia

    2006-06-01

    Full Text Available Physical forces affect both the function and phenotype of cells in the lung. Bronchial, alveolar, and other parenchymal cells, as well as fibroblasts and macrophages, are normally subjected to a variety of passive and active mechanical forces associated with lung inflation and vascular perfusion as a result of the dynamic nature of lung function. These forces include changes in stress (force per unit area or strain (any forced change in length in relation to the initial length and shear stress (the stress component parallel to a given surface. The responses of cells to mechanical forces are the result of the cell's ability to sense and transduce these stimuli into intracellular signaling pathways able to communicate the information to its interior. This review will focus on the modulation of intracellular pathways by lung mechanical forces and the intercellular signaling. A better understanding of the mechanisms by which lung cells transduce physical forces into biochemical and biological signals is of key importance for identifying targets for the treatment and prevention of physical force-related disorders.

  7. Oral fibroblasts produce more HGF and KGF than skin fibroblasts in response to co-culture with keratinocytes

    DEFF Research Database (Denmark)

    Grøn, Birgitte; Stoltze, Kaj; Andersson, Anders

    2002-01-01

    The production of hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF) in subepithelial fibroblasts from buccal mucosa, periodontal ligament, and skin was determined after co-culture with keratinocytes. The purpose was to detect differences between the fibroblast subpopulations...... days by ELISA. When cultured on polystyrene, the constitutive level of KGF and HGF in periodontal fibroblasts was higher than the level in buccal and skin fibroblasts. In the presence of keratinocytes, all three types of fibroblasts in general increased their HGF and KGF production 2-3 times. When...... cells were maintained in collagen, the level of HGF and KGF was decreased mainly in skin cultures. However, in oral fibroblasts, induction after stimulation was at a similar level in collagen compared to on polystyrene. Skin fibroblasts maintained in collagen produced almost no HGF whether...

  8. The use of chitosan-dextran gel shows anti-inflammatory, antibiofilm, and antiproliferative properties in fibroblast cell culture.

    Science.gov (United States)

    Paramasivan, Sathish; Jones, Damien; Baker, Leonie; Hanton, Lyall; Robinson, Simon; Wormald, Peter J; Tan, Lorwai

    2014-01-01

    Chitosan-dextran gel has been used as an antihemostatic agent and antiadhesive agent after endoscopic sinus surgery. Because Staphylococcus aureus biofilms have been implicated in recalcitrant chronic rhinosinusitis, this study aimed to further investigate the (i) anti-inflammatory, (ii) bacterial biofilm inhibition, (iii) antiproliferative effects, and (iv) wound-healing properties of chitosan and chitosan-dextran gel. Fibroblasts were isolated from human nasal tissue and were used to determine the effects of chitosan and chitosan-dextran gel on (i) cell proliferation, (ii) wound healing, (iii) inflammation in fibroblast cultures challenged with superantigens S. aureus enterotoxin B (SEB) and toxic shock syndrome toxin (TSST), and (iv) on S. aureus biofilms. Chitosan was highly effective at reducing IL-8 expression after TSST and SEB challenge. Chitosan was also effective at reducing IL-8 expression of nonchallenged fibroblasts showing its anti-inflammatory effects on fibroblasts in a diseased state. Chitosan-dextran gel showed strong antibiofilm properties at 50% (v/v) concentration in vitro. Dextran, on its own, showed antibiofilm properties at 1.25% (w/v) concentration. Chitosan, on its own, reduced proliferation of fibroblasts to 82% of control proliferation and chitosan-dextran gel reduced proliferation of the fibroblasts to 0.04% of control proliferation. Relative to the no treatment controls, chitosan-dextran gel significantly delayed the wound-healing rate over the first 48 hours of the experiment. Chitosan-dextran gel reduced fibroblast proliferation and wound-healing time, showing a possible mechanism of reducing adhesions in the postsurgical period. Chitosan reduced IL-8 levels, showing its anti-inflammatory properties. Chitosan-dextran gel and dextran treatment showed antibiofilm properties in our model.

  9. Survival of human diploid skin fibroblasts from normal individuals after X-irradiation

    International Nuclear Information System (INIS)

    Little, J.B.; Nove, J.; Strong, L.C.; Nichols, W.W.

    1988-01-01

    The cytotoxic effect of X-rays was measured by a colony formation assay in multiple experiments with fibroblast cell strains derived from 24 presumably normal individuals, received as 65 different coded and blinded samples. Each strain was received on two or more occasions at different times and bearing different codes. The means and standard deviations of the survival curve parameters for the 24 strains were: D 0 = 123 +- 23; D 10 = 273 +- 42 cGy. The D 0 ranged from 89 to 175 and the D 10 from 196 to 372 cGy. The degree of interexperimental variation, though generally minimal, differed considerably among cell strains. There was no systematic effect of passage level, cloning efficiency, serum lot, age or sex of the donor on X-ray survival. These results confirm that the intrinsic radiosensitivity varies significantly among skin fibroblasts isolated from clinically normal individuals, apparently owing to as yet unidentified genetic factors. (Author)

  10. Knockout of Myostatin by Zinc-finger Nuclease in Sheep Fibroblasts and Embryos

    Directory of Open Access Journals (Sweden)

    Xuemei Zhang

    2016-10-01

    Full Text Available Myostatin (MSTN can negatively regulate the growth and development of skeletal muscle, and natural mutations can cause “double-muscling” trait in animals. In order to block the inhibiting effect of MSTN on muscle growth, we transferred zinc-finger nucleases (ZFN which targeted sheep MSTN gene into cultured fibroblasts. Gene targeted colonies were isolated from transfected fibroblasts by serial dilution culture and screened by sequencing. Two colonies were identified with mono-allele mutation and one colony with bi-allelic deletion. Further, we introduced the MSTN-ZFN mRNA into sheep embryos by microinjection. Thirteen of thirty-seven parthenogenetic embryos were targeted by ZFN, with the efficiency of 35%. Our work established the technical foundation for generation of MSTN gene editing sheep by somatic cloning and microinjection ZFN into embryos.

  11. Gene targeting in adult rhesus macaque fibroblasts

    Directory of Open Access Journals (Sweden)

    Wolf Don P

    2008-03-01

    Full Text Available Abstract Background Gene targeting in nonhuman primates has the potential to produce critical animal models for translational studies related to human diseases. Successful gene targeting in fibroblasts followed by somatic cell nuclear transfer (SCNT has been achieved in several species of large mammals but not yet in primates. Our goal was to establish the protocols necessary to achieve gene targeting in primary culture of adult rhesus macaque fibroblasts as a first step in creating nonhuman primate models of genetic disease using nuclear transfer technology. Results A primary culture of adult male fibroblasts was transfected with hTERT to overcome senescence and allow long term in vitro manipulations. Successful gene targeting of the HPRT locus in rhesus macaques was achieved by electroporating S-phase synchronized cells with a construct containing a SV40 enhancer. Conclusion The cell lines reported here could be used for the production of null mutant rhesus macaque models of human genetic disease using SCNT technology. In addition, given the close evolutionary relationship and biological similarity between rhesus macaques and humans, the protocols described here may prove useful in the genetic engineering of human somatic cells.

  12. Stimulation of alveolar macrophages by BCG vaccine enhances the process of lung fibrosis induced by bleomycin.

    Science.gov (United States)

    Chyczewska, E; Chyczewski, L; Bańkowski, E; Sułkowski, S; Nikliński, J

    1993-01-01

    It was found that the BCG vaccine injected subcutaneously to the rats enhances the process of lung fibrosis induced by bleomycin. Pretreatment of rats with this vaccine results in accumulation of activated macrophages in lung interstitium and in the bronchoalveolar spaces. It may be suggested that the activated macrophages release various cytokines which may stimulate the proliferation of fibroblasts and biosynthesis of extracellular matrix components.

  13. Metalloproteinase 11, potential marker and molecular target in advanced and castration-resistant prostate cancer. Culture study of peritumoral fibroblasts.

    Science.gov (United States)

    Fernandez-Gomez, J M; Eiro, N; García-Rodríguez, J J; Quintás-Blanco, A; Gonzalez-Ruiz de León, C; Perez de Haro, M L; Vizoso-Piñero, F

    To analyze the expression of metalloprotein 11 (MMP11) in cultured fibroblasts obtained from human prostate tumors with different clinical and pathological characteristics. For this study we analyzed samples of transrectal prostate biopsies from tumors with different characteristics, treated with or whithout androgen deprivation (AD). After optimization of the culture method, fibroblasts were isolated and cultured to perform the study (PCR) of MMP11 mRNA. Finally, 37 cases were studied: 5 samples of benign prostatic hyperplasia, 14 cases with localized neoplasms (7 high-risk according to the D'Amico classification), 5 with metastasic tumors (bone metastases), and 13 treated with AD therapy, of which 6 fulfilled the requirements to be defined as resistant to castration. In tumors without AD therapy, MMP11 expression was significantly higher (P=.001) in fibroblasts of higher grade tumors. A significant (P=.001) correlation was found between PSA and expression of MMP11 in fibroblast s and a significant increase of MMP11 expression in metastatic tumors. In tumors with AD therapy, a significantly greater expression of MMP11 was observed in resistant to castration patients than in those sensitive to castration (P=.003). In advanced prostate tumors or in stages of increased tumor aggressiveness, the production of MMP11 by fibroblasts is significantly greater than in non-metastatic tumors or in AD sensitive tumors. Copyright © 2016 AEU. Publicado por Elsevier España, S.L.U. All rights reserved.

  14. The lungs

    International Nuclear Information System (INIS)

    Macey, D.J.; Marshall, R.

    1982-01-01

    Currently emission tomography of the lungs is only practical for perfusion images with sup(99m)Tc microaggregates and ventilation images with sup(81m)Kr. The following topics are touched on: the rotating gamma camera single photon ECT system, spatial resolution and linearity, resolution in phantom studies, area and volume studies, quantitation studies, with particular reference to the authors' experience of perfusion and ventilation in investigations of pulmonary embolism. (U.K.)

  15. Improved methods for reprogramming human dermal fibroblasts using fluorescence activated cell sorting.

    Directory of Open Access Journals (Sweden)

    David J Kahler

    Full Text Available Current methods to derive induced pluripotent stem cell (iPSC lines from human dermal fibroblasts by viral infection rely on expensive and lengthy protocols. One major factor contributing to the time required to derive lines is the ability of researchers to identify fully reprogrammed unique candidate clones from a mixed cell population containing transformed or partially reprogrammed cells and fibroblasts at an early time point post infection. Failure to select high quality colonies early in the derivation process results in cell lines that require increased maintenance and unreliable experimental outcomes. Here, we describe an improved method for the derivation of iPSC lines using fluorescence activated cell sorting (FACS to isolate single cells expressing the cell surface marker signature CD13(NEGSSEA4(POSTra-1-60(POS on day 7-10 after infection. This technique prospectively isolates fully reprogrammed iPSCs, and depletes both parental and "contaminating" partially reprogrammed fibroblasts, thereby substantially reducing the time and reagents required to generate iPSC lines without the use of defined small molecule cocktails. FACS derived iPSC lines express common markers of pluripotency, and possess spontaneous differentiation potential in vitro and in vivo. To demonstrate the suitability of FACS for high-throughput iPSC generation, we derived 228 individual iPSC lines using either integrating (retroviral or non- integrating (Sendai virus reprogramming vectors and performed extensive characterization on a subset of those lines. The iPSC lines used in this study were derived from 76 unique samples from a variety of tissue sources, including fresh or frozen fibroblasts generated from biopsies harvested from healthy or disease patients.

  16. Hyperlucent lung

    International Nuclear Information System (INIS)

    Jimenez-Gutierrez, Florana; Soto-Quiros, Manuel E.

    2007-01-01

    Unilateral hyperlucent lung is also known as Swyer-James Syndrome, Macleod Syndrome or lobular or unilateral emphysema. It is an uncommon disease characterized by lung or unilateral lobe hiperlucency associated to an air trapping upon expiration. As regards to etiology, this syndrome is considered to be an acquired disease that appears secondary to respiratory infections during the early years of life, probably bronchiolitis and/ or viral pneumonia. The clinical presentation varies among patients. Some of them are asymptomatic, others present a history of recurrent episodes of pulmonary infections from early years of life or present effort dyspnea. The diagnosis is usually made accidentally by a chest radiograph in a child with history of respiratory infections or in an adult during a routine chest x- ray in an asymptomatic person. It is important to differentiate this syndrome from other causes of unilateral pulmonary hiperlucency on conventional chest x-rays. Few cases of Swyer-James Syndrome in children have been reported, it is presented the clinical case of a patient who had a parainfluenza 3 bronchopneumonia when he was a month and eighteen days of age. The differential diagnosis of this syndrome should be done with other thoracic entities that diminish the radiological pulmonary unilateral density. A case of a child who is the bearer of hyperlucent lung is described. (author) [es

  17. Pentagalloyl glucose increases elastin deposition, decreases reactive oxygen species and matrix metalloproteinase activity in pulmonary fibroblasts under inflammatory conditions.

    Science.gov (United States)

    Parasaram, Vaideesh; Nosoudi, Nasim; Chowdhury, Aniqa; Vyavahare, Naren

    2018-04-30

    Emphysema is characterized by degradation of lung alveoli that leads to poor airflow in lungs. Irreversible elastic fiber degradation by matrix metalloproteinases (MMPs) and reactive oxygen species (ROS) activity leads to loss of elasticity and drives the progression of this disease. We investigated if a polyphenol, pentagalloyl glucose (PGG) can increase elastin production in pulmonary fibroblasts. We also studied the effect of PGG treatment in reducing MMP activity and ROS levels in cells. We exposed rat pulmonary fibroblasts to two different types of inflammatory environments i.e., tumor necrosis factor-α (TNF-α) and cigarette smoke extract (CSE) to mimic the disease. Parameters like lysyl oxidase (LOX) and elastin gene expression, MMP-9 activity in the medium, lysyl oxidase (LOX) activity and ROS levels were studied to assess the effect of PGG on pulmonary fibroblasts. CSE inhibited lysyl oxidase (LOX) enzyme activity that resulted in a decreased elastin formation. Similarly, TNF-α treated cells showed less elastin in the cell layers. Both these agents caused increase in MMP activity and ROS levels in cells. However, when supplemented with PGG treatment along with these two inflammatory agents, we saw a significant increase in elastin deposition, reduction in both MMP activity and ROS levels. Thus PGG, which has anti-inflammatory, anti-oxidant properties coupled with its ability to aid in elastic fiber formation, can be a multifunctional drug to potentially arrest the progression of emphysema. Copyright © 2018 Elsevier Inc. All rights reserved.

  18. Multimodal imaging of lung cancer and its microenvironment (Conference Presentation)

    Science.gov (United States)

    Hariri, Lida P.; Niederst, Matthew J.; Mulvey, Hillary; Adams, David C.; Hu, Haichuan; Chico Calero, Isabel; Szabari, Margit V.; Vakoc, Benjamin J.; Hasan, Tayyaba; Bouma, Brett E.; Engelman, Jeffrey A.; Suter, Melissa J.

    2016-03-01

    Despite significant advances in targeted therapies for lung cancer, nearly all patients develop drug resistance within 6-12 months and prognosis remains poor. Developing drug resistance is a progressive process that involves tumor cells and their microenvironment. We hypothesize that microenvironment factors alter tumor growth and response to targeted therapy. We conducted in vitro studies in human EGFR-mutant lung carcinoma cells, and demonstrated that factors secreted from lung fibroblasts results in increased tumor cell survival during targeted therapy with EGFR inhibitor, gefitinib. We also demonstrated that increased environment stiffness results in increased tumor survival during gefitinib therapy. In order to test our hypothesis in vivo, we developed a multimodal optical imaging protocol for preclinical intravital imaging in mouse models to assess tumor and its microenvironment over time. We have successfully conducted multimodal imaging of dorsal skinfold chamber (DSC) window mice implanted with GFP-labeled human EGFR mutant lung carcinoma cells and visualized changes in tumor development and microenvironment facets over time. Multimodal imaging included structural OCT to assess tumor viability and necrosis, polarization-sensitive OCT to measure tissue birefringence for collagen/fibroblast detection, and Doppler OCT to assess tumor vasculature. Confocal imaging was also performed for high-resolution visualization of EGFR-mutant lung cancer cells labeled with GFP, and was coregistered with OCT. Our results demonstrated that stromal support and vascular growth are essential to tumor progression. Multimodal imaging is a useful tool to assess tumor and its microenvironment over time.

  19. Efficient direct conversion of human fibroblasts into myogenic lineage induced by co-transduction with MYCL and MYOD1.

    Science.gov (United States)

    Wakao, Junko; Kishida, Tsunao; Fumino, Shigehisa; Kimura, Koseki; Yamamoto, Kenta; Kotani, Shin-Ichiro; Mizushima, Katsura; Naito, Yuji; Yoshikawa, Toshikazu; Tajiri, Tatsuro; Mazda, Osam

    2017-06-24

    The skeletal muscle consists of contractile myofibers and plays essential roles for maintenance of body posture, movement, and metabolic regulation. During the development and regeneration of the skeletal muscle tissue, the myoblasts fuse into multinucleated myotubes that subsequently form myofibers. Transplantation of myoblasts may make possible a novel regenerative therapy against defects or dysfunction of the skeletal muscle. It is reported that rodent fibroblasts are converted into myoblast-like cells and fuse to form syncytium after forced expression of exogenous myogenic differentiation 1 (MYOD1) that is a key transcription factor for myoblast differentiation. But human fibroblasts are less efficiently converted into myoblasts and rarely fused by MYOD1 alone. Here we found that transduction of v-myc avian myelocytomatosis viral oncogene lung carcinoma derived homolog (MYCL) gene in combination with MYOD1 gene induced myoblast-like phenotypes in human fibroblasts more strongly than MYOD1 gene alone. The rate of conversion was approximately 90%. The directly converted myoblasts (dMBs) underwent fusion in an ERK5 pathway-dependent manner. The dMBs also formed myofiber-like structure in vivo after an inoculation into mice at the subcutaneous tissue. The present results strongly suggest that the combination of MYCL plus MYOD1 may promote direct conversion of human fibroblasts into functional myoblasts that could potentially be used for regenerative therapy for muscle diseases and congenital muscle defects. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Extracellular ATP drives breast cancer cell migration and metastasis via S100A4 production by cancer cells and fibroblasts.

    Science.gov (United States)

    Liu, Ying; Geng, Yue-Hang; Yang, Hui; Yang, Han; Zhou, Yan-Ting; Zhang, Hong-Quan; Tian, Xin-Xia; Fang, Wei-Gang

    2018-05-04

    Our previous work has demonstrated that extracellular ATP is an important pro-invasive factor, and in this study, we tapped into a possible mechanism involved. We discovered that ATP could upregulate both the intracellular expression and secretion of S100A4 in breast cancer cells and fibroblasts. Apart from stimulating breast cancer cell motility via intracellular S100A4, ATP enhanced the ability of breast cancer cells to transform fibroblasts into cancer-associated fibroblast (CAF)-like cells, which in turn secreted S100A4 to further promote cancer cell motility. Both apyrase and niclosamide treatments could inhibit metastasis of inoculated tumors to lung, liver and kidney in mice model, and CAFs from these treated tumors exhibited weakened migration-stimulating capacity for breast cancer cells. Collectively, our data indicate that extracellular ATP promotes the interactions between breast cancer cells and fibroblasts, which work collaboratively via production of S100A4 to exacerbate breast cancer metastasis. Copyright © 2018. Published by Elsevier B.V.

  1. Inflammation and angiogenesis in fibrotic lung disease.

    Science.gov (United States)

    Keane, Michael P; Strieter, Robert M; Lynch, Joseph P; Belperio, John A

    2006-12-01

    The pathogenesis of pulmonary fibrosis is poorly understood. Although inflammation has been presumed to have an important role in the development of fibrosis this has been questioned recently, particularly with regard to idiopathic pulmonary fibrosis (IPF). It is, however, increasingly recognized that the polarization of the inflammatory response toward a type 2 phenotype supports fibroproliferation. Increased attention has been on the role of noninflammatory structural cells such as the fibroblast, myofibroblast, epithelial cell, and endothelial cells. Furthermore, the origin of these cells appears to be multifactorial and includes resident cells, bone marrow-derived cells, and epithelial to mesenchymal transition. Increasing evidence supports the presence of vascular remodeling in fibrotic lung disease, although the precise role in the pathogenesis of fibrosis remains to be determined. Therefore, the pathogenesis of pulmonary fibrosis is complex and involves the interaction of multiple cell types and compartments within the lung.

  2. Prevalence and Biotyping of Pasteurella Haemolytica Isolates from ...

    African Journals Online (AJOL)

    P. haemolytica isolated from Sahel sheep and goat in Maiduguri was characterized phenotypically. A total of 92 P. haemolytica isolates were obtained from the nasopharyngeal swabs while a total of 15 isolates came from pneumonic lung samples. The results showed that 37(20.22%) P. haemolytica isolates were obtained ...

  3. Early structural changes in sheep lung following thoracic irradiation

    International Nuclear Information System (INIS)

    Guerry-Force, M.L.; Perkett, E.A.; Brigham, K.L.; Meyrick, B.

    1988-01-01

    Using a large animal model of radiation lung injury--the sheep exposed to bilateral thoracic irradiation--we have recently shown the development of sustained pulmonary hypertension during the first 4 weeks following radiation. This is the period prior to the onset of pneumonitis and pulmonary fibrosis. In the present study, we have examined biopsy and autopsy lung tissue from these same sheep and assessed the sequential changes in lung morphology. Six unanesthetized sheep received bilateral thoracic irradiation (a total of 15 Gy); control sheep were sham irradiated. Lung biopsy tissue was taken prior to and at weekly or biweekly intervals during the 4 weeks immediately following radiation. The lungs were also removed at autopsy for light and electron microscopic examination. Our results show early (Week 1) interstitial and progressive intraalveolar edema accompanied by endothelial and epithelial injury. A gradual increase in number of interstitial mononuclear cells was evident from Week 1, both in the lung tissue and in perivascular cuffs. The number of peripheral lung interstitial mononuclear cells was twice baseline from Week 3 and included accumulation of lymphocytes, fibroblasts, and intravascular macrophages. The increased numbers of mononuclear cells paralleled the development of chronic pulmonary hypertension, perhaps suggesting their involvement in the pathogenesis of this disease. Alternatively, it may be that increased mononuclear cell number represents a stage of lung repair

  4. Multistep carcinogenesis of normal human fibroblasts. Human fibroblasts immortalized by repeated treatment with Co-60 gamma rays were transformed into tumorigenic cells with Ha-ras oncogenes.

    Science.gov (United States)

    Namba, M; Nishitani, K; Fukushima, F; Kimoto, T

    1988-01-01

    Two normal mortal human fibroblast cell strains were transformed into immortal cell lines, SUSM-1 and KMST-6, by treatment with 4-nitroquinoline 1-oxide (4NQO) and Co-60 gamma rays, respectively. These immortalized cell lines showed morphological changes of cells and remarkable chromosome aberrations, but neither of them grew in soft agar or formed tumors in nude mice. The immortal cell line, KMST-6, was then converted into neoplastic cells by treatment with Harvey murine sarcoma virus (Ha-MSV) or the c-Ha-ras oncogene derived from a human lung carcinoma. These neoplastically transformed cells acquired anchorage-independent growth potential and developed tumors when transplanted into nude mice. All the tumors grew progressively without regression until the animals died of tumors. In addition, the tumors were transplantable into other nude mice. Normal human fibroblasts, on the other hand, were not transformed into either immortal or tumorigenic cells by treatment with Ha-MSV or c-Ha-ras alone. Our present data indicate that (1) the chemical carcinogen, 4NQO, or gamma rays worked as an initiator of carcinogenesis in normal human cells, giving rise to immortality, and (2) the ras gene played a role in the progression of the immortally transformed cells to more malignant cells showing anchorage-independent growth and tumorigenicity. In other words, the immortalization process of human cells seems to be a pivotal or rate-limiting step in the carcinogenesis of human cells.

  5. Improved Fibroblast Functionalities by Microporous Pattern Fabricated by Microelectromechanical Systems

    OpenAIRE

    Wei, Hongbo; Zhao, Lingzhou; Chen, Bangdao; Bai, Shizhu; Zhao, Yimin

    2014-01-01

    Fibroblasts, which play an important role in biological seal formation and maintenance, determine the long-term success of percutaneous implants. In this study, well-defined microporous structures with micropore diameters of 10–60 µm were fabricated by microelectromechanical systems and their influence on the fibroblast functionalities was observed. The results show that the microporous structures with micropore diameters of 10–60 µm did not influence the initial adherent fibroblast number; ...

  6. Metastatic tumors of lungs

    International Nuclear Information System (INIS)

    Rozenshtraukh, L.C.; Rybakova, N.I.; Vinner, M.G.

    1987-01-01

    Roentgenologic semiotics of lung metastases and their complications, as well as peculiarities of lung metastases of separate localization tumours are presented. Definition table for primary tumour by roentgenologic aspect of lung metastases is given

  7. How Lungs Work

    Science.gov (United States)

    ... Diseases > How Lungs Work How Lungs Work The Respiratory System Your lungs are part of the respiratory system, ... your sense of smell. The Parts of the Respiratory System and How They Work Airways SINUSES are hollow ...

  8. Protecting Your Lungs

    Science.gov (United States)

    ... lung capacity. Specific breathing exercises can also help improve your lung function if you have certain lung diseases, like COPD. Exercise and breathing techniques are also great for improving your mood and helping you relax. Public Health and Your ...

  9. Urokinase-type plasminogen activator receptor (uPAR) ligation induces a raft-localized integrin signaling switch that mediates the hypermotile phenotype of fibrotic fibroblasts.

    Science.gov (United States)

    Grove, Lisa M; Southern, Brian D; Jin, Tong H; White, Kimberly E; Paruchuri, Sailaja; Harel, Efrat; Wei, Ying; Rahaman, Shaik O; Gladson, Candece L; Ding, Qiang; Craik, Charles S; Chapman, Harold A; Olman, Mitchell A

    2014-05-02

    The urokinase-type plasminogen activator receptor (uPAR) is a glycosylphosphatidylinositol-linked membrane protein with no cytosolic domain that localizes to lipid raft microdomains. Our laboratory and others have documented that lung fibroblasts from patients with idiopathic pulmonary fibrosis (IPF) exhibit a hypermotile phenotype. This study was undertaken to elucidate the molecular mechanism whereby uPAR ligation with its cognate ligand, urokinase, induces a motile phenotype in human lung fibroblasts. We found that uPAR ligation with the urokinase receptor binding domain (amino-terminal fragment) leads to enhanced migration of fibroblasts on fibronectin in a protease-independent, lipid raft-dependent manner. Ligation of uPAR with the amino-terminal fragment recruited α5β1 integrin and the acylated form of the Src family kinase, Fyn, to lipid rafts. The biological consequences of this translocation were an increase in fibroblast motility and a switch of the integrin-initiated signal pathway for migration away from the lipid raft-independent focal adhesion kinase pathway and toward a lipid raft-dependent caveolin-Fyn-Shc pathway. Furthermore, an integrin homologous peptide as well as an antibody that competes with β1 for uPAR binding have the ability to block this effect. In addition, its relative insensitivity to cholesterol depletion suggests that the interactions of α5β1 integrin and uPAR drive the translocation of α5β1 integrin-acylated Fyn signaling complexes into lipid rafts upon uPAR ligation through protein-protein interactions. This signal switch is a novel pathway leading to the hypermotile phenotype of IPF patient-derived fibroblasts, seen with uPAR ligation. This uPAR dependent, fibrotic matrix-selective, and profibrotic fibroblast phenotype may be amenable to targeted therapeutics designed to ameliorate IPF.

  10. Tumor-secreted LOXL2 activates fibroblasts through FAK signaling

    DEFF Research Database (Denmark)

    Barker, Holly E; Bird, Demelza; Lang, Georgina

    2013-01-01

    models. Here, we discovered that tumor-derived LOXL2 directly activated stromal fibroblasts in the tumor microenvironment. Genetic manipulation or antibody inhibition of LOXL2 in orthotopically grown mammary tumors reduced the expression of α-smooth muscle actin (α-SMA). Using a marker for reticular....... Importantly, in vitro assays revealed that tumor-derived LOXL2 and a recombinant LOXL2 protein induced fibroblast branching on collagen matrices, as well as increased fibroblast-mediated collagen contraction and invasion of fibroblasts through extracellular matrix. Moreover, LOXL2 induced the expression of α...

  11. In vivo excision of pyrimidine dimers is mediated by a DNA N-glycosylase in Micrococcus luteus but not in human fibroblasts

    International Nuclear Information System (INIS)

    La Belle, M.; Linn, S.

    1982-01-01

    It has been previously shown that Micrococcus luteus possesses a pyrimidine dimer-specific endonuclease which in vitro, functions as both an endonuclease and DNA-glycosylase. To determine if these combined activities function in vivo, the excision products of UV-irradiated M. luteus were isolated and examined. In addition, a procedure was devised to isolate and examine the excision products from UV-irradiated human fibroblasts to determine if an endonuclease/glycosylase activity functions in the excision of UV-induced pyrimidine dimers in human fibroblasts. It was shown that, in vivo, an endonuclease/glycosylase mechanism is utilized extensively in the repair of pyrimidine dimers by M. luteus, but that human fibroblasts do not appear to use this mechanism. (author)

  12. In vivo excision of pyrimidine dimers is mediated by a DNA N-glycosylase in Micrococcus luteus but not in human fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    La Belle, M; Linn, S [California Univ., Berkeley (USA). Dept. of Biochemistry

    1982-09-01

    It has been previously shown that Micrococcus luteus possesses a pyrimidine dimer-specific endonuclease which in vitro, functions as both an endonuclease and DNA-glycosylase. To determine if these combined activities function in vivo, the excision products of UV-irradiated M. luteus were isolated and examined. In addition, a procedure was devised to isolate and examine the excision products from UV-irradiated human fibroblasts to determine if an endonuclease/glycosylase activity functions in the excision of UV-induced pyrimidine dimers in human fibroblasts. It was shown that, in vivo, an endonuclease/glycosylase mechanism is utilized extensively in the repair of pyrimidine dimers by M. luteus, but that human fibroblasts do not appear to use this mechanism.

  13. Neuronal protein gene product 9.5 (IEF SSP 6104) is expressed in cultured human MRC-5 fibroblasts of normal origin and is strongly down-regulated in their SV40 transformed counterparts

    DEFF Research Database (Denmark)

    Honoré, B; Rasmussen, H H; Vandekerckhove, J

    1991-01-01

    of proteins recovered from 2D gels we have identified PGP 9.5 UCH-L1 as polypeptide IEF SSP 6104 (Mr = 27,000, pI = 5.49) in the comprehensive 2D gel cellular protein database of human embryonal lung MRC-5 fibroblasts [(1989) Electrophoresis 10, 76 115; (1990) Electrophoresis 11, 1072 1113]. This protein...

  14. Single-domain antibodies that compete with the natural ligand fibroblast growth factor block the internalization of the fibroblast growth factor receptor 1

    Energy Technology Data Exchange (ETDEWEB)

    Veggiani, Gianluca; Ossolengo, Giuseppe; Aliprandi, Marisa; Cavallaro, Ugo [IFOM-IEO Campus, Via Adamello 16, 20139 Milano (Italy); Marco, Ario de, E-mail: ario.demarco@ung.si [IFOM-IEO Campus, Via Adamello 16, 20139 Milano (Italy); Dept. Environmental Sciences, University of Nova Gorica (UNG), Vipavska 13, P.O. Box 301-SI-5000, Rozna Dolina, Nova Gorica (Slovenia)

    2011-05-20

    Highlights: {yields} Recombinant antibodies for FGFR1 were isolated from a llama naive library in VHH format. {yields} These antibodies compete with the natural ligand FGF-2 for the same epitope on FGFR1. {yields} The antibody competition inhibits the FGF-2-dependent internalization of FGFR1. -- Abstract: Single-domain antibodies in VHH format specific for fibroblast growth factor receptor 1 (FGFR1) were isolated from a phage-display llama naive library. In particular, phage elution in the presence of the natural receptor ligand fibroblast growth factor (FGF) allowed for the identification of recombinant antibodies that compete with FGF for the same region on the receptor surface. These antibodies posses a relatively low affinity for FGFR1 and were never identified when unspecific elution conditions favoring highly affine binders were applied to panning procedures. Two populations of competitive antibodies were identified that labeled specifically the receptor-expressing cells in immunofluorescence and recognize distinct epitopes. Antibodies from both populations effectively prevented FGF-dependent internalization and nuclear accumulation of the receptor in cultured cells. This achievement indicates that these antibodies have a capacity to modulate the receptor physiology and, therefore, constitute powerful reagents for basic research and a potential lead for therapeutic applications.

  15. Single-domain antibodies that compete with the natural ligand fibroblast growth factor block the internalization of the fibroblast growth factor receptor 1

    International Nuclear Information System (INIS)

    Veggiani, Gianluca; Ossolengo, Giuseppe; Aliprandi, Marisa; Cavallaro, Ugo; Marco, Ario de

    2011-01-01

    Highlights: → Recombinant antibodies for FGFR1 were isolated from a llama naive library in VHH format. → These antibodies compete with the natural ligand FGF-2 for the same epitope on FGFR1. → The antibody competition inhibits the FGF-2-dependent internalization of FGFR1. -- Abstract: Single-domain antibodies in VHH format specific for fibroblast growth factor receptor 1 (FGFR1) were isolated from a phage-display llama naive library. In particular, phage elution in the presence of the natural receptor ligand fibroblast growth factor (FGF) allowed for the identification of recombinant antibodies that compete with FGF for the same region on the receptor surface. These antibodies posses a relatively low affinity for FGFR1 and were never identified when unspecific elution conditions favoring highly affine binders were applied to panning procedures. Two populations of competitive antibodies were identified that labeled specifically the receptor-expressing cells in immunofluorescence and recognize distinct epitopes. Antibodies from both populations effectively prevented FGF-dependent internalization and nuclear accumulation of the receptor in cultured cells. This achievement indicates that these antibodies have a capacity to modulate the receptor physiology and, therefore, constitute powerful reagents for basic research and a potential lead for therapeutic applications.

  16. Immortalization of Werner syndrome and progeria fibroblasts

    International Nuclear Information System (INIS)

    Saito, H.; Moses, R.E.

    1991-01-01

    Human fibroblast cells from two different progeroid syndromes, Werner syndrome (WS) and progeria, were established as immortalized cell lines by transfection with plasmid DNA containing the SV40 early region. The lineage of each immortalized cell line was confirmed by VNTR analysis. Each of the immortalized cell lines maintained its original phenotype of slow growth. DNA repair ability of these cells was also studied by measuring sensitivity to killing by uv or the DNA-damaging drugs methyl methansulfonate, bleomycin, and cis-dichlorodiamine platinum. The results showed that both WS and progeria cells have normal sensitivity to these agents

  17. Coupling of cytoskeleton functions for fibroblast locomotion

    DEFF Research Database (Denmark)

    Couchman, J R; Lenn, M; Rees, D A

    1985-01-01

    caused visible protrusions in projected positions at the leading edge. We conclude that fibroblast locomotion may be driven coordinately by a common set of motility mechanisms and that this coordination may be lost as a result of physical or pharmacological disturbance. Taking our evidence with results...... from other Laboratories, we propose the following cytoskeleton functions. (i) Protrusive activity, probably based on solation--gelation cycles of the actin based cytoskeleton and membrane recycling which provides cellular and membrane components for streaming through the cell body to the leading edge...

  18. Differences in motility pattern between human buccal fibroblasts and periodontal and skin fibroblasts

    DEFF Research Database (Denmark)

    Lepekhin, Eugene; Grøn, Birgitte; Berezin, Vladimir

    2002-01-01

    at these sites can be explained by differences in the motile behavior of their respective fibroblast populations. The migratory characteristics were studied in a two-dimensional culture system. The migration of single cells was time-lapse video recorded at intervals of 15 min for a period of 6 h using a computer...

  19. Lung PET scan

    Science.gov (United States)

    ... Chest PET scan; Lung positron emission tomography; PET - chest; PET - lung; PET - tumor imaging; ... Grainger & Allison's Diagnostic Radiology: A Textbook of Medical Imaging . 6th ed. Philadelphia, ...

  20. Extravascular Lung Water and Acute Lung Injury

    Directory of Open Access Journals (Sweden)

    Ritesh Maharaj

    2012-01-01

    Full Text Available Acute lung injury carries a high burden of morbidity and mortality and is characterised by nonhydrostatic pulmonary oedema. The aim of this paper is to highlight the role of accurate quantification of extravascular lung water in diagnosis, management, and prognosis in “acute lung injury” and “acute respiratory distress syndrome”. Several studies have verified the accuracy of both the single and the double transpulmonary thermal indicator techniques. Both experimental and clinical studies were searched in PUBMED using the term “extravascular lung water” and “acute lung injury”. Extravascular lung water measurement offers information not otherwise available by other methods such as chest radiography, arterial blood gas, and chest auscultation at the bedside. Recent data have highlighted the role of extravascular lung water in response to treatment to guide fluid therapy and ventilator strategies. The quantification of extravascular lung water may predict mortality and multiorgan dysfunction. The limitations of the dilution method are also discussed.

  1. Replacement of murine fibroblasts by human fibroblasts irradiated in obtaining feeder layer for the culture of human keratinocytes

    International Nuclear Information System (INIS)

    Yoshito, Daniele; Sufi, Bianca S.; Santin, Stefany P.; Mathor, Monica B.; Altran, Silvana C.; Isaac, Cesar

    2011-01-01

    Human autologous epithelia cultivated in vitro, have been used successfully in treating damage to skin integrity. The methodology allowed the cultivation of these epithelia was described by Rheinwald and Green in 1975, this methodology consisted in seeding keratinocytes onto a feeder layer composed of lineage 3T3 murine fibroblasts, the proliferation rate is controlled through the action of ionizing radiation. However, currently there is a growing concern about the possibility of transmitting prions and murine viruses to transplanted patients. Taking into account this concern, in this present work, we replaced the feeder layer originally composed of murine fibroblasts by human fibroblasts. To obtain this new feeder layer was necessary to standardize the enough irradiation dose to inhibit the replication of human fibroblasts and the verification of effectiveness of the development of keratinocytes culture on a feeder layer thus obtained. According to the obtained results we can verify that the human fibroblasts irradiated at various tested doses (60, 70, 100, 200, 250 and 300 Gy) had their mitotic activity inactivated by irradiation, allowing the use of any of these doses to confection of the feeder layer, since these fibroblasts irradiated still showed viable until fourteen days of cultivation. In the test of colony formation efficiency was observed that keratinocytes seeded on irradiated human fibroblasts were able to develop satisfactorily, preserving their clonogenic potential. Therefore it was possible the replacement of murine fibroblasts by human fibroblasts in confection of the feeder layer, in order to eliminate this xenobiotic component of the keratinocytes culture. (author)

  2. Replacement of murine fibroblasts by human fibroblasts irradiated in obtaining feeder layer for the culture of human keratinocytes

    Energy Technology Data Exchange (ETDEWEB)

    Yoshito, Daniele; Sufi, Bianca S.; Santin, Stefany P.; Mathor, Monica B. [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil); Altran, Silvana C.; Isaac, Cesar [Universidade Sao Paulo (USP), Sao Paulo, SP (Brazil). Fac. de Medicina. Lab. de Microcirurgia Plastica; Esteves-Pedro, Natalia M. [Universidade Sao Paulo (USP), Sao Paulo, SP (Brazil). Fac. de Ciencias Farmaceuticas. Lab. de Controle Biologico; Herson, Marisa R. [DonorTissue Bank of Victoria (Australia)

    2011-07-01

    Human autologous epithelia cultivated in vitro, have been used successfully in treating damage to skin integrity. The methodology allowed the cultivation of these epithelia was described by Rheinwald and Green in 1975, this methodology consisted in seeding keratinocytes onto a feeder layer composed of lineage 3T3 murine fibroblasts, the proliferation rate is controlled through the action of ionizing radiation. However, currently there is a growing concern about the possibility of transmitting prions and murine viruses to transplanted patients. Taking into account this concern, in this present work, we replaced the feeder layer originally composed of murine fibroblasts by human fibroblasts. To obtain this new feeder layer was necessary to standardize the enough irradiation dose to inhibit the replication of human fibroblasts and the verification of effectiveness of the development of keratinocytes culture on a feeder layer thus obtained. According to the obtained results we can verify that the human fibroblasts irradiated at various tested doses (60, 70, 100, 200, 250 and 300 Gy) had their mitotic activity inactivated by irradiation, allowing the use of any of these doses to confection of the feeder layer, since these fibroblasts irradiated still showed viable until fourteen days of cultivation. In the test of colony formation efficiency was observed that keratinocytes seeded on irradiated human fibroblasts were able to develop satisfactorily, preserving their clonogenic potential. Therefore it was possible the replacement of murine fibroblasts by human fibroblasts in confection of the feeder layer, in order to eliminate this xenobiotic component of the keratinocytes culture. (author)

  3. Toll-like receptor 9 mediated responses in cardiac fibroblasts.

    Directory of Open Access Journals (Sweden)

    Ingrid Kristine Ohm

    Full Text Available Altered cardiac Toll-like receptor 9 (TLR9 signaling is important in several experimental cardiovascular disorders. These studies have predominantly focused on cardiac myocytes or the heart as a whole. Cardiac fibroblasts have recently been attributed increasing significance in mediating inflammatory signaling. However, putative TLR9-signaling through cardiac fibroblasts remains non-investigated. Thus, our aim was to explore TLR9-signaling in cardiac fibroblasts and investigate the consequence of such receptor activity on classical cardiac fibroblast cellular functions. Cultivated murine cardiac fibroblasts were stimulated with different TLR9 agonists (CpG A, B and C and assayed for the secretion of inflammatory cytokines (tumor necrosis factor α [TNFα], CXCL2 and interferon α/β. Expression of functional cardiac fibroblast TLR9 was proven as stimulation with CpG B and -C caused significant CXCL2 and TNFα-release. These responses were TLR9-specific as complete inhibition of receptor-stimulated responses was achieved by co-treatment with a TLR9-antagonist (ODN 2088 or chloroquine diphosphate. TLR9-stimulated responses were also found more potent in cardiac fibroblasts when compared with classical innate immune cells. Stimulation of cardiac fibroblasts TLR9 was also found to attenuate migration and proliferation, but did not influence myofibroblast differentiation in vitro. Finally, results from in vivo TLR9-stimulation with subsequent fractionation of specific cardiac cell-types (cardiac myocytes, CD45+ cells, CD31+ cells and cardiac fibroblast-enriched cell-fractions corroborated our in vitro data and provided evidence of differentiated cell-specific cardiac responses. Thus, we conclude that cardiac fibroblast may constitute a significant TLR9 responder cell within the myocardium and, further, that such receptor activity may impact important cardiac fibroblast cellular functions.

  4. Stochastic variation in telomere shortening rate causes heterogeneity of human fibroblast replicative life span.

    Science.gov (United States)

    Martin-Ruiz, Carmen; Saretzki, Gabriele; Petrie, Joanne; Ladhoff, Juliane; Jeyapalan, Jessie; Wei, Wenyi; Sedivy, John; von Zglinicki, Thomas

    2004-04-23

    The replicative life span of human fibroblasts is heterogeneous, with a fraction of cells senescing at every population doubling. To find out whether this heterogeneity is due to premature senescence, i.e. driven by a nontelomeric mechanism, fibroblasts with a senescent phenotype were isolated from growing cultures and clones by flow cytometry. These senescent cells had shorter telomeres than their cycling counterparts at all population doubling levels and both in mass cultures and in individual subclones, indicating heterogeneity in the rate of telomere shortening. Ectopic expression of telomerase stabilized telomere length in the majority of cells and rescued them from early senescence, suggesting a causal role of telomere shortening. Under standard cell culture conditions, there was a minor fraction of cells that showed a senescent phenotype and short telomeres despite active telomerase. This fraction increased under chronic mild oxidative stress, which is known to accelerate telomere shortening. It is possible that even high telomerase activity cannot fully compensate for telomere shortening in all cells. The data show that heterogeneity of the human fibroblast replicative life span can be caused by significant stochastic cell-to-cell variation in telomere shortening.

  5. Potentially lethal damage repair in cell lines of radioresistant human tumours and normal skin fibroblasts

    International Nuclear Information System (INIS)

    Marchese, M.J.; Minarik, L.; Hall, E.J.; Zaider, M.

    1985-01-01

    Radiation cell survival data were obtained in vitro for three cell lines isolated from human tumours traditionally considered to be radioresistant-two melanomas and one osteosarcoma-as well as from a diploid skin fibroblast cell line. One melanoma cell line was much more radioresistant than the other, while the osteosarcoma and fibroblast cell lines were more radiosensitive than either. For cells growing exponentially, little potentially lethal damage repair (PLDR) could be demonstrated by comparing survival data for cells in which subculture was delayed by 6 h with those sub-cultured immediately after treatment. For the malignant cells in plateau phase, which in these cells might be better termed 'slowed growth phase', since an appreciable fraction of the cells are still cycling, a small amount of PLDR was observed, but not as much as reported by other investigators in the literature. The normal fibroblasts, which achieved a truer plateau phase in terms of noncycling cells, showed a significantly larger amount of PLDR than the tumour cells. (author)

  6. L-Lactate Protects Skin Fibroblasts against Aging-Associated Mitochondrial Dysfunction via Mitohormesis

    Directory of Open Access Journals (Sweden)

    Jaroslav Zelenka

    2015-01-01

    Full Text Available A moderate elevation of reactive oxygen species (ROS production and a mild inhibition of mitochondrial respiratory chain have been associated with a health promotion and a lifespan extension in several animal models of aging. Here, we tested whether this phenomenon called mitohormesis could be mediated by L-lactate. The treatment with 5 mM L-lactate significantly increased H2O2 production and slightly inhibited the respiration in cultured skin fibroblasts and in isolated mitochondria. The L-lactate exposure was associated with oxidation of intracellular glutathione, phosphorylation of 5′AMP-activated protein kinase (AMPK, and induction of peroxisome proliferator-activated receptor gamma coactivator 1α (PGC1α transcription. A replicative aging of fibroblasts (L0 with a constant (LC, or intermittent 5 mM L-lactate (LI in media showed that the high-passage LI fibroblasts have higher respiration, lower H2O2 release, and lower secretion of L-lactate compared to L0 and LC. This protection against mitochondrial dysfunction in LI cells was associated with lower activity of mechanistic target of rapamycin complex 1 (mTORC1, less signs of cellular senescence, and increased autophagy compared to L0 and LC. In conclusion, we demonstrated that intermittent but not constant exposure to L-lactate triggers mitohormesis, prevents aging-associated mitochondrial dysfunction, and improves other markers of aging.

  7. Fibroblastic connective tissue nevus: Clinicopathological and immunohistochemical study of 14 cases.

    Science.gov (United States)

    Pennacchia, Ilaria; Kutzner, Heinz; Kazakov, Dmitry V; Mentzel, Thomas

    2017-10-01

    We present herein a series of 14 lesions showing overlapping features with the newly defined benign cutaneous mesenchymal neoplasm labeled as fibroblastic connective tissue nevus (FCTN). Total of 8 patients were male and 5 were female, ranging in age from 1 to 56 years. Lesions appeared as isolated nodules or plaques on the trunk (7 cases), the limbs (4 cases) and the neck (2 cases). Histologically, all cases were composed of bundles of bland spindle cells of fibroblastic/myofibroblastic lineage irregularly branching within the reticular dermis and along fibrous septa in the subcutis. Adnexal structures and dermal adipocytes were entrapped by the fascicles, the epidermis was often papillomatous and elastic fibers were decreased and fragmented. Expression of CD34 and ASMA was found in 8 and 7 cases, respectively. Follow-up was available for 7 patients (mean follow-up, 5 years; range, 1-10 years). None of the cases metastasized or recurred, even when incompletely excised. The differential diagnosis of FCTN is broad and includes hypertrophic scar, dermatofibroma, dermatomyofibroma, pilar leiomyoma, plaque-stage DFSP, CD34-positive plaque-like dermal fibroma, fibroblastic-predominant plexiform fibrohistiocytic tumor, lipofibromatosis, superficial desmoid fibromatosis and fibrous hamartoma of infancy, of which it represents probably the monophasic variant. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  8. Determining the pharmacological activity of Physalis peruviana fruit juice on rabbit eyes and fibroblast primary cultures.

    Science.gov (United States)

    Pardo, Juan Manuel; Fontanilla, Marta Raquel; Ospina, Luis Fernando; Espinosa, Lady

    2008-07-01

    The pharmacologic activity of compounds isolated from Physalis peruviana has been demonstrated. The use of this fruit juice for treating pterygium has been reported in Colombian traditional medicine. However, studies demonstrating the fruit juice's pharmacologic activity when used in this disease have not been published to date. In the present study the anti-inflammatory and cytostatic activities of P. peruviana fruit juice in a rabbit eye inflammatory model were investigated. A novel rabbit eye inflammation model was developed for studying the juice's anti-inflammatory activity (based on an adaptation of the Draize test). Cytostatic activity was evaluated by measuring and comparing growth rates of cultured fibroblasts exposed and not exposed to various fruit juice concentrations. P. peruviana fruit juice exhibited a mild anti-inflammatory activity compared with methylprednisolone, a known anti-inflammatory drug. An interesting dose-dependent cytostatic effect on cultured fibroblasts was also established. The data found suggest that the P. peruviana fruit juice anti-pterygium effect described in traditional medicine may be related to its inhibiting fibroblast growth. The present study contributes to the pharmacologic knowledge regarding a remedy commonly used in Colombian traditional medicine.

  9. Establishment of the first humpback whale fibroblast cell lines and their application in chemical risk assessment

    Energy Technology Data Exchange (ETDEWEB)

    Burkard, Michael, E-mail: Michael.burkard@eawag.ch [Griffith University, Environmental Futures Research Institute, Southern Ocean Persistent Organic Pollutants Program, Brisbane, QLD (Australia); Eawag, Swiss Federal Institute of Technology, Dübendorf (Switzerland); Whitworth, Deanne [The University of Queensland, School of Veterinary Science, Gatton, QLD (Australia); Schirmer, Kristin [Eawag, Swiss Federal Institute of Technology, Dübendorf (Switzerland); ETH Zürich, Institute of Biogechemistry and Pollutant Dynamics, Zürich (Switzerland); EPF Lausanne, School of Architecture, Civil and Environmental Engineering, Lausanne (Switzerland); Nash, Susan Bengtson [Griffith University, Environmental Futures Research Institute, Southern Ocean Persistent Organic Pollutants Program, Brisbane, QLD (Australia)

    2015-10-15

    Highlights: • We established and characterised the first humpback whale fibroblast cell lines. • Cell lines have a stable karyotype with 2n = 44. • Exposure to p,p′-DDE resulted in a concentration-dependent loss of cell viability. • p,p′-DDE sensitivity differed considerably from human fibroblasts. • Exposure to a whale blubber extract showed higher sensitivity than to p,p′-DDE alone. - Abstract: This paper reports the first successful derivation and characterization of humpback whale fibroblast cell lines. Primary fibroblasts were isolated from the dermal connective tissue of skin biopsies, cultured at 37 °C and 5% CO{sub 2} in the standard mammalian medium DMEM/F12 supplemented with 10% fetal bovine serum (FBS). Of nine initial biopsies, two cell lines were established from two different animals and designated HuWa1 and HuWa2. The cells have a stable karyotype with 2n = 44, which has commonly been observed in other baleen whale species. Cells were verified as being fibroblasts based on their spindle-shaped morphology, adherence to plastic and positive immunoreaction to vimentin. Population doubling time was determined to be ∼41 h and cells were successfully cryopreserved and thawed. To date, HuWa1 cells have been propagated 30 times. Cells proliferate at the tested temperatures, 30, 33.5 and 37 °C, but show the highest rate of proliferation at 37 °C. Short-term exposure to para,para′-dichlorodiphenyldichloroethylene (p,p′-DDE), a priority compound accumulating in southern hemisphere humpback whales, resulted in a concentration-dependent loss of cell viability. The effective concentration which caused a 50% reduction in HuWa1 cell viability (EC{sub 50} value) was approximately six times greater than the EC{sub 50} value for the same chemical measured with human dermal fibroblasts. HuWa1 exposed to a natural, p,p′-DDE-containing, chemical mixture extracted from whale blubber showed distinctively higher sensitivity than to p,p′-DDE alone

  10. Establishment of the first humpback whale fibroblast cell lines and their application in chemical risk assessment

    International Nuclear Information System (INIS)

    Burkard, Michael; Whitworth, Deanne; Schirmer, Kristin; Nash, Susan Bengtson

    2015-01-01

    Highlights: • We established and characterised the first humpback whale fibroblast cell lines. • Cell lines have a stable karyotype with 2n = 44. • Exposure to p,p′-DDE resulted in a concentration-dependent loss of cell viability. • p,p′-DDE sensitivity differed considerably from human fibroblasts. • Exposure to a whale blubber extract showed higher sensitivity than to p,p′-DDE alone. - Abstract: This paper reports the first successful derivation and characterization of humpback whale fibroblast cell lines. Primary fibroblasts were isolated from the dermal connective tissue of skin biopsies, cultured at 37 °C and 5% CO_2 in the standard mammalian medium DMEM/F12 supplemented with 10% fetal bovine serum (FBS). Of nine initial biopsies, two cell lines were established from two different animals and designated HuWa1 and HuWa2. The cells have a stable karyotype with 2n = 44, which has commonly been observed in other baleen whale species. Cells were verified as being fibroblasts based on their spindle-shaped morphology, adherence to plastic and positive immunoreaction to vimentin. Population doubling time was determined to be ∼41 h and cells were successfully cryopreserved and thawed. To date, HuWa1 cells have been propagated 30 times. Cells proliferate at the tested temperatures, 30, 33.5 and 37 °C, but show the highest rate of proliferation at 37 °C. Short-term exposure to para,para′-dichlorodiphenyldichloroethylene (p,p′-DDE), a priority compound accumulating in southern hemisphere humpback whales, resulted in a concentration-dependent loss of cell viability. The effective concentration which caused a 50% reduction in HuWa1 cell viability (EC_5_0 value) was approximately six times greater than the EC_5_0 value for the same chemical measured with human dermal fibroblasts. HuWa1 exposed to a natural, p,p′-DDE-containing, chemical mixture extracted from whale blubber showed distinctively higher sensitivity than to p,p′-DDE alone. Thus, we

  11. Pirfenidone inhibits the proliferation of fibroblasts from patients with active Crohn's disease.

    Science.gov (United States)

    Kadir, Sara-Irini; Wenzel Kragstrup, Tue; Dige, Anders; Kok Jensen, Simon; Dahlerup, Jens Frederik; Kelsen, Jens

    2016-11-01

    One-third of Crohn's disease (CD) patients develop intestinal strictures that require repeated surgical intervention. Current anti-inflammatory therapies have limited effect on stricture development, which necessitates the exploration of new pharmacological approaches. Pirfenidone (PFD), a novel anti-fibrotic agent, was recently approved in Europe for the treatment of idiopathic pulmonary fibrosis (IPF). We hypothesized that observations in IPF could be transferable to intestinal fibrosis and that PFD inhibits the proliferation and extracellular matrix (ECM) turnover of gut-derived fibroblasts from CD patients. Fibroblasts were isolated from biopsies of inflamed (n = 8) and non-inflamed (n = 5) colonic mucosa. Expression of CD90 and alpha-smooth muscle actin (αSMA) expression was determined by flow cytometry. The fibroblasts were cultured with PFD (0.5, 1.0 and 2.0 mg/ml). Proliferation was evaluated with CellTiter 96(®) AQueous One Solution Cell Proliferation Assay. Production of matrix metalloproteinase-3 (MMP-3), tissue inhibitor of metalloproteinases-1 (TIMP-1) and collagen were assessed using ELISA and calorimetric assays, respectively. The majority of the fibroblasts were αSMA-positive myofibroblasts. PFD inhibited fibroblast proliferation [0.94 (PFD 0.5 mg/ml); 0.76 (1.0 mg/ml); 0.58 (2.0 mg/ml)] and production of MMP-3 [0.85 (0.5 mg/ml); 0.74 (1.0 mg/ml); 0.63 (2.0 mg/ml)] dose-dependently (both p = 0.0001). The anti-proliferative effect of PFD was reversible (p = 0.0001), indicating that PFD does not act by an irreversible cytotoxic mechanism. PFD did not influence neither TIMP-1 nor collagen production. PFD inhibited the proliferation and the production of MMP-3 dose-dependently in gut-derived fibroblast from CD patients. Our observations support further studies on PFD in stricturing CD.

  12. Role of Matrix Metalloproteinases-1 and -2 in Interleukin-13–Suppressed Elastin in Airway Fibroblasts in Asthma

    Science.gov (United States)

    Slade, David; Church, Tony D.; Francisco, Dave; Heck, Karissa; Sigmon, R. Wesley; Ghio, Michael; Murillo, Anays; Firszt, Rafael; Lugogo, Njira L.; Que, Loretta; Sunday, Mary E.; Kraft, Monica

    2016-01-01

    Elastin synthesis and degradation in the airway and lung parenchyma contribute to airway mechanics, including airway patency and elastic recoil. IL-13 mediates many features of asthma pathobiology, including airway remodeling, but the effects of IL-13 on elastin architecture in the airway wall are not known. We hypothesized that IL-13 modulates elastin expression in airway fibroblasts from subjects with allergic asthma. Twenty-five subjects with mild asthma (FEV1, 89 ± 3% predicted) and 30 normal control subjects (FEV1, 102 ± 2% predicted) underwent bronchoscopy with endobronchial biopsy. Elastic fibers were visualized in airway biopsy specimens using Weigert’s resorcin-fuchsin elastic stain. Airway fibroblasts were exposed to IL-13; a pan-matrix metalloproteinase (MMP) inhibitor (GM6001); specific inhibitors to MMP-1, -2, -3, and -8; and combinations of IL-13 with MMP inhibitors in separate conditions in serum-free media for 48 hours. Elastin (ELN) expression as well as MMP secretion and activity were quantified. Results of this study show that elastic fiber staining of airway biopsy tissue was significantly associated with methacholine PC20 (i.e., the provocative concentration of methacholine resulting in a 20% fall in FEV1 levels) in patients with asthma. IL-13 significantly suppressed ELN expression in asthmatic airway fibroblasts as compared with normal control fibroblasts. The effect of IL-13 on ELN expression was significantly correlated with postbronchodilator FEV1/FVC in patients with asthma. MMP inhibition significantly stimulated ELN expression in patients with asthma as compared with normal control subjects. Specific inhibition of MMP-1 and MMP-2, but not MMP-3 or MMP-8, reversed the IL-13–induced suppression of ELN expression. In asthma, MMP-1 and MMP-2 mediate IL-13–induced suppression of ELN expression in airway fibroblasts. PMID:26074138

  13. Role of Matrix Metalloproteinases-1 and -2 in Interleukin-13-Suppressed Elastin in Airway Fibroblasts in Asthma.

    Science.gov (United States)

    Ingram, Jennifer L; Slade, David; Church, Tony D; Francisco, Dave; Heck, Karissa; Sigmon, R Wesley; Ghio, Michael; Murillo, Anays; Firszt, Rafael; Lugogo, Njira L; Que, Loretta; Sunday, Mary E; Kraft, Monica

    2016-01-01

    Elastin synthesis and degradation in the airway and lung parenchyma contribute to airway mechanics, including airway patency and elastic recoil. IL-13 mediates many features of asthma pathobiology, including airway remodeling, but the effects of IL-13 on elastin architecture in the airway wall are not known. We hypothesized that IL-13 modulates elastin expression in airway fibroblasts from subjects with allergic asthma. Twenty-five subjects with mild asthma (FEV1, 89 ± 3% predicted) and 30 normal control subjects (FEV1, 102 ± 2% predicted) underwent bronchoscopy with endobronchial biopsy. Elastic fibers were visualized in airway biopsy specimens using Weigert's resorcin-fuchsin elastic stain. Airway fibroblasts were exposed to IL-13; a pan-matrix metalloproteinase (MMP) inhibitor (GM6001); specific inhibitors to MMP-1, -2, -3, and -8; and combinations of IL-13 with MMP inhibitors in separate conditions in serum-free media for 48 hours. Elastin (ELN) expression as well as MMP secretion and activity were quantified. Results of this study show that elastic fiber staining of airway biopsy tissue was significantly associated with methacholine PC20 (i.e., the provocative concentration of methacholine resulting in a 20% fall in FEV1 levels) in patients with asthma. IL-13 significantly suppressed ELN expression in asthmatic airway fibroblasts as compared with normal control fibroblasts. The effect of IL-13 on ELN expression was significantly correlated with postbronchodilator FEV1/FVC in patients with asthma. MMP inhibition significantly stimulated ELN expression in patients with asthma as compared with normal control subjects. Specific inhibition of MMP-1 and MMP-2, but not MMP-3 or MMP-8, reversed the IL-13-induced suppression of ELN expression. In asthma, MMP-1 and MMP-2 mediate IL-13-induced suppression of ELN expression in airway fibroblasts.

  14. Pharmaceutical studies for gene therapy: expression of human Cu, Zn-superoxide dismutase gene transfected by lipofection in rat skin fibroblasts.

    Science.gov (United States)

    Nishiguchi, K; Ishida, K; Nakajima, M; Maeda, T; Komada, F; Iwakawa, S; Tanigawara, Y; Okumura, K

    1996-08-01

    To evaluate whether lipofection using Lipofectin is suitable for delivering foreign genes into skin fibroblasts as target cells, we performed experiments using human superoxide dismutase (hSOD) and neomycin-resistance (Neo) genes as models in rat skin fibroblasts (FR and primary cells) in vitro. The amounts of DNA used in the lipofection procedure significantly affected the transfection efficiencies, and the optimal amounts were determined for all cells used. However, the efficiencies in rat skin fibroblasts were about 20-fold higher than that in rat lung epithelial-like cells (L2 cells). The differences in plasmid vectors (pRc/RSV-SOD and pRc/CMV-SOD) hardly affected the transfection efficiencies. The amounts of Lipofectin significantly affected the transfection efficiencies, and the optimal amounts were determined for both types of skin fibroblasts. However, cytotoxic effects in both skin fibroblasts were observed with high doses of Lipofectin. On the other hand, with optimal amounts of DNA and Lipofectin, the reporter gene (NeoT) introduced into cells was mainly integrated into the host cell chromosome. Western blot analysis showed the continuous expression of hSOD protein for at least 45 d in skin fibroblasts transfected with the expression plasmid for hSOD by Lipofectin under the optimal conditions, and the cellular SOD activity fluctuated in parallel with the expression of hSOD protein. Differences in the type of cells also affected the expression of hSOD. These results indicate that it is necessary to set up optimal conditions for transfection using Lipofectin for each cell type, and that transfection with Lipofectin under optimal conditions may be an efficient method for introduction of foreign genes into skin fibroblasts for use as a clinical delivery system of therapeutic protein.

  15. Degranulating mast cells in fibrotic regions of human tumors and evidence that mast cell heparin interferes with the growth of tumor cells through a mechanism involving fibroblasts

    International Nuclear Information System (INIS)

    Samoszuk, Michael; Kanakubo, Emi; Chan, John K

    2005-01-01

    The purpose of this study was to test the hypothesis that mast cells that are present in fibrotic regions of cancer can suppress the growth of tumor cells through an indirect mechanism involving peri-tumoral fibroblasts. We first immunostained a wide variety of human cancers for the presence of degranulated mast cells. In a subsequent series of controlled in vitro experiments, we then co-cultured UACC-812 human breast cancer cells with normal fibroblasts in the presence or absence of different combinations and doses of mast cell tryptase, mast cell heparin, a lysate of the human mast cell line HMC-1, and fibroblast growth factor-7 (FGF-7), a powerful, heparin-binding growth factor for breast epithelial cells. Degranulating mast cells were localized predominantly in the fibrous tissue of every case of breast cancer, head and neck cancer, lung cancer, ovarian cancer, non-Hodgkin's lymphoma, and Hodgkin's disease that we examined. Mast cell tryptase and HMC-1 lysate had no significant effect on the clonogenic growth of cancer cells co-cultured with fibroblasts. By contrast, mast cell heparin at multiple doses significantly reduced the size and number of colonies of tumor cells co-cultured with fibroblasts, especially in the presence of FGF-7. Neither heparin nor FGF-7, individually or in combination, produced any significant effect on the clonogenic growth of breast cancer cells cultured without fibroblasts. Degranulating mast cells are restricted to peri-tumoral fibrous tissue, and mast cell heparin is a powerful inhibitor of clonogenic growth of tumor cells co-cultured with fibroblasts. These results may help to explain the well-known ability of heparin to inhibit the growth of primary and metastatic tumors

  16. Degranulating mast cells in fibrotic regions of human tumors and evidence that mast cell heparin interferes with the growth of tumor cells through a mechanism involving fibroblasts

    Directory of Open Access Journals (Sweden)

    Kanakubo Emi

    2005-09-01

    Full Text Available Abstract Background The purpose of this study was to test the hypothesis that mast cells that are present in fibrotic regions of cancer can suppress the growth of tumor cells through an indirect mechanism involving peri-tumoral fibroblasts. Methods We first immunostained a wide variety of human cancers for the presence of degranulated mast cells. In a subsequent series of controlled in vitro experiments, we then co-cultured UACC-812 human breast cancer cells with normal fibroblasts in the presence or absence of different combinations and doses of mast cell tryptase, mast cell heparin, a lysate of the human mast cell line HMC-1, and fibroblast growth factor-7 (FGF-7, a powerful, heparin-binding growth factor for breast epithelial cells. Results Degranulating mast cells were localized predominantly in the fibrous tissue of every case of breast cancer, head and neck cancer, lung cancer, ovarian cancer, non-Hodgkin's lymphoma, and Hodgkin's disease that we examined. Mast cell tryptase and HMC-1 lysate had no significant effect on the clonogenic growth of cancer cells co-cultured with fibroblasts. By contrast, mast cell heparin at multiple doses significantly reduced the size and number of colonies of tumor cells co-cultured with fibroblasts, especially in the presence of FGF-7. Neither heparin nor FGF-7, individually or in combination, produced any significant effect on the clonogenic growth of breast cancer cells cultured without fibroblasts. Conclusion Degranulating mast cells are restricted to peri-tumoral fibrous tissue, and mast cell heparin is a powerful inhibitor of clonogenic growth of tumor cells co-cultured with fibroblasts. These results may help to explain the well-known ability of heparin to inhibit the growth of primary and metastatic tumors.

  17. Intersections of lung progenitor cells, lung disease and lung cancer.

    Science.gov (United States)

    Kim, Carla F

    2017-06-30

    The use of stem cell biology approaches to study adult lung progenitor cells and lung cancer has brought a variety of new techniques to the field of lung biology and has elucidated new pathways that may be therapeutic targets in lung cancer. Recent results have begun to identify the ways in which different cell populations interact to regulate progenitor activity, and this has implications for the interventions that are possible in cancer and in a variety of lung diseases. Today's better understanding of the mechanisms that regulate lung progenitor cell self-renewal and differentiation, including understanding how multiple epigenetic factors affect lung injury repair, holds the promise for future better treatments for lung cancer and for optimising the response to therapy in lung cancer. Working between platforms in sophisticated organoid culture techniques, genetically engineered mouse models of injury and cancer, and human cell lines and specimens, lung progenitor cell studies can begin with basic biology, progress to translational research and finally lead to the beginnings of clinical trials. Copyright ©ERS 2017.

  18. Intersections of lung progenitor cells, lung disease and lung cancer

    Directory of Open Access Journals (Sweden)

    Carla F. Kim

    2017-06-01

    Full Text Available The use of stem cell biology approaches to study adult lung progenitor cells and lung cancer has brought a variety of new techniques to the field of lung biology and has elucidated new pathways that may be therapeutic targets in lung cancer. Recent results have begun to identify the ways in which different cell populations interact to regulate progenitor activity, and this has implications for the interventions that are possible in cancer and in a variety of lung diseases. Today's better understanding of the mechanisms that regulate lung progenitor cell self-renewal and differentiation, including understanding how multiple epigenetic factors affect lung injury repair, holds the promise for future better treatments for lung cancer and for optimising the response to therapy in lung cancer. Working between platforms in sophisticated organoid culture techniques, genetically engineered mouse models of injury and cancer, and human cell lines and specimens, lung progenitor cell studies can begin with basic biology, progress to translational research and finally lead to the beginnings of clinical trials.

  19. Myogenic conversion of bladder fibroblasts by construction and ...

    African Journals Online (AJOL)

    The cultured primary bladder fibroblasts were transfected by pEGFP-Myod1 with Lipofection 2000 reagent. The results showed that expression of Myod1 could cause myogenic differentiation of bladder fibroblasts. These findings support the possibility of an alternative approach to exploit the capacity of Myod1 to activate ...