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Sample records for tobacco cells exposed

  1. Lung cell reactions in guinea pigs exposed to tobacco smoke and silica dust or bacterial lipopolysaccharides

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    Sjoestrand, M; Rylander, R

    1984-02-01

    In order to investigate the possibility of the synergistic effects of tobacco smoke and/or silica dust (SiO2) or bacterial endotoxins (LPS), guinea pigs were exposed to combinations of these agents. A 15-day exposure to SiO2 alone caused a decrease in intracellular lysosomal enzymes of alveolar macrophages (AM) and an increase of lysosomal enzymes detected in lung lavage fluid which was present 16 weeks after exposure. The effect was the same in animals which received SiO2 in combination with tobacco smoke. Exposure to LPS caused an increase in the number of neutrophils recovered in lavage fluid. The increase in neutrophils was less in animals previously exposed to tobacco smoke alone or in combination with LPS. Acute exposure to LPS also caused an increase in lactate dehydrogenase, N-acetyl-beta-D-glucosaminidase and acid phosphatase activity detectable in lung lavage fluid. The increase was less pronounced in animals previously exposed to smoke. Cathepsin D was increased in AM after tobacco smoke exposure alone and was decreased to below control values of the animals which received an acute LPS exposure.

  2. Are students exposed to tobacco smoke in German schools?

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    Heinrich, Joachim

    2005-12-01

    Full Text Available The aim of this study was to investigate to which extent 6th grade school children are exposed to tobacco smoke by others. As biomarker for the exposure to tobacco smoke nicotine and cotinine were measured in the urine. Our study population consisted of 771 schoolchildren aged 11-14 years who according to a questionnaire did not smoke. In addition we analysed the data of 459 school children who were not exposed to tobacco smoke at home. The nicotine and cotinine concentrations in the spontaneous urine sample were determined by HPLC methods.On average in about 20% of all non-smoking children, who were not exposed to tobacco smoke at home, biomarker (nicotine or cotinine were detected in the urine. The percentage of the detected biomarker values (nicotine and/or cotinine in the urine of the school children varied between 0% and 50% between schools. In addition we determined the proportion of smoking classmates per school. No positive association was found between the detected biomarker values of the non-smoking school children not exposed to tobacco smoke at home and the proportion of smokers per school. The concentration of biomarker depending on the time of day the urine samples were collected showed higher nicotine and cotinine values when the urine sample was collected between 10 and 12 o'clock in the morning compared to urine samples collected between 7 and 10 a.m.In spite of the limitations our study provides some evidence that children are exposed involuntarily to tobacco smoke by others at school. That is why our results support the requirement of a general legal ban on smoking for teachers, the school staff and students.

  3. Role of Oxidative Stress in the Suppression of Immune Responses in Peripheral Blood Mononuclear Cells Exposed to Combustible Tobacco Product Preparation.

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    Arimilli, Subhashini; Schmidt, Eckhardt; Damratoski, Brad E; Prasad, G L

    2017-10-01

    Cigarette smoking is a major risk factor for several human diseases. Chronic inflammation, resulting from increased oxidative stress, has been suggested as a mechanism that contributes to the increased susceptibility of smokers to cancer and microbial infections. We have previously shown that whole-smoke conditioned medium (WS-CM) and total particulate matter (TPM) prepared from Kentucky 3R4F reference cigarettes [collectively called as combustible tobacco product preparations (TPPs)] potently suppressed agonist-stimulated cytokine secretion and target cell killing in peripheral blood mononuclear cells (PBMCs). Here we have investigated the role of oxidative stress from TPPs, which alters inflammatory responses in vitro. Particularly, we investigated the mechanisms of WS-CM-induced suppression of select cytokine secretions in Toll-like receptor (TLR) agonist-stimulated cells and target cell killing by effector cells in PBMCs. Pretreatment with N-acetyl cysteine (NAC), a precursor of reduced glutathione and an established anti-oxidant, protected against DNA damage and cytotoxicity caused by exposure to WS-CM. Similarly, secretion of tumor necrosis factor (TNF), interleukin (IL)-6, and IL-8 in response to TLR-4 stimulation was restored by pretreatment with NAC. Target cell killing, a functional measure of cytolytic cells in PBMCs, is suppressed by WS-CM. Pretreatment with NAC restored the target cell killing in WS-CM treated PBMCs. This was accompanied by higher perforin levels in the effector cell populations. Collectively, these data suggest that reducing oxidative stress caused by cigarette smoke components restores select immune responses in this ex vivo model.

  4. Antioxidant status of neonates exposed in utero to tobacco smoke.

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    Fayol, L; Gulian, J M; Dalmasso, C; Calaf, R; Simeoni, U; Millet, V

    2005-01-01

    To investigate the influence of maternal smoke exposure on neonatal and maternal antioxidant status, 39 mothers who were active smokers, 14 mothers exposed to environmental tobacco smoke (ETS), 17 controls, and their newborns were included in a prospective, controlled study. Plasma total antioxidant capacity, measured as total radical-trapping antioxidant parameter (TRAP) and ferric reducing antioxidant power (FRAP), and concentrations of specific antioxidants were measured in cord and in maternal blood. A similar, significant increase in ceruloplasmin concentration was observed in neonates born to actively smoking mothers and in those born to ETS exposed mothers. Uric acid and TRAP concentrations were significantly increased in ETS-exposed newborns and their mothers, compared to newborns and mothers from the active smoking and no-exposure groups with a trend towards increased uric acid, TRAP and FRAP concentrations being observed in the active smokers group. Neonatal and maternal antioxidant concentrations correlated significantly, except for ceruloplasmin. Cord blood vitamin A, E and C concentrations were unaffected by smoke exposure. These results show that maternal active smoking as well as ETS exposure significantly affect neonatal and maternal antioxidant status. Copyright (c) 2005 S. Karger AG, Basel

  5. Exposing and addressing tobacco industry conduct in low-income and middle-income countries.

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    Gilmore, Anna B; Fooks, Gary; Drope, Jeffrey; Bialous, Stella Aguinaga; Jackson, Rachel Rose

    2015-03-14

    The tobacco industry's future depends on increasing tobacco use in low-income and middle-income countries (LMICs), which face a growing burden of tobacco-related disease, yet have potential to prevent full-scale escalation of this epidemic. To drive up sales the industry markets its products heavily, deliberately targeting non-smokers and keeps prices low until smoking and local economies are sufficiently established to drive prices and profits up. The industry systematically flaunts existing tobacco control legislation and works aggressively to prevent future policies using its resource advantage to present highly misleading economic arguments, rebrand political activities as corporate social responsibility, and establish and use third parties to make its arguments more palatable. Increasingly it is using domestic litigation and international arbitration to bully LMICs from implementing effective policies and hijacking the problem of tobacco smuggling for policy gain, attempting to put itself in control of an illegal trade in which there is overwhelming historical evidence of its complicity. Progress will not be realised until tobacco industry interference is actively addressed as outlined in Article 5.3 of the Framework Convention on Tobacco Control. Exemplar LMICs show this action can be achieved and indicate that exposing tobacco industry misconduct is an essential first step. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Inhibition of HMGB1 Translocation by Green Tea Extract in Rats Exposed to Environmental Tobacco Smoke

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    Sirintip Chaichalotornkul

    2012-01-01

    Full Text Available Environmental tobacco smoke (ETS exposure is linked to carcinogenic, oxidative and inflammatory cellular reactions. Green tea polyphenol reportedly plays a role in the prevention of inflammation-related diseases. To evaluate the effects of green tea extract (GTE on cellular location of High Mobility Group Box-1 (HMGB1 protein, we studied the lung tissue in rats exposed to cigarette smoke (CS. Rats were divided into three groups; CS, CSG, and C, which were groups of CS-treated only, CS-treated with GTE dietary supplement, and the control, respectively. Our findings by immunocytochemistry showed that abundant HMGB1 translocated from the nucleus to the cytoplasm in the lung tissues of rats that were exposed to CS, whereas HMGB1 was localized to the nuclei of CSG and C group. For in vitro studies, cotinine stimulated the secretion of HMGB1 in a dose and time dependent manner and the HMGB1 level was suppressed by GTE in murine macrophage cell lines. Our results could suggest that GTE supplementation which could suppress HMGB1 may offer a beneficial effect against diseases.

  7. Inhibition of HMGB1 Translocation by Green Tea Extract in Rats Exposed to Environmental Tobacco Smoke

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    Sirintip Chaichalotornkul; Wisuda Suvitayavat; Vanida Sangalangkarn; Yuko Nawa; Kiyoshi Kikuchi; Koichi Kawahara; Tawee Saiwichai; Somphong Narkpinit; Pratap Singhasivanon; Ikuro Maruyama; Salunya Tancharoen

    2012-01-01

    Environmental tobacco smoke (ETS) exposure is linked to carcinogenic, oxidative and inflammatory cellular reactions. Green tea polyphenol reportedly plays a role in the prevention of inflammation-related diseases. To evaluate the effects of green tea extract (GTE) on cellular location of High Mobility Group Box-1 (HMGB1) protein, we studied the lung tissue in rats exposed to cigarette smoke (CS). Rats were divided into three groups; CS, CSG, and C, which were groups of CS-treated only, CS-tre...

  8. Is Tobacco Smoke a Germ-Cell Mutagen?

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    Although no international organization exists to declare whether an agent is a germ-cell mutagen, tobacco smoke may be a human germ-cell mutagen. In the mouse, tobacco smoke induces a significant increase in the mutation frequency at an expanded simple tandem repeat (ESTR) locus....

  9. Biological and environmental risk factors of children exposed or not to environmental tobacco pollution

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    Alice Stenzel de Pina Ferreira

    2017-12-01

    Full Text Available We aimed to investigate the biological and environmental risk facotrs of children exposed or not to environmental tobacco pollution (ETP. A cross-sectional study with 670 children of both sexes, aged between eight and 12 years, from schools located in Anápolis (GO. We used an adapted questionnaire directed to parents/guardians. The parents of children of the non-exposed to ETP group (NETP were more educated. The group of children exposed to ETP (EETP had a higher history of respiratory disease. The EETP resides with a smoker, commonly fathers, who smoke up to 20 cigarretes a day. The EETP lived in houses with fewer windows, less air circulation and more registries of mold. The EETP presents more respiratory diseases and unfavourable socioeconomic conditions. Therefore, there is a need for more care for the exposure and the environment where they live. Health professionals and educators should promote protection, education and stimulate the abandonment of parent smoking.

  10. Cell cycle-dependent O-GlcNAc modification of tobacco histones and their interaction with the tobacco lectin.

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    Delporte, Annelies; De Zaeytijd, Jeroen; De Storme, Nico; Azmi, Abdelkrim; Geelen, Danny; Smagghe, Guy; Guisez, Yves; Van Damme, Els J M

    2014-10-01

    The Nicotiana tabacum agglutinin or Nictaba is a nucleocytoplasmic lectin that is expressed in tobacco after the plants have been exposed to jasmonate treatment or insect herbivory. Nictaba specifically recognizes GlcNAc residues. Recently, it was shown that Nictaba is interacting in vitro with the core histone proteins from calf thymus. Assuming that plant histones - similar to their animal counterparts - undergo O-GlcNAcylation, this interaction presumably occurs through binding of the lectin to the O-GlcNAc modification present on the histones. Hereupon, the question was raised whether this modification also occurs in plants and if it is cell cycle dependent. To this end, histones were purified from tobacco BY-2 suspension cells and the presence of O-GlcNAc modifications was checked. Concomitantly, O-GlcNAcylation of histone proteins was studied. Our data show that similar to animal histones plant histones are modified by O-GlcNAc in a cell cycle-dependent fashion. In addition, the interaction between Nictaba and tobacco histones was confirmed using lectin chromatography and far Western blot analysis. Collectively these findings suggest that Nictaba can act as a modulator of gene transcription through its interaction with core histones. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  11. Exposure of Human Lung Cells to Tobacco Smoke Condensate Inhibits the Nucleotide Excision Repair Pathway.

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    Nathaniel Holcomb

    Full Text Available Exposure to tobacco smoke is the number one risk factor for lung cancer. Although the DNA damaging properties of tobacco smoke have been well documented, relatively few studies have examined its effect on DNA repair pathways. This is especially true for the nucleotide excision repair (NER pathway which recognizes and removes many structurally diverse DNA lesions, including those introduced by chemical carcinogens present in tobacco smoke. The aim of the present study was to investigate the effect of tobacco smoke on NER in human lung cells. We studied the effect of cigarette smoke condensate (CSC, a surrogate for tobacco smoke, on the NER pathway in two different human lung cell lines; IMR-90 lung fibroblasts and BEAS-2B bronchial epithelial cells. To measure NER, we employed a slot-blot assay to quantify the introduction and removal of UV light-induced 6-4 photoproducts and cyclobutane pyrimidine dimers. We find a dose-dependent inhibition of 6-4 photoproduct repair in both cell lines treated with CSC. Additionally, the impact of CSC on the abundance of various NER proteins and their respective RNAs was investigated. The abundance of XPC protein, which is required for functional NER, is significantly reduced by treatment with CSC while the abundance of XPA protein, also required for NER, is unaffected. Both XPC and XPA RNA levels are modestly reduced by CSC treatment. Finally, treatment of cells with MG-132 abrogates the reduction in the abundance of XPC protein produced by treatment with CSC, suggesting that CSC enhances proteasome-dependent turnover of the protein that is mediated by ubiquitination. Together, these findings indicate that tobacco smoke can inhibit the same DNA repair pathway that is also essential for the removal of some of the carcinogenic DNA damage introduced by smoke itself, increasing the DNA damage burden of cells exposed to tobacco smoke.

  12. Tobacco

    Science.gov (United States)

    ... Second-hand smoke is the smoke that fills restaurants, offices or other enclosed spaces when people burn ... as smuggling, illicit manufacturing and counterfeiting. The tobacco industry and others often argue that high tobacco product ...

  13. Reduced number of alpha- and beta-adrenergic receptors in the myocardium of rats exposed to tobacco smoke

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    Larue, D.; Kato, G.

    1981-04-09

    The concentration of alpha- and beta-adrenergic receptors--as measured by specific (/sup 3/H)WB-4101 and (-)-(/sup 3/H)dihydroalprenolol binding--was diminished by 60% below control values in the hearts of rats exposed to tobacco smoke. These changes in receptor numbers took place almost immediately after tobacco smoke exposure and were rapidly reversible after termination of the exposure. The dissociation constant, KD, for (/sup 3/H)WB-4101 was identical in exposed (KD . 0.34 +/- 0.09 nM) and control (KD . 0.35 +/- 0.07 nM) hearts but was significantly different in the case of (-)-(3H)dihydroalprenolol binding (exposed, KD . 2.83 +/- 0.30 mM vs. control KD . 5.22 +/- 0.61 nM). For beta-receptor binding there was no significant difference between exposed and control animals in the Ki values for (-)-epinephrine, (-)-norepinephrine, (-)-alprenolol, (+/-)-propranolol or timolol. (-)-Isoproterenol, however, was found to bind with lower affinity in exposed compared with control hearts. For alpha-receptor binding there was no significant difference between control and 'smoked' animals in the Ki values for (-)-epinephrine, (-0)-norepinephrine or phentolamine. The decrease in alpha- and beta-adrenergic receptor concentration may be related to the phenomenon of receptor desensitization resulting from a release of catecholamines in rats exposed to tobacco smoke.

  14. Acrolein Exposure in Hookah Smokers and Non-Smokers Exposed to Hookah Tobacco Secondhand Smoke: Implications for Regulating Hookah Tobacco Products.

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    Kassem, Nada O F; Kassem, Noura O; Liles, Sandy; Zarth, Adam T; Jackson, Sheila R; Daffa, Reem M; Chatfield, Dale A; Carmella, Steven G; Hecht, Stephen S; Hovell, Melbourne F

    2018-03-06

    Acrolein is a highly ciliatoxic agent, a toxic respiratory irritant, a cardiotoxicant, and a possible carcinogen present in tobacco smoke including hookah tobacco. 105 hookah smokers and 103 non-smokers attended exclusively hookah smoking social events at either a hookah lounge or private home, and provided urine samples the morning of and the morning after the event. Samples were analyzed for 3-hydroxypropylmercapturic acid (3-HPMA), a metabolite of acrolein. Geometric mean (GM) urinary 3-HPMA levels in hookah smokers and non-smokers exposed to secondhand smoke (SHS) increased significantly, 1.41 times, 95% CI = 1.15 to 1.74 and 1.39 times, 95% CI = 1.16 to 1.67, respectively, following a hookah social event. The highest increase (1.68 times, 95% CI = 1.15 to 2.45; p = 0.007) in 3-HPMA post a hookah social event was among daily hookah smokers (GM, from 1991 pmol/mg to 3348 pmol/mg). Pre-to-post event change in urinary 3-HPMA was significantly positively correlated with pre-to-post event change in urinary cotinine among hookah smokers at either location of hookah event, (ρ = 0.359, p = 0.001), and among non-smokers in hookah lounges (ρ = 0.369, p = 0.012). Hookah tobacco smoke is a source of acrolein exposure. Findings support regulating hookah tobacco products including reducing humectants and sugar additives, which are precursors of acrolein under certain pyrolysis conditions. We suggest posting health warning signs for indoor smoking in hookah lounges, and encouraging voluntary bans of smoking hookah tobacco in private homes. Our study is the first to quantify the increase in acrolein exposure in hookah smokers and non-smokers exposed to exclusively hookah tobacco SHS at hookah social events in homes or hookah lounges. Our findings provide additional support for regulating hookah tobacco product content, protecting non-smokers' health by posting health warning signs for indoor smoking in hookah lounges, and encouraging home bans on hookah tobacco smoking to

  15. Toxicological responses in SW mice exposed to inhaled pyrolysates of polymer/tobacco mixtures and blended tobacco.

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    Werley, Michael S; Lee, K Monika; Lemus-Olalde, Ranulfo

    2009-12-01

    Modern cigarette manufacturing is highly automated and produces millions of cigarettes per day. The potential for small inclusions of non-cigarette materials such as wood, cardboard packaging, plastic, and other materials exists as a result of bulk handling and high-speed processing of tobacco. Many non-tobacco inclusions such as wood, paper, and cardboard would be expected to yield similar pyrolysis products as a burning cigarette. The aircraft industry has developed an extensive literature on the pyrolysis products of plastics, however, that have been reported to yield toxic by-products upon burning, by-products that have been lethal in animals and humans upon acute exposure under some exposure conditions. Some of these smoke constituents have also been reported in cigarette smoke. Five synthetic polymers, nylon 6, acrylonitrile-butadiene-styrene (ABS), nylon 12, nylon 6,6, and acrylonitrile-butadiene (AB), and the natural polymer wool were evaluated by adding them to tobacco at a 3, 10, and 30% inclusion level and then pyrolyzing the mixture. The validated smoke generation and exposure system have been described previously. We used the DIN 53-436 tube furnace and nose-only exposure chamber in combination to conduct exposures in Swiss-Webster mice. Potentially useful biological endpoints for predicting hazards in humans included sensory irritation and pulmonary irritation, respiratory function, clinical signs, body weights, bronchoalveolar lavage (BAL) fluid analysis, carboxyhemoglogin, blood cyanide concentrations, and histopathology of the respiratory tract. Chemical analysis of selected smoke constituents in the test atmosphere was also performed in order to compare the toxicological responses with exposure to the test atmospheres. Under the conditions of these studies, biological responses considered relevant and useful for prediction of effects in humans were found for sensory irritation, body weights, BAL fluid analysis, and histopathology of the nose

  16. Principal Component Analysis of Chlorophyll Content in Tobacco, Bean and Petunia Plants Exposed to Different Tropospheric Ozone Concentrations

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    Borowiak, Klaudia; Zbierska, Janina; Budka, Anna; Kayzer, Dariusz

    2014-06-01

    Three plant species were assessed in this study - ozone-sensitive and -resistant tobacco, ozone-sensitive petunia and bean. Plants were exposed to ambient air conditions for several weeks in two sites differing in tropospheric ozone concentrations in the growing season of 2009. Every week chlorophyll contents were analysed. Cumulative ozone effects on the chlorophyll content in relation to other meteorological parameters were evaluated using principal component analysis, while the relation between certain days of measurements of the plants were analysed using multivariate analysis of variance. Results revealed variability between plant species response. However, some similarities were noted. Positive relations of all chlorophyll forms to cumulative ozone concentration (AOT 40) were found for all the plant species that were examined. The chlorophyll b/a ratio revealed an opposite position to ozone concentration only in the ozone-resistant tobacco cultivar. In all the plant species the highest average chlorophyll content was noted after the 7th day of the experiment. Afterwards, the plants usually revealed various responses. Ozone-sensitive tobacco revealed decrease of chlorophyll content, and after few weeks of decline again an increase was observed. Probably, due to the accommodation for the stress factor. While during first three weeks relatively high levels of chlorophyll contents were noted in ozone-resistant tobacco. Petunia revealed a slow decrease of chlorophyll content and the lowest values at the end of the experiment. A comparison between the plant species revealed the highest level of chlorophyll contents in ozone-resistant tobacco.

  17. Tobacco

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    ... 1 in 3 countries, representing 39% of the world's population, monitors tobacco use by repeating nationally representative youth ... 1.4 billion people, or 20% of the world's population, are protected by comprehensive national smoke-free laws. ...

  18. Lethal impacts of cigarette smoke in cultured tobacco cells

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    Kawano Tomonori

    2011-07-01

    Full Text Available Abstract Background In order to understand and generalize the toxic mechanism of cigarette smoke in living cells, comparison of the data between animal systems and other biological system such as microbial and plant systems is highly beneficial. Objective By employing the tobacco cells as model materials for cigarette smoke toxicity assay, the impacts of the combustion by-products such as nitrogen oxides could be highlighted as the toxic impacts of the plant-derived endogenous chemicals could be excluded in the plant cells. Methods Cigarette smoke-induced cell death was assessed in tobacco cell suspension cultures in the presence and absence of pharmacological inhibitors. Results Cigarette smoke was effective in induction of cell death. The smoke-induced cell death could be partially prevented by addition of nitric oxide (NO scavenger, suggesting the role for NO as the cell death mediator. Addition of NO donor to tobacco cells also resulted in development of partial cell death further confirming the role of NO as cell death mediator. Members of reactive oxygen species and calcium ion were shown to be protecting the cells from the toxic action of smoke-derived NO.

  19. Advertising of tobacco products at point of sale: who are more exposed in Brazil?

    Science.gov (United States)

    Ferreira-Gomes, Adriana Bacelar; Moura, Lenildo de; Araújo-Andrade, Silvânia Suely de; Lacerda-Mendes, Felipe; Perez, Cristina A; Abaakouk, Zohra

    2017-01-01

    To describe the adult population perception of cigarette advertising at point of sale, according their tobacco-use status and socio-demographic characteristics such as sex, age, race/color, region, household location and schooling. A multivariable analysis was carried out using data from the Global Adult Tobacco Survey in 2008 and the National Health Survey in 2013. Both surveys showed that among nonsmokers: women, young adults and those who had over 10 years of schooling had more frequently noticed advertising of cigarettes at point of sale. It was also observed that among the population with fewer years of schooling these proportions increased significantly. A measure that completely bans tobacco advertising would be more effective to protect the vulnerable groups from tobacco consumption.

  20. Cluster Headache Clinical Phenotypes: Tobacco Nonexposed (Never Smoker and No Parental Secondary Smoke Exposure as a Child) versus Tobacco-Exposed: Results from the United States Cluster Headache Survey.

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    Rozen, Todd D

    2018-05-01

    To present results from the United States Cluster Headache Survey comparing the clinical presentation of tobacco nonexposed and tobacco-exposed cluster headache patients. Cluster headache is uniquely tied to a personal history of tobacco usage/cigarette smoking and, if the individual cluster headache sufferer did not smoke, it has been shown that their parent(s) typically did and that individual had significant secondary smoke exposure as a child. The true nontobacco exposed (no personal or secondary exposure) cluster headache sufferer has never been fully studied. The United States Cluster Headache Survey consisted of 187 multiple choice questions related to cluster headache including: patient demographics, clinical headache characteristics, family history, triggers, smoking history (personal and secondary), and headache-related disability. The survey was placed on a website from October through December 2008. One thousand one hundred thirty-four individuals completed the survey. One hundred thirty-three subjects or 12% of the surveyed population had no personal smoking/tobacco use history and no secondary smoke exposure as an infant/child, thus a nontobacco exposed population. In the nonexposed population, there were 87 males and 46 females with a gender ratio of 1.9:1. Episodic cluster headache occurred in 80% of nonexposed subjects. One thousand and one survey responders or 88% were tobacco-exposed (729 males and 272 females) with a gender ratio of 2.7:1. Eighty-three percent had a personal smoking history, while only 17% just had parents who smoked with secondary smoke exposure. Eighty-five percent of smokers had double exposure with a personal smoking history and secondary exposure as a child. Nonexposed cluster headache subjects are significantly more likely to develop cluster headache at ages 40 years and younger, while the exposed sufferers are significantly more likely to develop cluster headache at 40 years of age and older. Nonexposed patients have a

  1. Respiratory effects in people exposed to arsenic via the drinking water and tobacco smoking in southern part of Pakistan

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    Arain, Muhammad Balal, E-mail: bilal_ku2004@yahoo.com [Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080 (Pakistan); Kazi, Tasneem Gul, E-mail: tgkazi@yahoo.com [Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080 (Pakistan); Baig, Jameel Ahmed, E-mail: jab_mughal@yahoo.com [Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080 (Pakistan); Jamali, Muhammad Khan, E-mail: mkhanjamali@yahoo.com [Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080 (Pakistan); Afridi, Hassan Imran, E-mail: hassanimranafridi@yahoo.com [Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080 (Pakistan); Jalbani, Nusrat, E-mail: nusratjalbani_21@yahoo.com [Pakistan Council for Scientific and Industrial Research, University Road Karachi-75280 (Pakistan); Sarfraz, Raja Adil, E-mail: rajaadilsarfraz@gmail.com [Department of Chemistry, University of Education, Lahore, Okara Campus, Okara (Pakistan); Shah, Abdul Qadir, E-mail: aqshah07@yahoo.com [Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080 (Pakistan); Kandhro, Ghulam Abbas, E-mail: gakandhro@yahoo.com [Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080 (Pakistan)

    2009-10-15

    In this study, a survey has been conducted during 2005-2007 on surface and groundwater arsenic (As) contamination and its impact on the health of local population, of villages located on the banks of Manchar lake, southern part of Sindh, Pakistan. We have also assessed the relationship between arsenic exposure through respiratory disorders in male subjects with drinking water and smoking cigarettes made from tobacco grown in agricultural land irrigated with As contaminated lake water. The biological samples (blood and scalp hair) were collected from As exposed subjects (100% smokers) and age matched healthy male subjects (40.2% smoker and 59.8% non smokers) belong to unexposed areas for comparison purposes. The As concentration in drinking water (surface and underground water), agricultural soil, cigarette tobacco and biological samples were determined by electrothermal atomic absorption spectrometry. The range of As concentrations in lake water was 35.2-158 {mu}g/L (average 97.5 {mu}g/L), which is 3-15 folds higher than permissible limit of World Health Organization (WHO, 2004). While the As level in local cigarette tobacco was found to be 3-6 folds higher than branded cigarettes (0.37-0.79 {mu}g/g). Arsenic exposed subjects (with and without RD) had significantly elevated levels of As in their biological samples as compared to referent male subject of unexposed area. These respiratory effects were more pronounced in individuals who had also As induced skin lesions. The linear regressions showed good correlations between As concentrations in water versus hair and blood samples of exposed subjects with and without respiratory problems.

  2. Respiratory effects in people exposed to arsenic via the drinking water and tobacco smoking in southern part of Pakistan

    International Nuclear Information System (INIS)

    Arain, Muhammad Balal; Kazi, Tasneem Gul; Baig, Jameel Ahmed; Jamali, Muhammad Khan; Afridi, Hassan Imran; Jalbani, Nusrat; Sarfraz, Raja Adil; Shah, Abdul Qadir; Kandhro, Ghulam Abbas

    2009-01-01

    In this study, a survey has been conducted during 2005-2007 on surface and groundwater arsenic (As) contamination and its impact on the health of local population, of villages located on the banks of Manchar lake, southern part of Sindh, Pakistan. We have also assessed the relationship between arsenic exposure through respiratory disorders in male subjects with drinking water and smoking cigarettes made from tobacco grown in agricultural land irrigated with As contaminated lake water. The biological samples (blood and scalp hair) were collected from As exposed subjects (100% smokers) and age matched healthy male subjects (40.2% smoker and 59.8% non smokers) belong to unexposed areas for comparison purposes. The As concentration in drinking water (surface and underground water), agricultural soil, cigarette tobacco and biological samples were determined by electrothermal atomic absorption spectrometry. The range of As concentrations in lake water was 35.2-158 μg/L (average 97.5 μg/L), which is 3-15 folds higher than permissible limit of World Health Organization (WHO, 2004). While the As level in local cigarette tobacco was found to be 3-6 folds higher than branded cigarettes (0.37-0.79 μg/g). Arsenic exposed subjects (with and without RD) had significantly elevated levels of As in their biological samples as compared to referent male subject of unexposed area. These respiratory effects were more pronounced in individuals who had also As induced skin lesions. The linear regressions showed good correlations between As concentrations in water versus hair and blood samples of exposed subjects with and without respiratory problems.

  3. Evaluation of genetic damage in tobacco and arsenic exposed population of Southern Assam, India using buccal cytome assay and comet assay.

    Science.gov (United States)

    Roy, Prasenjit; Mukherjee, Anita; Giri, Sarbani

    2016-02-01

    Ground water is the principal source of drinking water in Assam. Ground water contamination of arsenic in drinking water is a great concern for human health and considered as a human carcinogen. The present cytogenetic biomonitoring study was undertaken to investigate the genotoxic effects associated with people of southern Assam consuming arsenic contaminated water and chewing tobacco. Employing the buccal cytome assay, exfoliated cells were analyzed in 138 individuals of age range 22-42 years and divided into four groups. Group I (n=54) are participants residing in localities where ground water contains arsenic concentration below the permissible limit (comet assay, percent of tail DNA gradually increases among the groups and has statistical significance. Spearman correlation revealed strong positive correlation between the arsenic exposed peoples and the binucleated cells (r=0.4763; Pcomet assay. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Tobacco and e-cigarette products initiate Kupffer cell inflammatory responses.

    Science.gov (United States)

    Rubenstein, David A; Hom, Sarah; Ghebrehiwet, Berhane; Yin, Wei

    2015-10-01

    Kupffer cells are liver resident macrophages that are responsible for screening and clearing blood of pathogens and foreign particles. It has recently been shown that Kupffer cells interact with platelets, through an adhesion based mechanism, to aid in pathogen clearance and then these platelets re-enter the general systemic circulation. Thus, a mechanism has been identified that relates liver inflammation to possible changes in the systemic circulation. However, the role that Kupffer cells play in cardiovascular disease initiation/progression has not been elucidated. Thus, our objective was to determine whether or not Kupffer cells are responsive to a classical cardiovascular risk factor and if these changes can be transmitted into the general systemic circulation. If Kupffer cells initiate inflammatory responses after exposure to classical cardiovascular risk factors, then this provides a potential alternative/synergistic pathway for cardiovascular disease initiation. We aimed to elucidate the prevalence of this potential pathway. We hypothesized that Kupffer cells would initiate a robust inflammatory response after exposure to tobacco cigarette or e-cigarette products and that the inflammatory response would have the potential to antagonize other salient cells for cardiovascular disease progression. To test this, Kupffer cells were incubated with tobacco smoke extracts, e-cigarette vapor extracts or pure nicotine. Complement deposition onto Kupffer cells, Kupffer cell complement receptor expression, oxidative stress production, cytokine release and viability and density were assessed after the exposure. We observed a robust inflammatory response, oxidative stress production and cytokine release after Kupffer cells were exposed to tobacco or e-cigarette extracts. We also observed a marginal decrease in cell viability coupled with a significant decrease in cell density. In general, this was not a function of the extract formulation (e.g. tobacco vs. e

  5. Different genome maintenance strategies in human and tobacco cells.

    Science.gov (United States)

    Pelczar, Pawel; Kalck, Véronique; Kovalchuk, Igor

    2003-08-22

    In this work, genome maintenance strategies of organisms belonging to different kingdoms (animals versus plants) but of similar genome size were investigated using a novel, universal double-strand break (DSB) repair assay. Different plasmids linearised with KpnI, Acc65I or EcoRV yielding either 3' or 5' protruding or blunt DNA termini, respectively, were transfected into HeLa cells and Nicotiana plumbaginifolia protoplasts and assayed for the efficiency and fidelity of DSB repair. We show that the mechanism of break sealing is similar but that drastic differences are seen in the fidelity of repair: in HeLa cells, 50-55% DSBs were repaired precisely, compared to as little as 15-30% in tobacco cells. Moreover, the DSB repair in plants resulted in 30-40% longer deletions and significantly shorter insertions. Combined, these led to more than twofold larger net DNA loss in tobacco cells. Our observations point to possible differences in the strategies of DSB repair and genome maintenance in plants and animals.

  6. Expression of Plant Receptor Kinases in Tobacco BY-2 Cells.

    Science.gov (United States)

    Shinohara, Hidefumi; Matsubayashi, Yoshikatsu

    2017-01-01

    Although more than 600 single-transmembrane receptor kinase genes have been found in the Arabidopsis genome, only a few of them have known physiological functions, and even fewer plant receptor kinases have known specific ligands. Ligand-binding analysis must be operated using the functionally expressed receptor form. However, the relative abundance of native receptor kinase molecules in the plasma membrane is often quite low. Here, we present a method for stable and functional expression of plant receptor kinases in tobacco BY-2 cells that allows preparation of microsomal fractions containing the receptor. This procedure provides a sufficient amount of receptor proteins while maintaining its ligand-binding activities.

  7. A novel cell division factor from tobacco 2B-13 cells that induced cell division in auxin-starved tobacco BY-2 cells

    Science.gov (United States)

    Shimizu, Takashi; Eguchi, Kentaro; Nishida, Ikuo; Laukens, Kris; Witters, Erwin; van Onckelen, Harry; Nagata, Toshiyuki

    2006-06-01

    Effects of auxin as plant hormones are widespread; in fact in almost all aspects of plant growth and development auxin plays a pivotal role. Although auxin is required for propagating cell division in plant cells, its effect upon cell division is least understood. If auxin is depleted from the culture medium, cultured cells cease to divide. It has been demonstrated in this context that the addition of auxin to auxin-starved nondividing tobacco BY-2 cells induced semisynchronous cell division. On the other hand, there are some cell lines, named habituated cells, that can grow without auxin. The cause and reason for the habituated cells have not been clarified. A habituated cell line named 2B-13 is derived from the tobacco BY-2 cell line, which has been most intensively studied among plant cell lines. When we tried to find the difference between two cell lines of BY-2 and 2B-13 cells, we found that the addition of culture filtrated from the auxin-habituated 2B-13 cells induced semisynchronous cell division in auxin-starved BY-2 cells. The cell division factor (CDF) that is responsible for inducing cell division in auxin-starved BY-2 cells was purified to near-homogeneity by sequential passage through a hydroxyapatite column, a ConA Sepharose column and a Sephadex gel filtration column. The resulting purified fraction appeared as a single band of high molecular weight on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels by silver staining and was able to induce cell division in auxin-starved BY-2 cells. Identification of the protein by MALD-TOF-MS/MS revealed that it is structurally related to P-glycoprotein from Gossypioides kirkii, which belongs to ATP-binding cassette (ABC)-transporters. The significance of CDF as a possible ABC-transporter is discussed in relationship to auxin-autotrophic growth and auxin-signaling pathway.

  8. Isolation of Autolysosomes from Tobacco BY-2 Cells.

    Science.gov (United States)

    Takatsuka, Chihiro; Inoue-Aono, Yuko; Moriyasu, Yuji

    2017-01-01

    Autolysosomes are organelles that sequester and degrade a portion of the cytoplasm during autophagy. Although autophagosomes are short lived compared to other organelles such as mitochondria, plastids, and peroxisomes, many autolysosomes accumulate in tobacco BY-2 cells cultured under sucrose starvation conditions in the presence of a cysteine protease inhibitor. We here describe our methodology for isolating autolysosomes from BY-2 cells by conventional cell fractionation using a Percoll gradient. The autolysosome fraction separates clearly from fractions containing mitochondria and peroxisomes. It contains acid phosphatase, vacuolar H + -ATPase, and protease activity. Electron micrographs show that the fraction contains partially degraded cytoplasm seen in autolysosomes before isolation although an autolysosome structure is only partially preserved.

  9. Oxidative stress is reduced in Wistar rats exposed to smoke from tobacco and treated with specific broad-band pulse electromagnetic fields

    Directory of Open Access Journals (Sweden)

    Bajić V.

    2009-01-01

    Full Text Available There have been a number of attempts to reduce the oxidative radical burden of tobacco. A recently patented technology, pulse electromagnetic technology, has been shown to induce differential action of treated tobacco products versus untreated products on the production of reactive oxygen species (ROS in vivo. In a 90-day respiratory toxicity study, Wistar rats were exposed to cigarette smoke from processed and unprocessed tobacco and biomarkers of oxidative stress were compared with pathohistological analysis of rat lungs. Superoxide dismutase (SOD activity was decreased in a dose-dependent manner to 81% in rats exposed to smoke from normal cigarettes compared to rats exposed to treated smoke or the control group. These results correspond to pathohistological analysis of rat lungs, in which those rats exposed to untreated smoke developed initial signs of emphysema, while rats exposed to treated smoke showed no pathology, as in the control group. The promise of inducing an improved health status in humans exposed to smoke from treated cigarettes merits further investigation.

  10. Interaction of the Tobacco mosaic virus movement protein with microtubules during the cell cycle in tobacco BY-2 cells.

    Science.gov (United States)

    Boutant, Emmanuel; Fitterer, Chantal; Ritzenthaler, Christophe; Heinlein, Manfred

    2009-10-01

    Cell-to-cell movement of Tobacco mosaic virus (TMV) involves the interaction of virus-encoded 30-kDa movement protein (MP) with microtubules. In cells behind the infection front that accumulate high levels of MP, this activity is reflected by the formation of stabilized MP/microtubule complexes. The ability of MP to bind along and stabilize microtubules is conserved upon expression in mammalian cells. In mammalian cells, the protein also leads to inhibition of mitosis and cell division through a microtubule-independent process correlated with the loss of centrosomal gamma-tubulin and of centrosomal microtubule-nucleation activity. Since MP has the capacity to interact with plant factors involved in microtubule nucleation and dynamics, we used inducible expression in BY-2 cells to test whether MP expression inhibits mitosis and cell division also in plants. We demonstrate that MP:GFP associates with all plant microtubule arrays and, unlike in mammalian cells, does not interfere with mitosis. Thus, MP function and the interaction of MP with factors of the cytoskeleton do not entail an inhibition of mitosis in plants. We also report that the protein targets primary plasmodesmata in BY-2 cells immediately upon or during cytokinesis and that the accumulation of MP in plasmodesmata occurs in the presence of inhibitors of the cytoskeleton and the secretory pathway.

  11. Biological behaviour of buccal cells exposed to blue light

    International Nuclear Information System (INIS)

    Gritsch, Kerstin; Ponsonnet, Laurence; Schembri, Catherine; Farge, Pierre; Pourreyron, Laurence; Grosgogeat, Brigitte

    2008-01-01

    Blue light is used in dental practise to cure resin-based materials, but the path of the light often includes oral tissues such as gingival tissues. While adverse effects of blue light exposure on cells - such as retina cells - are well known, few studies have investigated the impact of blue light exposure on oral cells. The aim of the present in vitro study was to assess the biological effects of blue light emitted by two dental curing devices (a plasma-arc and a light-emitting diode curing unit) on human gingival fibroblasts. Light intensities and light-induced temperature rise were respectively measured with a radiometer and a thermocouple. Cellular response to blue light exposure was assessed by the observation of cell morphology (scanning electron microscopy) and the estimation of cell mitochondrial activity (MTT assay). Light intensities measured at the clinical distance were 488 ± 42 mW/cm 2 for the plasma-arc unit and ranged from 61 ± 5 to 140 ± 16 mW/cm 2 for the light-emitting diodes unit, according to the curing program used. The highest temperature rise was 0.5 and 3.5 deg. C for exposure to the plasma-arc light and to the light-emitting diodes light, respectively. Results showed no differences between exposed- and non-exposed cells in regards to cell morphology. However, cells exposed to blue light presented an increased mitochondrial activity compared to control cells (non-exposed), and mostly those exposed to plasma-arc light

  12. Analyzing Cell Wall Elasticity After Hormone Treatment: An Example Using Tobacco BY-2 Cells and Auxin.

    Science.gov (United States)

    Braybrook, Siobhan A

    2017-01-01

    Atomic force microscopy, and related nano-indentation techniques, is a valuable tool for analyzing the elastic properties of plant cell walls as they relate to changes in cell wall chemistry, changes in development, and response to hormones. Within this chapter I will describe a method for analyzing the effect of the phytohormone auxin on the cell wall elasticity of tobacco BY-2 cells. This general method may be easily altered for different experimental systems and hormones of interest.

  13. Aphidicolin-induced nuclear elongation in tobacco BY-2 cells.

    Science.gov (United States)

    Yasuhara, Hiroki; Kitamoto, Kazuki

    2014-05-01

    Plant nuclei are known to differentiate into various shapes within a single plant. However, little is known about the mechanisms of nuclear morphogenesis. We found that nuclei of tobacco BY-2 cells were highly elongated on long-term treatment with 5 mg l⁻¹ aphidicolin, an inhibitor of DNA polymerase α. In aphidicolin-treated cells, the nuclear length was correlated with the cell length. During culture in the presence of aphidicolin, the nuclei were elongated in parallel with cell elongation. Nuclear elongation was inhibited by the inhibition of cell elongation with 2,6-dichlorobenzonitrile, a cellulose synthesis inhibitor. However, cell elongation induced in the auxin-depleted medium in the absence of aphidicolin did not cause nuclear elongation, indicating that cell elongation alone is not sufficient for nuclear elongation. Treatment with either latrunculin B or propyzamide inhibited the aphidicolin-induced nuclear elongation, indicating that both actin filaments and microtubules (MTs) are required for nuclear elongation. Observations using BY-YTHCLR2 cells, in which actin filaments, MTs and nuclei were simultaneously visualized, revealed that the longitudinally arranged MT bundles associated with the nucleus play an important role in nuclear elongation, and that actin filaments affect the formation of these MT bundles. In aphidicolin-treated cells, the nuclear DNA contents of the elongated nuclei exceeded 4C, and the nuclear length was highly correlated with the nuclear DNA content. In cells treated with 50 mg l⁻¹ aphidicolin, cells were elongated and nucleus-associated longitudinal MT bundles were formed, but the nuclear DNA contents did not exceed 4C and the nuclei did not elongate. These results indicate that an increase in the nuclear DNA content above 4C is also required for nuclear elongation.

  14. Elemental analysis in cultured cells, tobacco and grape, treated with aluminum

    International Nuclear Information System (INIS)

    Tanoi, K.; Iikura, H; Nakanishi, T.M.

    2001-01-01

    Relationship between Al toxicity and the Al, Fe and B amount of element in tobacco and grape cells are discussed. Al and Fe were analyzed by neutron activation analysis and B was analyzed by prompt gamma-ray analysis. Callose content was also measured as an indicator of cell damage induced Al toxicity. When tobacco cells were incubated in 1 mM and 300 μM Al solution, the pattern of callose formation was much similar to that of Fe accumulation than that of Al accumulation in tobacco cells, suggesting that the increase of Fe content induced toxic effect along with Al incorporated into the cells. However, this tendency was not observed in grape cells. Boron content did not show any relation to those of Al or Fe throughout the Al treatment in both tobacco and grape cells. (author)

  15. Action spectra in mammalian cells exposed to ultraviolet radiation

    International Nuclear Information System (INIS)

    Anon.

    1981-01-01

    A review is given of the literature published since 1977 on action spectra in mammalian cells exposed to ultraviolet radiation in the wavelength region above 220 nm. Action spectra for lethal events are discussed for cell inactivation in normal cells, growth arrested cells and photosensitive cells. Action spectra for non-lethal events are also discussed in relation to pyrimidine dimer formation, photoreactivation and the use of photosensitisers. It was concluded from these studies that damage to the DNA, and the extent of the repair of this damage, seems to determine a cell's response to such parameters as inactivation, mutation, transformation, latent viral activation, cellular viral capacity and ultraviolet enhanced viral reactivation. In addition to the direct effects of UV on DNA, photosensitization of cellular responses with chemicals such as 8-MOP extend the wavelength range at which damage can be demonstrated. (U.K.)

  16. The cell biology of Tobacco mosaic virus replication and movement

    Directory of Open Access Journals (Sweden)

    Chengke eLiu

    2013-02-01

    Full Text Available Successful systemic infection of a plant by Tobacco mosaic virus (TMV requires three processes that repeat over time: initial establishment and accumulation in invaded cells, intercellular movement and systemic transport. Accumulation and intercellular movement of TMV necessarily involves intracellular transport by complexes containing virus and host proteins and virus RNA during a dynamic process that can be visualized. Multiple membranes appear to assist TMV accumulation, while membranes, microfilaments and microtubules appear to assist TMV movement. Here we review cell biological studies that describe TMV-membrane, -cytoskeleton and -other host protein interactions which influence virus accumulation and movement in leaves and callus tissue. The importance of understanding the developmental phase of the infection in relationship to the observed virus-membrane or -host protein interaction is emphasized. Utilizing the latest observations of TMV-membrane and -host protein interactions within our evolving understanding of the infection ontogeny, a model for TMV accumulation and intracellular spread in a cell biological context is provided.

  17. Genetic transformation of tobacco NT1 cells with Agrobacterium tumefaciens.

    Science.gov (United States)

    Mayo, Kristin J; Gonzales, Barbara J; Mason, Hugh S

    2006-01-01

    This protocol is used to produce stably transformed tobacco (Nicotiana tabacum) NT1 cell lines, using Agrobacterium tumefaciens-mediated DNA delivery of a binary vector containing a gene encoding hepatitis B surface antigen and a gene encoding the kanamycin selection marker. The NT1 cultures, at the appropriate stage of growth, are inoculated with A. tumefaciens containing the binary vector. A 3-day cocultivation period follows, after which the cultures are rinsed and placed on solid selective medium. Transformed colonies ('calli') appear in approximately 4 weeks; they are subcultured until adequate material is obtained for analysis of antigen production. 'Elite' lines are selected based on antigen expression and growth characteristics. The time required for the procedure from preparation of the plant cell materials to callus development is approximately 5 weeks. Growth of selected calli to sufficient quantities for antigen screening may require 4-6 weeks beyond the initial selection. Creation of the plasmid constructs, transformation of the A. tumefaciens line, and ELISA and Bradford assays to assess protein production require additional time.

  18. Plant Cell Division Analyzed by Transient Agrobacterium-Mediated Transformation of Tobacco BY-2 Cells.

    Science.gov (United States)

    Buschmann, Henrik

    2016-01-01

    The continuing analysis of plant cell division will require additional protein localization studies. This is greatly aided by GFP-technology, but plant transformation and the maintenance of transgenic lines can present a significant technical bottleneck. In this chapter I describe a method for the Agrobacterium-mediated genetic transformation of tobacco BY-2 cells. The method allows for the microscopic analysis of fluorescence-tagged proteins in dividing cells in within 2 days after starting a coculture. This transient transformation procedure requires only standard laboratory equipment. It is hoped that this rapid method would aid researchers conducting live-cell localization studies in plant mitosis and cytokinesis.

  19. Dedifferentiation of tobacco cells is associated with ribosomal RNA gene hypomethylation, increased transcription, and chromatin alterations

    Czech Academy of Sciences Publication Activity Database

    Koukalová, Blažena; Fojtová, Miloslava; Lim, Yoong Kar; Fulneček, Jaroslav; Leitch, Rowland Andrew; Kovařík, Aleš

    2005-01-01

    Roč. 139, - (2005), s. 275-286 ISSN 0032-0889 Institutional research plan: CEZ:AV0Z50040507 Keywords : pluripotent tobacco cells * epigenetic changes Subject RIV: BO - Biophysics Impact factor: 6.114, year: 2005

  20. Introduction of transformed chloroplasts from tobacco into petunia by asymmetric cell fusion.

    Science.gov (United States)

    Sigeno, Asako; Hayashi, Sugane; Terachi, Toru; Yamagishi, Hiroshi

    2009-11-01

    Plastid engineering technique has been established only in Nicotiana tabacum, and the widespread application is severely limited so far. In order to exploit a method to transfer the genetically transformed plastomes already obtained in tobacco into other plant species, somatic cell fusion was conducted between a plastome transformant of tobacco and a cultivar of petunia (Petunia hybrida). A tobacco strain whose plastids had been transformed with aadA (a streptomycin/spectinomycin adenylyltransferase gene) and mdar [a gene for monodehydroascorbate reductase (MDAR)] and a petunia variety, 'Telstar', were used as cell fusion partners. An efficient regeneration system from the protoplasts of both the parents, and effectiveness of selection for the aadA gene with spectinomycin were established before the cell fusion. In addition, the influence of UV irradiation on the callus development from the protoplasts and shoot regeneration of tobacco was investigated. Protoplasts were cultured after cell fusion treatment with polyethylene glycol, and asymmetric somatic cybrids were selected using the aadA gene as a marker. Although many shoots of tobacco that had escaped the UV irradiation regenerated, several shoots possessing the morphology of petunia and the resistance to spectinomycin were obtained. Molecular analyses of the petunia type regenerants demonstrated that they had the nuclear and mitochondrial genomes derived from petunia besides the chloroplasts of tobacco transformed with aadA and mdar. Furthermore, it was ascertained that mdar was transcribed in the somatic cybrids. The results indicate the success in intergeneric transfer of transformed plastids of tobacco into petunia.

  1. The efficiency of photovoltaic cells exposed to pulsed laser light

    Science.gov (United States)

    Lowe, R. A.; Landis, G. A.; Jenkins, P.

    1993-01-01

    Future space missions may use laser power beaming systems with a free electron laser (FEL) to transmit light to a photovoltaic array receiver. To investigate the efficiency of solar cells with pulsed laser light, several types of GaAs, Si, CuInSe2, and GaSb cells were tested with the simulated pulse format of the induction and radio frequency (RF) FEL. The induction pulse format was simulated with an 800-watt average power copper vapor laser and the RF format with a frequency-doubled mode-locked Nd:YAG laser. Averaged current vs bias voltage measurements for each cell were taken at various optical power levels and the efficiency measured at the maximum power point. Experimental results show that the conversion efficiency for the cells tested is highly dependent on cell minority carrier lifetime, the width and frequency of the pulses, load impedance, and the average incident power. Three main effects were found to decrease the efficiency of solar cells exposed to simulated FEL illumination: cell series resistance, LC 'ringing', and output inductance. Improvements in efficiency were achieved by modifying the frequency response of the cell to match the spectral energy content of the laser pulse with external passive components.

  2. Sulfated lentinan induced mitochondrial dysfunction leads to programmed cell death of tobacco BY-2 cells.

    Science.gov (United States)

    Wang, Jie; Wang, Yaofeng; Shen, Lili; Qian, Yumei; Yang, Jinguang; Wang, Fenglong

    2017-04-01

    Sulphated lentinan (sLTN) is known to act as a resistance inducer by causing programmed cell death (PCD) in tobacco suspension cells. However, the underlying mechanism of this effect is largely unknown. Using tobacco BY-2 cell model, morphological and biochemical studies revealed that mitochondrial reactive oxygen species (ROS) production and mitochondrial dysfunction contribute to sLNT induced PCD. Cell viability, and HO/PI fluorescence imaging and TUNEL assays confirmed a typical cell death process caused by sLNT. Acetylsalicylic acid (an ROS scavenger), diphenylene iodonium (an inhibitor of NADPH oxidases) and protonophore carbonyl cyanide p-trifluoromethoxyphenyl hydrazone (a protonophore and an uncoupler of mitochondrial oxidative phosphorylation) inhibited sLNT-induced H 2 O 2 generation and cell death, suggesting that ROS generation linked, at least partly, to a mitochondrial dysfunction and caspase-like activation. This conclusion was further confirmed by double-stained cells with the mitochondria-specific marker MitoTracker RedCMXRos and the ROS probe H 2 DCFDA. Moreover, the sLNT-induced PCD of BY-2 cells required cellular metabolism as up-regulation of the AOX family gene transcripts and induction of the SA biosynthesis, the TCA cycle, and miETC related genes were observed. It is concluded that mitochondria play an essential role in the signaling pathway of sLNT-induced ROS generation, which possibly provided new insight into the sLNT-mediated antiviral response, including PCD. Copyright © 2016. Published by Elsevier Inc.

  3. Biolistic transformation of tobacco and maize suspension cells using bacterial cells as microprojectiles.

    Science.gov (United States)

    Rasmussen, J L; Kikkert, J R; Roy, M K; Sanford, J C

    1994-01-01

    We have used both Escherichia coli cells and Agrobacterium tumefaciens cells as microprojectiles to deliver DNA into suspension-cultured tobacco (Nicotiana tabacum L. line NT1) cells using a helium powered biolistic device. In addition, E. coli cells were used as microprojectiles for the transformation of suspension-cultured maize (Zea mays cv. Black Mexican Sweet) cells. Pretreating the bacterial cells with phenol at a concentration of 1.0%, and combining the bacterial cells with tungsten particles increased the rates of transformation. In N. tabacum, we obtained hundreds of transient transformants per bombardment, but were unable to recover any stable transformants. In Z. mays we obtained thousands of transient transformants and an average of six stable transformants per bombardment. This difference is discussed.

  4. Nitric oxide modulates cadmium influx during cadmium-induced programmed cell death in tobacco BY-2 cells.

    Science.gov (United States)

    Ma, Wenwen; Xu, Wenzhong; Xu, Hua; Chen, Yanshan; He, Zhenyan; Ma, Mi

    2010-07-01

    Nitric oxide (NO) is a bioactive gas and functions as a signaling molecule in plants exposed to diverse biotic and abiotic stresses including cadmium (Cd(2+)). Cd(2+) is a non-essential and toxic heavy metal, which has been reported to induce programmed cell death (PCD) in plants. Here, we investigated the role of NO in Cd(2+)-induced PCD in tobacco BY-2 cells (Nicotiana tabacum L. cv. Bright Yellow 2). In this work, BY-2 cells exposed to 150 microM CdCl(2) underwent PCD with TUNEL-positive nuclei, significant chromatin condensation and the increasing expression of a PCD-related gene Hsr203J. Accompanied with the occurring of PCD, the production of NO increased significantly. The supplement of NO by sodium nitroprusside (SNP) had accelerated the PCD, whereas the NO synthase inhibitor Nomega-nitro-L-arginine methyl ester hydrochloride (L-NAME) and NO-specific scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) alleviated this toxicity. To investigate the mechanism by which NO exerted its function, Cd(2+) concentration was measured subsequently. SNP led more Cd(2+) content than Cd(2+) treatment alone. By contrast, the prevention of NO by L-NAME decreased Cd(2+) accumulation. Using the scanning ion-selective electrode technique, we analyzed the pattern and rate of Cd(2+) fluxes. This analysis revealed the promotion of Cd(2+) influxes into cells by application of SNP, while L-NAME and cPTIO reduced the rate of Cd(2+) uptake or even resulted in net Cd(2+) efflux. Based on these founding, we concluded that NO played a positive role in CdCl(2)-induced PCD by modulating Cd(2+) uptake and thus promoting Cd(2+) accumulation in BY-2 cells.

  5. Synthesis of protein in intestinal cells exposed to cholera toxin

    International Nuclear Information System (INIS)

    Peterson, J.W.; Berg, W.D. Jr.; Coppenhaver, D.H.

    1987-01-01

    The mechanism by which cyclic adenosine monophosphate (AMP), formed by intestinal epithelial cells in response to cholera toxin, ultimately results in alterations in water and electrolyte transport is poorly understood. Several studies have indicated that inhibitors of transcription or translation block much of the transport of ions and water in the intestine and edema formation in tissue elicited by cholera toxin. Data presented in this study confirmed the inhibitory effects of cycloheximide on cholera toxin-induced fluid accumulation in the rabbit intestinal loop model. Neither cycloheximide nor actinomycin D altered the amount of cyclic AMP that accumulated in intestinal cells and Chinese hamster ovary cells exposed to cholera toxin. An increase in [ 3 H] leucine incorporation was readily demonstrable in intestinal epithelial cells from rabbits challenged with Vibrio cholerae. Similarly, intestinal epithelial cells incubated with cholera toxin for 4 hr synthesized substantially more protein than controls as determined by relative incorporation of [ 35 S] methionine. Most of the new protein synthesized in response to cholera toxin was membrane associated and of high molecular weight. The possible significance of the toxin-induced protein relative to cholera pathogenesis was discussed

  6. A fundamental research of growth, metabolism and product formation of tobacco suspension cells at different scales

    OpenAIRE

    Ullisch, David

    2012-01-01

    For over two decades, plant cell cultures have been promising hosts for the expression of recombinant proteins such as hormones, growth factors, full-size antibodies and antigens. So far, over 700 different plant cell cultures are stored in the German Collection of Microorganisms and Cell Cultures (DSMZ) in Braunschweig. Among these plant cell cultures, the tobacco cell line Nicotiana tabacum Bright Yellow 2 (BY-2) was chosen as a good host cell line for the production of recombinant proteins...

  7. How does tobacco smoke influence the morphometry of the fetus and the umbilical cord?-Research on pregnant women with intrauterine growth restriction exposed to tobacco smoke.

    Science.gov (United States)

    Milnerowicz-Nabzdyk, Ewa; Bizoń, Anna

    2015-12-01

    Proper structure of the umbilical cord is important for the fetal development. We evaluated effects of toxic factors from tobacco smoke on fetal and umbilical cord morphometry. 109 women in weeks 29-40 of pregnancy (31 smokers with intrauterine growth restriction (IUGR); 28 non-smoking women with IUGR; 50 healthy pregnancies) were included. In smokers with IUGR, cotinine, cadmium and lead concentrations were significantly higher than in controls (mean 55.23ng/l; 1.52ng/ml; 14.85ng/ml vs 1.07; 0.34; 9.42) and inverse correlation between lead concentration and uncoiled umbilical cord was significant (r=-0.80). In smokers with IUGR, area of Wharton's jelly was increased compared to nonsmokers and controls. Inverse correlations occurred between cotinine and cadmium concentration and fetal percentile in smokers (r=-0.87; r=-0.87) and non-smokers (r=-0.47; r=-0.78) with IUGR. Exposure to tobacco smoke measured by cotinine, cadmium and lead concentration has an impact on fetal growth and umbilical cord morphometry and correlates with intensity of IUGR. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. Polyamines and polyamine biosynthesis in cells exposed to hyperthermia

    Energy Technology Data Exchange (ETDEWEB)

    Gerner, E.W.; Stickney, D.G.; Herman, T.S.; Fuller, D.J.

    1983-02-01

    The issue of how polyamines act to sensitize cultured cells to the lethal effects of hyperthermia was investigated using Chinese hamster cells which were induced to express thermotolerance. Intracellular levels of these naturally occurring polycations were manipulated in certain situations by treating whole cells with methylglyoxal bis-(guanylhydrazone), an inhibitor of the S-adenosyl-L-methionine decarboxylases. Exogenous spermine as low as 100 ..mu..M in the culture media dramatically sensitized cells expressing thermotolerance to the lethal effects of subsequent 42/sup 0/C exposures. When thermotolerance was differentially induced in cultures exposed to 42.4/sup 0/C by varying the rate of heating from 37 to 42.4/sup 0/C, the most resistant cells and the highest levels of intracellular spermidine and spermine. This finding was explainable in part by the observation that the putrescine-dependent S-adenosyl-L-methionine decarboxylase activity was minimally affected in cells expressng the greatest degree of thermotolerance. When this enzyme activity was inhibited by drug, lowered intracellular polyamine levels did not correspond with subsequent survival responses to heat. Interestingly, cultures treated with methylglyoxal bis-(guanylhydrazone) 24 hr previous to heat exposure showed a reduced capacity to express rate of heating-induced thermotolerance. Together, these results demonstrate that the polyamines, especially spermidine and spermine, enhance hyperthermia-induced cell killing by some mechanism involving the plasma membrane. Further, our data suggest that methylglyoxal bis-(guanylhydrazone) can act to affect thermal responses by a mechanism(s) other than modification of intracellular polyamine levels.

  9. Why does anatabine, but not nicotine, accumulate in jasmonate-elicited cultured tobacco BY-2 cells?

    Science.gov (United States)

    Shoji, Tsubasa; Hashimoto, Takashi

    2008-08-01

    Suspension-cultured cells of Nicotiana tabacum cv. Bright Yellow-2 (BY-2) grow rapidly in a highly homogenous population and still exhibit the general behavior of plant cells, and thus are often used as model systems in several areas of plant molecular and cellular biology, including secondary metabolism. While the parental tobacco variety synthesizes nicotine as a major alkaloid, the cultured tobacco cells mainly produce a related alkaloid anatabine, instead of nicotine, when elicited with jasmonates. We report here that cultured BY-2 cells scarcely express N-methylputrescine oxidase (MPO) genes even after jasmonate elicitation. MPO is the second enzyme in the biosynthetic pathway that supplies the pyrrolidine moiety of nicotine and nornicotine, but is predicted to be dispensable for the biosynthesis of anatabine, anabasine and anatalline, which do not contain the pyrrolidine moiety. When MPO was overexpressed in tobacco BY-2 cells, nicotine synthesis was dramatically enhanced while anatabine formation was effectively suppressed. As a complementary approach, we suppressed MPO expression by RNA interference in tobacco hairy roots that normally accumulate nicotine. In the MPO-suppressed roots, the contents of anatabine, anabasine and anatalline, as well as N-methylputrescine and putrescine, markedly increased to compensate for suppressed formation of nicotine and nornicotine. These results identify the transcriptional regulation of MPO as a critical rate-limiting step that restricts nicotine formation in cultured tobacco BY-2 cells.

  10. Mammalian cells exposed to ionizing radiation: structural and biochemical aspects

    International Nuclear Information System (INIS)

    Sabanero, M.; Flores V, L. L.; Azorin V, J. C.; Vallejo, M. A.; Cordova F, T.; Sosa A, M.; Castruita D, J. P.; Barbosa S, G.

    2015-10-01

    Acute or chronic exposure to ionizing radiation is a factor that may be hazardous to health. It has been reported that exposure to low doses of radiation (less than 50 mSv / year) and subsequently exposure to high doses have greater effects in people. However, it is unknown molecular and biochemical level alteration. This study, analyzes the susceptibility of a biological system (HeLa Atcc CCL-2 human cervix cancer cell line) to ionizing radiation (6 and 60 mSv/ 90). Our evaluate multiple variables such as: total protein profile, mitochondrial metabolic activity (XTT assay), cell viability (Trypan blue exclusion assay), cytoskeleton (actin micro filaments), nuclei (D API), genomic DNA. The results indicate, that cells exposed to ionizing radiation structurally show alterations in nuclear phenotype and aneuploidy, further disruption in the tight junctions and consequently on the distribution of actin micro filaments. Similar alterations were observed in cells treated with a genotoxic agent (200μM H 2 O 2 /1 h). In conclusion, this multi-criteria assessment enables precise comparisons of the effects of radiation between any biological systems. However, it is necessary to determine stress markers for integration of the effects of ionizing radiation. (Author)

  11. Mammalian cells exposed to ionizing radiation: structural and biochemical aspects

    Energy Technology Data Exchange (ETDEWEB)

    Sabanero, M.; Flores V, L. L. [Universidad de Guanajuato, Departamento de Biologia, DCNE, Noria Alta s/n, 36250 Guanajuato, Gto. (Mexico); Azorin V, J. C.; Vallejo, M. A.; Cordova F, T.; Sosa A, M. [Universidad de Guanajuato, Departamento de Ingenieria Fisica, DCI, Loma del Bosque 103, Col. Lomas del Campestre, 37150 Leon, Guanajuato (Mexico); Castruita D, J. P. [Universidad de Guadalajara, Departamento de Ecologia, CUCBA, Las Agujas, 45100 Zapopan, Jalisco (Mexico); Barbosa S, G., E-mail: myrna.sabanero@gmail.com [Universidad de Guanajuato, Departamento de Ciencias Medicas, DCS, 20 de Enero No. 929, Col. Obregon, 37000 Leon, Guanajuato (Mexico)

    2015-10-15

    Acute or chronic exposure to ionizing radiation is a factor that may be hazardous to health. It has been reported that exposure to low doses of radiation (less than 50 mSv / year) and subsequently exposure to high doses have greater effects in people. However, it is unknown molecular and biochemical level alteration. This study, analyzes the susceptibility of a biological system (HeLa Atcc CCL-2 human cervix cancer cell line) to ionizing radiation (6 and 60 mSv/ 90). Our evaluate multiple variables such as: total protein profile, mitochondrial metabolic activity (XTT assay), cell viability (Trypan blue exclusion assay), cytoskeleton (actin micro filaments), nuclei (D API), genomic DNA. The results indicate, that cells exposed to ionizing radiation structurally show alterations in nuclear phenotype and aneuploidy, further disruption in the tight junctions and consequently on the distribution of actin micro filaments. Similar alterations were observed in cells treated with a genotoxic agent (200μM H{sub 2}O{sub 2}/1 h). In conclusion, this multi-criteria assessment enables precise comparisons of the effects of radiation between any biological systems. However, it is necessary to determine stress markers for integration of the effects of ionizing radiation. (Author)

  12. Characterization of the programmed cell death induced by metabolic products of Alternaria alternata in tobacco BY-2 cells.

    Science.gov (United States)

    Cheng, Dan-Dan; Jia, Yu-Jiao; Gao, Hui-Yuan; Zhang, Li-Tao; Zhang, Zi-Shan; Xue, Zhong-Cai; Meng, Qing-Wei

    2011-02-01

    Alternaria alternata has received considerable attention in current literature and most of the studies are focused on its pathogenic effects on plant chloroplasts, but little is known about the characteristics of programmed cell death (PCD) induced by metabolic products (MP) of A. alternata, the effects of the MP on mitochondrial respiration and its relation to PCD. The purpose of this study was to explore the mechanism of MP-induced PCD in non-green tobacco BY-2 cells and to explore the role of mitochondrial inhibitory processes in the PCD of tobacco BY-2 cells. MP treatment led to significant cell death that was proven to be PCD by the concurrent cytoplasm shrinkage, chromatin condensation and DNA laddering observed in the cells. Moreover, MP treatment resulted in the overproduction of reactive oxygen species (ROS), rapid ATP depletion and a respiratory decline in the tobacco BY-2 cells. It was concluded that the direct inhibition of the mitochondrial electron transport chain (ETC), alternative pathway (AOX) capacity and catalase (CAT) activity by the MP might be the main contributors to the MP-induced ROS burst observed in tobacco BY-2 cells. The addition of adenosine together with the MP significantly inhibited ATP depletion without preventing PCD; however, when the cells were treated with the MP plus CAT, ROS overproduction was blocked and PCD did not occur. The data presented here demonstrate that the ROS burst played an important role in MP-induced PCD in the tobacco BY-2 cells.

  13. Adenine nucleotide depletion from endothelial cells exposed to xanthine oxidase

    International Nuclear Information System (INIS)

    Aalto, T.K.; Raivio, K.O.

    1990-01-01

    Hypoxia causes breakdown of cellular nucleotides, accumulation of hypoxanthine (HX), and conversion of xanthine dehydrogenase into xanthine oxidase (XO). Upon reoxygenation, the HX-XO reaction generates free radicals, one potential mechanism of tissue damage. Because endothelial cells contain XO and are exposed to circulating HX, they are a likely target for damage. We studied the effect of XO and/or HX at physiologically relevant concentrations on nucleotide metabolism of cultured endothelial cells from human umbilical veins. Cells were labeled with [14C]adenine and incubated for up to 6 h with HX, XO, or both, in the absence or presence of serum. Adenine nucleotides from cell extracts and nucleotide breakdown products (HX, xanthine, and urate) from the medium were separated and counted. HX alone had no effect. XO (80 mU/ml) alone caused a 70% (no serum) or 40% (with serum) fall in adenine nucleotides and an equivalent increase of xanthine and urate. The combination of HX and XO caused a 90% (no serum) or 70% (with serum) decrease in nucleotides, decrease in energy charge, and detachment of cells from the culture plate. Nucleotide depletion was not accounted for by proteolytic activity in the XO preparation. Albumin was only half as effective as serum in preventing nucleotide loss. Thus exogenous XO, in the presence of endogenous HX, triggers adenine nucleotide catabolism, but endogenous XO activity is too low to influence nucleotide levels even at high exogenous HX concentrations. Serum limits the catabolic effect of XO and thus protects cells from free radical damage

  14. Genotoxic Effects of Tobacco on Buccal Epithelium: Cell Nuclear Anomalies as Biomarker

    Directory of Open Access Journals (Sweden)

    Sohini Das Biswas

    2014-12-01

    Full Text Available Background: Tobacco use has toxic effects on different organs. This study was carried out to assess the effect of indigenous tobacco both in smoking (bidi and smokeless (gutkha, zarda and khaini forms on buccal cells at chromosomal level, through assessment of different nuclear anomalies as biomarker. Methods:This study was done on people living in Durgapur and its adjacent areas, West Bengal, India during January to July 2011. The samples were collected from 50 smokers (case group, 50 smokeless tobacco consumers or chewers (case group and 50 non-tobacco consumers (control group. Micronucleus assay was used to assess buccal cell nuclear changes. Buccal smears collected from study subjects were prepared on a grease free slide. Prepared slides were observed under light microscope and 2 to 5 fields were observed randomly for counting the different anomalies. In each field, the frequency of each anomaly was assessed in 100 cells and reported with percentage. Results:Chewers had significantly the highest frequency of all nuclear anomalies compared to smokers and healthy controls (HCs. Smokers also had significantly more anomalies compared to HCs. Condensed chromatin (CC, karyolysis (KL and bi-nucleation (BN in chewers and CC, pyknosis and BN in smokers were the most frequent anomalies. KL was significantly more frequent in chewers compared to smokers (59.8 ± 6.4 vs. 24.2 ± 12.4%, P < 0.001, however, the frequency of other nuclear anomalies were not significantly different in these two study groups. Presence of each nuclear anomaly was significantly greater in older ages in all study groups. Conclusion:Tobacco can cause and increase the rate of nuclear anomalies in both smoking and smokeless forms compared to HCs. The genotoxic effects of tobacco on buccal cells are partly age-related. Cell nuclear anomalies in buccal tissue can be used as biomarker indicating the detrimental effects of tobacco.

  15. Transient expression of P-type ATPases in tobacco epidermal cells

    DEFF Research Database (Denmark)

    Pedas, Lisbeth Rosager; Palmgren, Michael Broberg; Lopez Marques, Rosa Laura

    2016-01-01

    Transient expression in tobacco cells is a convenient method for several purposes such as analysis of protein-protein interactions and the subcellular localization of plant proteins. A suspension of Agrobacterium tumefaciens cells carrying the plasmid of interest is injected into the intracellula...... for example protein-protein interaction studies. In this chapter, we describe the procedure to transiently express P-type ATPases in tobacco epidermal cells, with focus on subcellular localization of the protein complexes formed by P4-ATPases and their β-subunits....

  16. Assessment of Organophosphate and Carbamate Pesticide Residues in Cigarette Tobacco with a Novel Cell Biosensor

    Directory of Open Access Journals (Sweden)

    Spiridon Kintzios

    2008-04-01

    Full Text Available The conventional analysis of pesticide residues in analytical commodities, such as tobacco and tobacco products is a labor intensive procedure, since it is necessary to cover a wide range of different chemicals, using a single procedure. Standard analysis methods include extensive sample pretreatment (with solvent extraction and partitioning phases and determination by GC and HPLC to achieve the necessary selectivity and sensitivity for the different classes of compounds under detection. As a consequence, current methods of analysis provide a limited sample capacity. In the present study, we report on the development of a novel cell biosensor for detecting organophosphate and carbamate pesticide residues in tobacco. The sensor is based on neuroblastoma N2a cells and the measurement of changes of the cell membrane potential, according to the working principle of the Bioelectric Recognition Assay (BERA. The presence of pesticide residues is detected by the degree of inhibition of acetylcholine esterase (AChE. The sensor instantly responded to both the organophoshate pesticide chlorpyriphos and the carbamate carbaryl in a concentration-dependent pattern, being able to detect one part per billion (1 ppb. Additionally, tobacco leaf samples (in blended dry form were analyzed with both the novel biosensor and conventional methods, according to a double-blind protocol. Pesticide residues in tobacco samples caused a considerable cell membrane hyperpolarization to neuroblastoma cells immobilized in the sensor, as indicated by the increase of the negative sensor potential, which was clearly distinguishable from the sensor’s response against pesticide-free control samples. The observed response was quite reproducible, with an average variation of +5,6%. Fluorescence microscopy observations showed that treatment of the cells with either chlorpyrifos or carbaryl was associated with increased [Ca2+]cyt . The novel biosensor offers fresh

  17. Exosomes from asbestos-exposed cells modulate gene expression in mesothelial cells.

    Science.gov (United States)

    Munson, Phillip; Lam, Ying-Wai; Dragon, Julie; MacPherson, Maximilian; Shukla, Arti

    2018-03-19

    Asbestos exposure is a determinate cause of many diseases, such as mesothelioma, fibrosis, and lung cancer, and poses a major human health hazard. At this time, there are no identified biomarkers to demarcate asbestos exposure before the presentation of disease and symptoms, and there is only limited understanding of the underlying biology that governs asbestos-induced disease. In our study, we used exosomes, 30-140 nm extracellular vesicles, to gain insight into these knowledge gaps. As inhaled asbestos is first encountered by lung epithelial cells and macrophages, we hypothesize that asbestos-exposed cells secrete exosomes with signature proteomic cargo that can alter the gene expression of mesothelial cells, contributing to disease outcomes like mesothelioma. In the present study using lung epithelial cells (BEAS2B) and macrophages (THP-1), we first show that asbestos exposure causes changes in abundance of some proteins in the exosomes secreted from these cells. Furthermore, exposure of human mesothelial cells (HPM3) to these exosomes resulted in gene expression changes related to epithelial-to-mesenchymal transition and other cancer-related genes. This is the first report to indicate that asbestos-exposed cells secrete exosomes with differentially abundant proteins and that those exosomes have a gene-altering effect on mesothelial cells.-Munson, P., Lam, Y.-W., Dragon, J. MacPherson, M., Shukla, A. Exosomes from asbestos-exposed cells modulate gene expression in mesothelial cells.

  18. Biosynthesis of callose and cellulose by detergent extracts of tobacco cell membranes and quantification of the polymers synthesized in vivo.

    NARCIS (Netherlands)

    Cifuentes Espitia, C.C.; Bulone, V.; Emons, A.M.C.

    2010-01-01

    The conditions that favor the in vitro synthesis of cellulose from tobacco BY-2 cell extracts were determined. The procedure leading to the highest yield of cellulose consisted of incubating digitonin extracts of membranes from 11-day-old tobacco BY-2 cells in the presence of 1 mM UDP-glucose, 8 mM

  19. Vacuolar and cytoskeletal dynamics during elicitor-induced programmed cell death in tobacco BY-2 cells.

    Science.gov (United States)

    Higaki, Takumi; Kadota, Yasuhiro; Goh, Tatsuaki; Hayashi, Teruyuki; Kutsuna, Natsumaro; Sano, Toshio; Hasezawa, Seiichiro; Kuchitsu, Kazuyuki

    2008-09-01

    Responses of plant cells to environmental stresses often involve morphological changes, differentiation and redistribution of various organelles and cytoskeletal network. Tobacco BY-2 cells provide excellent model system for in vivo imaging of these intracellular events. Treatment of the cell cycle-synchronized BY-2 cells with a proteinaceous oomycete elicitor, cryptogein, induces highly synchronous programmed cell death (PCD) and provide a model system to characterize vacuolar and cytoskeletal dynamics during the PCD. Sequential observation revealed dynamic reorganization of the vacuole and actin microfilaments during the execution of the PCD. We further characterized the effects cryptogein on mitotic microtubule organization in cell cycle-synchronized cells. Cryptogein treatment at S phase inhibited formation of the preprophase band, a cortical microtubule band that predicts the cell division site. Cortical microtubules kept their random orientation till their disruption that gradually occurred during the execution of the PCD twelve hours after the cryptogein treatment. Possible molecular mechanisms and physiological roles of the dynamic behavior of the organelles and cytoskeletal network in the pathogenic signal-induced PCD are discussed.

  20. Modification of cell volume and proliferative capacity of Pseudokirchneriella subcapitata cells exposed to metal stress

    International Nuclear Information System (INIS)

    Machado, Manuela D.; Soares, Eduardo V.

    2014-01-01

    Highlights: •Metals induce morphological alterations on P. subcapitata. •Algal cell cycle consists: mother cell growth; cell division, with two nucleus divisions; release of four autospores. •Cu(II) and Cr(VI) arrest cell growth before the first nuclear division. •Cd(II) arrests cell growth after the second nuclear division but before the cytokinesis. •The approach used can be useful in the elucidation of different modes of action of pollutants. -- Abstract: The impact of metals (Cd, Cr, Cu and Zn) on growth, cell volume and cell division of the freshwater alga Pseudokirchneriella subcapitata exposed over a period of 72 h was investigated. The algal cells were exposed to three nominal concentrations of each metal: low (closed to 72 h-EC 10 values), intermediate (closed to 72 h-EC 50 values) and high (upper than 72 h-EC 90 values). The exposure to low metal concentrations resulted in a decrease of cell volume. On the contrary, for the highest metal concentrations an increase of cell volume was observed; this effect was particularly notorious for Cd and less pronounced for Zn. Two behaviours were found when algal cells were exposed to intermediate concentrations of metals: Cu(II) and Cr(VI) induced a reduction of cell volume, while Cd(II) and Zn(II) provoked an opposite effect. The simultaneous nucleus staining and cell image analysis, allowed distinguishing three phases in P. subcapitata cell cycle: growth of mother cell; cell division, which includes two divisions of the nucleus; and, release of four autospores. The exposure of P. subcapitata cells to the highest metal concentrations resulted in the arrest of cell growth before the first nucleus division [for Cr(VI) and Cu(II)] or after the second nucleus division but before the cytokinesis (release of autospores) when exposed to Cd(II). The different impact of metals on algal cell volume and cell-cycle progression, suggests that different toxicity mechanisms underlie the action of different metals

  1. Transient foreign gene expression in chloroplasts of cultured tobacco cells after biolistic delivery of chloroplast vectors.

    Science.gov (United States)

    Daniell, H; Vivekananda, J; Nielsen, B L; Ye, G N; Tewari, K K; Sanford, J C

    1990-01-01

    Expression of chloramphenicol acetyltransferase (cat) by suitable vectors in chloroplasts of cultured tobacco cells, delivered by high-velocity microprojectiles, is reported here. Several chloroplast expression vectors containing bacterial cat genes, placed under the control of either psbA promoter region from pea (pHD series) or rbcL promoter region from maize (pAC series) have been used in this study. In addition, chloroplast expression vectors containing replicon fragments from pea, tobacco, or maize chloroplast DNA have also been tested for efficiency and duration of cat expression in chloroplasts of tobacco cells. Cultured NT1 tobacco cells collected on filter papers were bombarded with tungsten particles coated with pUC118 (negative control), 35S-CAT (nuclear expression vector), pHD312 (repliconless chloroplast expression vector), and pHD407, pACp18, and pACp19 (chloroplast expression vectors with replicon). Sonic extracts of cells bombarded with pUC118 showed no detectable cat activity in the autoradiograms. Nuclear expression of cat reached two-thirds of the maximal 48 hr after bombardment and the maximal at 72 hr. Cells bombarded with chloroplast expression vectors showed a low level of expression until 48 hr of incubation. A dramatic increase in the expression of cat was observed 24 hr after the addition of fresh medium to cultured cells in samples bombarded with pHD407; the repliconless vector pHD312 showed about 50% of this maximal activity. The expression of nuclear cat and the repliconless chloroplast vector decreased after 72 hr, but a high level of chloroplast cat expression was maintained in cells bombarded with pHD407. Organelle-specific expression of cat in appropriate compartments was checked by introducing various plasmid constructions into tobacco protoplasts by electroporation. Although the nuclear expression vector 35S-CAT showed expression of cat, no activity was observed with any chloroplast vectors.

  2. Scale-up of hydrophobin-assisted recombinant protein production in tobacco BY-2 suspension cells.

    Science.gov (United States)

    Reuter, Lauri J; Bailey, Michael J; Joensuu, Jussi J; Ritala, Anneli

    2014-05-01

    Plant suspension cell cultures are emerging as an alternative to mammalian cells for production of complex recombinant proteins. Plant cell cultures provide low production cost, intrinsic safety and adherence to current regulations, but low yields and costly purification technology hinder their commercialization. Fungal hydrophobins have been utilized as fusion tags to improve yields and facilitate efficient low-cost purification by surfactant-based aqueous two-phase separation (ATPS) in plant, fungal and insect cells. In this work, we report the utilization of hydrophobin fusion technology in tobacco bright yellow 2 (BY-2) suspension cell platform and the establishment of pilot-scale propagation and downstream processing including first-step purification by ATPS. Green fluorescent protein-hydrophobin fusion (GFP-HFBI) induced the formation of protein bodies in tobacco suspension cells, thus encapsulating the fusion protein into discrete compartments. Cultivation of the BY-2 suspension cells was scaled up in standard stirred tank bioreactors up to 600 L production volume, with no apparent change in growth kinetics. Subsequently, ATPS was applied to selectively capture the GFP-HFBI product from crude cell lysate, resulting in threefold concentration, good purity and up to 60% recovery. The ATPS was scaled up to 20 L volume, without loss off efficiency. This study provides the first proof of concept for large-scale hydrophobin-assisted production of recombinant proteins in tobacco BY-2 cell suspensions. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  3. Mitochondrial electron transport chain is involved in microcystin-RR induced tobacco BY-2 cells apoptosis.

    Science.gov (United States)

    Huang, Wenmin; Li, Dunhai; Liu, Yongding

    2014-09-01

    Microcystin-RR (MC-RR) has been suggested to induce apoptosis in tobacco BY-2 cells through mitochondrial dysfunction including the loss of mitochondrial membrane potential (ΔΨm). To further elucidate the mechanisms involved in MC-RR induced apoptosis in tobacco BY-2 cells, we have investigated the role of mitochondrial electron transport chain (ETC) as a potential source for reactive oxygen species (ROS). Tobacco BY-2 cells after exposure to MC-RR (60mg/L) displayed apoptotic changes in association with an increased production of ROS and loss of ΔΨm. All of these adverse effects were significantly attenuated by ETC inhibitors including Rotenone (2μmol/L, complex I inhibitor) and antimycin A (0.01μmol/L, complex III inhibitor), but not by thenoyltrifluoroacetone (5μmol/L, complex II inhibitor). These results suggest that mitochondrial ETC plays a key role in mediating MC-RR induced apoptosis in tobacco BY-2 cells through an increased mitochondrial production of ROS. Copyright © 2014. Published by Elsevier B.V.

  4. Cell growth, intracellular calcium concentration and metabolic cooperation measured in cells exposed to 50 Hz electromagnetic fields

    International Nuclear Information System (INIS)

    Skauli, K.S.

    1996-08-01

    Colony-forming efficiency, DNA/protein and DNA/cell were measured in cells exposed to magnetic fields of 0.2 and 1 mT at a frequency of 50 Hz. Intracellular calcium concentrations were measured in cells exposed to 0.3 and 1 mT at 50 Hz. Metabolic cooperation was measured in cells exposed to 1 mT at 50 Hz. No significant effects of the fields were observed. 20 refs., 10 figs

  5. Silicon does not mitigate cell death in cultured tobacco BY-2 cells subjected to salinity without ethylene emission.

    Science.gov (United States)

    Liang, Xiaolei; Wang, Huahua; Hu, Yanfeng; Mao, Lina; Sun, Lili; Dong, Tian; Nan, Wenbin; Bi, Yurong

    2015-02-01

    Silicon induces cell death when ethylene is suppressed in cultured tobacco BY-2 cells. There is a crosstalk between Si and ethylene signaling. Silicon (Si) is beneficial for plant growth. It alleviates both biotic and abiotic stresses in plants. How Si works in plants is still mysterious. This study investigates the mechanism of Si-induced cell death in tobacco BY-2 cell cultures when ethylene is suppressed. Results showed that K2SiO3 alleviated the damage of NaCl stress. Si treatment rapidly increased ethylene emission and the expression of ethylene biosynthesis genes. Treatments with Si + Ag and Si + aminooxyacetic acid (AOA, ethylene biosynthesis inhibitor) reduced the cell growth and increased cell damage. The treatment with Si + Ag induced hydrogen peroxide (H2O2) generation and ultimately cell death. Some nucleus of BY-2 cells treated with Si + Ag appeared TUNEL positive. The inhibition of H2O2 and nitric oxide (NO) production reduced the cell death rate induced by Si + Ag treatment. Si eliminated the up-regulation of alternative pathway by Ag. These data suggest that ethylene plays an important role in Si function in plants. Without ethylene, Si not only failed to enhance plant resistance, but also elevated H2O2 generation and further induced cell death in tobacco BY-2 cells.

  6. Nitric oxide production is not required for dihydrosphingosine-induced cell death in tobacco BY-2 cells.

    Science.gov (United States)

    Da Silva, Daniel; Lachaud, Christophe; Cotelle, Valérie; Brière, Christian; Grat, Sabine; Mazars, Christian; Thuleau, Patrice

    2011-05-01

    Sphinganine or dihydrosphingosine (d18:0, DHS), one of the most abundant free sphingoid Long Chain Base (LCB) in plants, is known to induce a calcium dependent programmed cell death (PCD) in tobacco BY-2 cells. In addition, we have recently shown that DHS triggers a production of H2O2, via the activation of NADPH oxidase(s). However, this production of H2O2 is not correlated with the DHS-induced cell death but would rather be associated with basal cell defense mechanisms. In the present study, we extend our current knowledge of the DHS signaling pathway, by demonstrating that DHS also promotes a production of nitric oxide (NO) in tobacco BY-2 cells. As for H2O2, this NO production is not necessary for cell death induction. 

  7. Potassium ion influx measurements on cultured Chinese hamster cells exposed to 60-hertz electromagnetic fields

    International Nuclear Information System (INIS)

    Stevenson, A.P.; Tobey, R.A.

    1985-01-01

    Potassium ion influx was measured by monitoring 42 KCl uptake by Chinese hamster ovary (CHO) cells grown in suspension culture and exposed in the culture medium to 60-Hz electromagnetic fields up to 2.85 V/m. In the presence of the field CHO cells exhibited two components of uptake, the same as previously observed for those grown under normal conditions; both these components of influx were decreased when compared to sham-exposed cells. Although decreases were consistently observed in exposed cells when plotted as loge of uptake, the differences between the means of the calculated fluxes of exposed and sham-exposed cells were quite small (on the order of 4-7%). When standard deviations were calculated, there was no significant difference between these means; however, when time-paired uptake data were analyzed, the differences were found to be statistically significant. Cells exposed only to the magnetic field exhibited similar small decreases in influx rates when compared to sham-exposed cells, suggesting that the reduction in K+ uptake could be attributed to the magnetic field. Additionally, intracellular K+ levels were measured over a prolonged exposure period (96 h), and no apparent differences in intracellular K+ levels were observed between field-exposed and sham-exposed cultures. These results indicate that high-strength electric fields have a small effect on the rate of transport of potassium ions but no effect on long-term maintenance of intracellular K+

  8. Effects of tobacco-smoke on radiation-induced pneumonitis in rats

    International Nuclear Information System (INIS)

    Nilsson, K.; Henriksson, R.; Cai, Y.-Q.; Hellstroem, S.; Bjermer, L.; Hoernqvist Bylunds, S.

    1992-01-01

    To investigate the effect of exposure to tobacco smoke (TS) on the development of irradiation-induced pneumonitis in rats, five groups of animals were investigated including controls (C), tobacco smoke exposed (S), irradiated (RNS) and irradiated and tobacco smoke exposed (RS). An additional group (RS/NS) was exposed to tobacco before irradiation but not afterwards. Results indicate that smoking suppresses the radiation-induced inflammation but to a lesser degree affects the radiation-induced increase in membrane permeability as reflected by increased protein levels in BAL. Moreover, the marked effects on the numbers of mast cells and neutrophils in the RS group may indicate that these cells play an important role in the mechanism by which tobacco smoke modulates the effects of irradiation. When exposure to tobacco smoke was terminated immediately after irradiation (RS/NS), the inflammatory response was unaffected. (author)

  9. Internalisation of cell-penetrating peptides into tobacco protoplasts.

    Science.gov (United States)

    Mäe, Maarja; Myrberg, Helena; Jiang, Yang; Paves, Heiti; Valkna, Andres; Langel, Ulo

    2005-05-20

    Cells are protected from the surrounding environment by plasma membrane which is impenetrable for most hydrophilic molecules. In the last 10 years cell-penetrating peptides (CPPs) have been discovered and developed. CPPs enter mammalian cells and carry cargo molecules over the plasma membrane with a molecular weight several times their own. Known transformation methods for plant cells have relatively low efficiency and require improvement. The possibility to use CPPs as potential delivery vectors for internalisation in plant cells has been studied in the present work. We analyse and compare the uptake of the fluorescein-labeled CPPs, transportan, TP10, penetratin and pVEC in Bowes human melanoma cells and Nicotiana tabacum cultivar (cv.) SR-1 protoplasts (plant cells without cell wall). We study the internalisation efficiency of CPPs with fluorescence microscopy, spectrofluorometry and fluorescence-activated cell sorter (FACS). All methods indicate, for the first time, that these CPPs can internalise into N. tabacum cv. SR-1 protoplasts. Transportan has the highest uptake efficacy among the studied peptides, both in mammalian cells and plant protoplast. The internalisation of CPPs by plant protoplasts may open up a new effective method for transfection in plants.

  10. Hemicellulose biosynthesis and degradation in tobacco cell walls

    NARCIS (Netherlands)

    Compier, M.G.M.

    2005-01-01

    Natural fibres have a wide range of technological applications, such as in paper and textile industries. The basic properties and the quality of plant fibres are determined by the composition of the plant cell wall. Characteristic for fibres are thick secondary cell walls, which consist of cellulose

  11. Cell culture-induced gradual and frequent epigenetic reprogramming of invertedly repeated tobacco transgene epialleles

    Czech Academy of Sciences Publication Activity Database

    Křížová, Kateřina; Fojtová, Miloslava; Depicker, A.; Kovařík, Aleš

    2009-01-01

    Roč. 149, č. 3 (2009), s. 1493-1504 ISSN 0032-0889 R&D Projects: GA AV ČR(CZ) IAA600040611; GA ČR(CZ) GD204/05/H505; GA ČR(CZ) GA521/07/0116 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : tobacco * cell culture * transgene silencing Subject RIV: BO - Biophysics Impact factor: 6.235, year: 2009

  12. Transient foreign gene expression in chloroplasts of cultured tobacco cells after biolistic delivery of chloroplast vectors.

    OpenAIRE

    Daniell, H; Vivekananda, J; Nielsen, B L; Ye, G N; Tewari, K K; Sanford, J C

    1990-01-01

    Expression of chloramphenicol acetyltransferase (cat) by suitable vectors in chloroplasts of cultured tobacco cells, delivered by high-velocity microprojectiles, is reported here. Several chloroplast expression vectors containing bacterial cat genes, placed under the control of either psbA promoter region from pea (pHD series) or rbcL promoter region from maize (pAC series) have been used in this study. In addition, chloroplast expression vectors containing replicon fragments from pea, tobacc...

  13. Induction of sesquiterpenoid biosynthesis in tobacco cell suspension cultures by fungal elicitor

    International Nuclear Information System (INIS)

    Chappell, J.; Nable, R.

    1987-01-01

    Large amounts of the sesquiterpenoid capsidiol accumulated in the media of tobacco (Nicotiana tabacum L. cv KY14) cell suspension cultures upon addition of fungal elicitor. Capsidiol accumulation was proportional to the amount of elicitor added. The accumulation of capsidiol was preceded by a transient increase in the capsidiol de novo synthesis rate as measured by the incorporation of exogenous [ 14 C]acetate. Changes in 3-hydroxy-3-methylglutaryl-CoA reductase activity, an enzyme of general isoprenoid metabolism, paralleled the changes in [ 14 C]acetate incorporation into capsidiol. Incubation of the cell cultures with mevinolin, a potent in vitro inhibitor of the tobacco HMGR enzyme activity, inhibited the elicitor-induced capsidiol accumulation in a concentration dependent manner. [ 14 C]Acetate incorporation into capsidiol was likewise inhibited by mevinolin treatment. Unexpectedly, [ 3 H] mevalonate incorporation into capsidiol was also partially inhibited by mevinolin, suggesting that mevinolin may effect secondary sites of sesquiterpenoid biosynthesis in vivo beyond HMGR. The data indicated the importance of the induced HMGR activity for capsidiol production in elicitor-treated tobacco cell suspension cultures

  14. Interleukin-16-producing NK cells and T-cells in the blood of tobacco smokers with and without COPD

    Directory of Open Access Journals (Sweden)

    Andersson A

    2016-09-01

    Full Text Available Anders Andersson,1,* Carina Malmhäll,2,* Birgitta Houltz,1 Sara Tengvall,1 Margareta Sjöstrand,2 Ingemar Qvarfordt,1 Anders Lindén,3 Apostolos Bossios2 1Respiratory Medicine and Allergology, Department of Internal Medicine and Clinical Nutrition, Institute of Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden; 2Krefting Research Center, Department of Internal Medicine and Clinical Nutrition, Institute of Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden; 3Unit for Lung and Airway Research, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden *These authors contributed equally to this work Background: Long-term exposure to tobacco smoke causes local inflammation in the airways that involves not only innate immune cells, including NK cells, but also adaptive immune cells such as cytotoxic (CD8+ and helper (CD4+ T-cells. We have previously demonstrated that long-term tobacco smoking increases extracellular concentration of the CD4+-recruiting cytokine interleukin (IL-16 locally in the airways. Here, we hypothesized that tobacco smoking alters IL-16 biology at the systemic level and that this effect involves oxygen free radicals (OFR.Methods: We quantified extracellular IL-16 protein (ELISA and intracellular IL-16 in NK cells, T-cells, B-cells, and monocytes (flow cytometry in blood samples from long-term tobacco smokers with and without chronic obstructive pulmonary disease (COPD and in never-smokers. NK cells from healthy blood donors were stimulated with water-soluble tobacco smoke components (cigarette smoke extract with or without an OFR scavenger (glutathione in vitro and followed by quantification of IL-16 protein.Results: The extracellular concentrations of IL-16 protein in blood did not display any substantial differences between groups. Notably, intracellular IL-16 protein was detected in all types of blood leukocytes. All long-term smokers displayed

  15. Delineating miRNA profile induced by chewing tobacco in oral keratinocytes

    Directory of Open Access Journals (Sweden)

    Mohd Younis Bhat

    2017-10-01

    Full Text Available The major established etiologic risk factor for oral cancer is tobacco (chewed, smoked and snuffed forms. Chewing form of tobacco is predominantly used in India making it the leading cause of oral cancer. Despite being one of the leading causes of oral cancer, the molecular alterations induced by chewing tobacco remains largely unclear. Carcinogenic effect of chewing tobacco is through chronic and not acute exposure. To understand the molecular alterations induced by chewing tobacco, we developed a cell line model where non-neoplastic oral keratinocytes were chronically exposed to chewing tobacco for a period of 6 months. This resulted in increased cellular proliferation and invasive ability of normal oral keratinocytes. Using this cellular model we studied the differential expression of miRNAs associated with chewing tobacco and the altered signaling pathways through which the aberrantly expressed miRNAs affect tumorigenesis. miRNA sequencing  was carried out using Illumina HiSeq 2500 platform  which resulted in the identification of 427 annotated miRNAs of which 10 were significantly dysregulated (≥ 4 fold; p-value ≤ 0.05 in tobacco exposed cells compared to untreated parental cells. To study the altered signaling in oral keratinocytes chronically exposed to chewing tobacco, we employed quantitative proteomics to characterize the dysregulated proteins. Integration of miRNA sequencing data with proteomic data resulted in identification of 36 proven protein targets which (≥1.5 fold; p-value ≤ 0.05 showed expression correlation with the 10 significantly dysregulated miRNAs. Pathway analysis of the dysregulated targets revealed enrichment of interferon signaling and mRNA processing related pathways in the chewing tobacco exposed cells. In addition, we also identified 6 novel miRNA in oral keratinocytes chronically exposed to chewing tobacco extract. Our study provides a framework to understand the oncogenic transformation induced by

  16. Analysis of endoplasmic reticulum of tobacco cells using confocal microscopy

    Czech Academy of Sciences Publication Activity Database

    Radochová, Barbora; Janáček, Jiří; Schwarzerová, K.; Demjénová, E.; Tomori, Z.; Karen, Petr; Kubínová, Lucie

    2005-01-01

    Roč. 24, č. 11 (2005), s. 181-185 ISSN 1580-3139 R&D Projects: GA AV ČR(CZ) KJB6011309 Institutional research plan: CEZ:AV0Z50110509 Keywords : confocal microscopy * endoplasmic reticulum * image analysis Subject RIV: EA - Cell Biology

  17. Evidence for alteration of the membrane-bound ribosomes in Micrococcus luteus cells exposed to lead

    Energy Technology Data Exchange (ETDEWEB)

    Barrow, W; Himmel, M; Squire, P G; Tornabene, T G

    1978-01-01

    Micrococcus luteus cells exposed to Pb(NO/sub 3/)/sub 2/ contained cytosol ribosomal particles and disaggregated membranal ribosomal particles as determined by ultracentrifugation and spectral studies. Approximately 60% of the membrane ribosome fraction from lead exposed cells had a sedimentation value of 8.4S. Cytosol ribosome from lead exposed cells as well as membranal and cytosol ribosomes from control cells were comparable by their contents of predominantly the 70S type with the 50S and 100S present in relatively small amounts. The lead content of the 8.4S components was more than 200 times higher than the components with higher sedimentation coefficients from lead exposed cells and approximately 650 times more than that of control cell ribosomes. The cells exposed to lead, however, showed no adverse effects from the lead in respect to their growth rates and cellular yields. These results indicate that lead is interacting only at specific sites of the membrane and is inducing events initiated only in strategic cellular regions. These data further substantiate that subtle changes do occur in lead exposed cells that show no obvious effects. It is assumed that these minor alterations are, in toto, biologically significant. 24 references, 2 figures, 1 table.

  18. B cells exposed to enterobacterial components suppress development of experimental colitis

    DEFF Research Database (Denmark)

    Schmidt, Esben Gjerløff Wedebye; Larsen, Hjalte List; Kristensen, Nanna Ny

    2012-01-01

    ). RESULTS: We demonstrate that splenic B cells exposed to ebx produce large amounts of IL-10 in vitro and express CD1d and CD5 previously known to be associated with regulatory B cells. In SCID mice transplanted with colitogenic CD4(+) CD25(-) T cells, co-transfer of ebx-B cells significantly suppressed...... development of colitis. Suppression was dependent on B cell-derived IL-10, as co-transfer of IL-10 knockout ebx-B cells failed to suppress colitis. Ebx-B cell-mediated suppression of colitis was associated with a decrease in interferon gamma (IFN-¿)-producing T(H) 1 cells and increased frequencies of Foxp3......-expressing T cells. CONCLUSIONS: These data demonstrate that splenic B cells exposed to enterobacterial components acquire immunosuppressive functions by which they can suppress development of experimental T cell-mediated colitis in an IL-10-dependent way. (Inflamm Bowel Dis 2011;)....

  19. Proliferation of human mammary cancer cells exposed to 27-hydroxycholesterol

    OpenAIRE

    CRUZ, PAMELA; TORRES, CRISTIAN; RAMÍREZ, MARÍA EUGENIA; EPUÑÁN, MARÍA JOSÉ; VALLADARES, LUIS EMILIO; SIERRALTA, WALTER DANIEL

    2010-01-01

    The aim of the present study was to identify the possible mechanisms by which certain estradiol receptor (ER)-positive mammary tumor cells remain resistant to treatment with anti-estrogens or inhibitors of local estradiol (E2) production. To this end, we compared the proliferative effects on mammary cancer cells of the novel selective ER modulator 27-hydroxycholesterol (27OHC) to those of E2, and evaluated their inhibition by ICI 182,780 (ICI). Analysis of the effects on the cell cycle of 27O...

  20. Proteins associated with adaptation of cultured tobacco cells to NaCl

    International Nuclear Information System (INIS)

    Singh, N.K.; Handa, A.K.; Hasegawa, P.M.; Bressan, R.A.

    1985-01-01

    Cultured tobacco cells (Nicotiana tabacum L. cv Wisconsin 38) adapted to grow in medium containing high levels of NaCl or polyethylene glycol (PEG) produce several new or enhanced polypeptide bands on sodium dodecyl sulfate-polyarylamide gel electrophoresis. The intensities of some of the polypeptide bands increase with increasing levels of NaCl adaptation, while the intensities of other polypeptide bands are reduced. Synthesis of 26-kilodalton polypeptide(s) occurs at two different periods during culture growth of NaCl adapted cells. Unadapted cells also incorporate 35 S into a 26-kilodalton polypeptide during the later stage of culture growth beginning at midlog phase. The 26-kilodalton polypeptides from adapted and unadapted cells have similar partial proteolysis peptide maps and are immunologically cross-reactive. During adaptation to NaCl, unadapted cells synthesize and accumulate a major 26-kilodalton polypeptide, and the beginning of synthesis corresponds to the period of osmotic adjustment and culture growth. From their results, the authors suggest an involvement of the 26-kilodalton polypeptide in the adaptation of cultured tobacco cells to NaCl and water stress. 38 references, 11 figures, 2 tables

  1. Ergosterol-induced sesquiterpenoid synthesis in tobacco Cells

    CSIR Research Space (South Africa)

    Tugizimana, F

    2012-02-01

    Full Text Available below [46,47]. 2.2.1. HPTLC Fractionation of Extracts and Initial Characterisation HPTLC analysis of the chloroform extracts allowed partial characterisation of the multi-component cell extracts. The image of the HPTLC plate (Figure 3) shows the six.... The corrected empirical formula is thus C15H22O; and searching in databases (structural correlation to MS spectrum), the compound was putatively identified as solavetivone (C15H22O, 218.340 Da). Molecules 2012, 17 1706 Figure 5. The identification...

  2. Cotton Ascorbate Oxidase Promotes Cell Growth in Cultured Tobacco Bright Yellow-2 Cells through Generation of Apoplast Oxidation

    Science.gov (United States)

    Li, Rong; Xin, Shan; Tao, Chengcheng; Jin, Xiang; Li, Hongbin

    2017-01-01

    Ascorbate oxidase (AO) plays an important role in cell growth through the modulation of reduction/oxidation (redox) control of the apoplast. Here, a cotton (Gossypium hirsutum) apoplastic ascorbate oxidase gene (GhAO1) was obtained from fast elongating fiber tissues. GhAO1 belongs to the multicopper oxidase (MCO) family and includes a signal peptide and several transmembrane regions. Analyses of quantitative real-time polymerase chain reaction (QRT-PCR) and enzyme activity showed that GhAO1 was expressed abundantly in 15-day post-anthesis (dpa) wild-type (WT) fibers in comparison with fuzzless-lintless (fl) mutant ovules. Subcellular distribution analysis in onion cells demonstrated that GhAO1 is localized in the cell wall. In transgenic tobacco bright yellow-2 (BY-2) cells with ectopic overexpression of GhAO1, the enhancement of cell growth with 1.52-fold increase in length versus controls was indicated, as well as the enrichment of both total ascorbate in whole-cells and dehydroascorbate acid (DHA) in apoplasts. In addition, promoted activities of AO and monodehydroascorbate reductase (MDAR) in apoplasts and dehydroascorbate reductase (DHAR) in whole-cells were displayed in transgenic tobacco BY-2 cells. Accumulation of H2O2, and influenced expressions of Ca2+ channel genes with the activation of NtMPK9 and NtCPK5 and the suppression of NtTPC1B were also demonstrated in transgenic tobacco BY-2 cells. Finally, significant induced expression of the tobacco NtAO gene in WT BY-2 cells under indole-3-acetic acid (IAA) treatment appeared; however, the sensitivity of the NtAO gene expression to IAA disappeared in transgenic BY-2 cells, revealing that the regulated expression of the AO gene is under the control of IAA. Taken together, these results provide evidence that GhAO1 plays an important role in fiber cell elongation and may promote cell growth by generating the oxidation of apoplasts, via the auxin-mediated signaling pathway. PMID:28644407

  3. Mitochondrial DNA damage and oxidative damage in HL-60 cells exposed to 900 MHz radiofrequency fields

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Yulong; Zong, Lin; Gao, Zhen [School of Public Health, Soochow University, Suzhou, Jiangsu Province (China); Zhu, Shunxing [Laboratory Animal Center, Nantong University, Nantong, Jiangsu Province (China); Tong, Jian [School of Public Health, Soochow University, Suzhou, Jiangsu Province (China); Cao, Yi, E-mail: yicao@suda.edu.cn [School of Public Health, Soochow University, Suzhou, Jiangsu Province (China)

    2017-03-15

    Highlights: • Increased reactive oxygen species. • Decreased mitochondrial transcription Factor A and polymerase gamma. • Decreased mitochondrial transcripts (ND1 and 16S) and mtDNA copy number. • Increased 8-hydroxy-2′deoxyguanosine. • Decreased adenosine triphosphate. - Abstract: HL-60 cells, derived from human promyelocytic leukemia, were exposed to continuous wave 900 MHz radiofrequency fields (RF) at 120 μW/cm{sup 2} power intensity for 4 h/day for 5 consecutive days to examine whether such exposure is capable damaging the mitochondrial DNA (mtDNA) mediated through the production of reactive oxygen species (ROS). In addition, the effect of RF exposure was examined on 8-hydroxy-2′-dexoyguanosine (8-OHdG) which is a biomarker for oxidative damage and on the mitochondrial synthesis of adenosine triphosphate (ATP) which is the energy required for cellular functions. The results indicated a significant increase in ROS and significant decreases in mitochondrial transcription factor A, mtDNA polymerase gamma, mtDNA transcripts and mtDNA copy number in RF-exposed cells compared with those in sham-exposed control cells. In addition, there was a significant increase in 8-OHdG and a significant decrease in ATP in RF-exposed cells. The response in positive control cells exposed to gamma radiation (GR, which is also known to induce ROS) was similar to those in RF-exposed cells. Thus, the overall data indicated that RF exposure was capable of inducing mtDNA damage mediated through ROS pathway which also induced oxidative damage. Prior-treatment of RF- and GR-exposed the cells with melatonin, a well-known free radical scavenger, reversed the effects observed in RF-exposed cells.

  4. Proteinaceous and oligosaccharidic elicitors induce different calcium signatures in the nucleus of tobacco cells.

    Science.gov (United States)

    Lecourieux, David; Lamotte, Olivier; Bourque, Stéphane; Wendehenne, David; Mazars, Christian; Ranjeva, Raoul; Pugin, Alain

    2005-12-01

    We previously reported elevated cytosolic calcium levels in tobacco cells in response to elicitors [D. Lecourieux, C. Mazars, N. Pauly, R. Ranjeva, A. Pugin, Analysis and effects of cytosolic free calcium elevations in response to elicitors in Nicotiana plumbaginifolia cells, Plant Cell 14 (2002) 2627-2641]. These data suggested that in response to elicitors, Ca2+, as a second messenger, was involved in both systemic acquired resistance (RSA) and/or hypersensitive response (HR) depending on calcium signature. Here, we used transformed tobacco cells with apoaequorin expressed in the nucleus to monitor changes in free nuclear calcium concentrations ([Ca2+](nuc)) in response to elicitors. Two types of elicitors are compared: proteins leading to necrosis including four elicitins and harpin, and non-necrotic elicitors including flagellin (flg22) and two oligosaccharidic elicitors, namely the oligogalacturonides (OGs) and the beta-1,3-glucan laminarin. Our data indicate that the proteinaceous elicitors induced a pronounced and sustainable [Ca2+](nuc) elevation, relative to the small effects of oligosaccharidic elicitors. This [Ca2+](nuc) elevation, which seems insufficient to induce cell death, is unlikely to result directly from the diffusion of calcium from the cytosol. The [Ca2+](nuc) rise depends on free cytosolic calcium, IP3, and active oxygen species (AOS) but is independent of nitric oxide.

  5. Proliferation of human mammary cancer cells exposed to 27-hydroxycholesterol.

    Science.gov (United States)

    Cruz, Pamela; Torres, Cristian; Ramírez, María Eugenia; Epuñán, María José; Valladares, Luis Emilio; Sierralta, Walter Daniel

    2010-05-01

    The aim of the present study was to identify the possible mechanisms by which certain estradiol receptor (ER)-positive mammary tumor cells remain resistant to treatment with anti-estrogens or inhibitors of local estradiol (E(2)) production. To this end, we compared the proliferative effects on mammary cancer cells of the novel selective ER modulator 27-hydroxycholesterol (27OHC) to those of E(2), and evaluated their inhibition by ICI 182,780 (ICI). Analysis of the effects on the cell cycle of 27OHC and E(2) in the absence or presence of ICI was conducted. In ER-positive mammary tumor cells, we detected the blocking of 27OHC proliferation-stimulatory activity by simvastatin, as well as the inhibition of E(2)-stimulated proliferation by an α-fetoprotein-derived cyclic nonapeptide. The effects reported herein may be extrapolated to infiltrating mammary cancer, where the activity of local macrophages may stimulate tumor growth. We suggest that increased breast cancer growth in obese patients may be related to increased 27OHC circulatory levels.

  6. 1,8-cineole inhibits both proliferation and elongation of BY-2 cultured tobacco cells.

    Science.gov (United States)

    Yoshimura, Hiroko; Sawai, Yu; Tamotsu, Satoshi; Sakai, Atsushi

    2011-03-01

    Volatile monoterpenes such as 1,8-cineole inhibit the growth of Brassica campestris seedlings in a dose-dependent manner, and the growth-inhibitory effects are more severe for roots than hypocotyls. The preferential inhibition of root growth may be explained if the compounds inhibit cell proliferation more severely than cell elongation because root growth requires both elongation and proliferation of the constituent cells, whereas hypocotyl growth depends exclusively on elongation of existing cells. In order to examine this possibility, BY-2 suspension-cultured tobacco (Nicotiana tabacum) cells were treated with 1,8-cineole, and the inhibitory effects on cell proliferation and on cell elongation were assessed quantitatively. Treatment with 1,8-cineole lowered both the mitotic index and elongation of the cells in a dose-dependent manner, and the half-maximal inhibitory concentration (IC₅₀) for cell elongation was lower than that for cell proliferation. Moreover, 1,8-cineole also inhibited starch synthesis, with IC₅₀ lower than that for cell proliferation. Thus, the inhibitory effects of 1,8-cineole were not specific to cell proliferation; rather, 1,8-cineole seemed inhibitory to a variety of physiological activities when it was in direct contact with target cells. Based on these results, possible mechanisms for the mode of action of 1,8-cineole and for its preferential inhibition on root growth are discussed.

  7. Actin filaments regulate the adhesion between the plasma membrane and the cell wall of tobacco guard cells.

    Science.gov (United States)

    Yu, Qin; Ren, Jing-Jing; Kong, Lan-Jing; Wang, Xiu-Ling

    2018-01-01

    During the opening and closing of stomata, guard cells undergo rapid and reversible changes in their volume and shape, which affects the adhesion of the plasma membrane (PM) to the cell wall (CW). The dynamics of actin filaments in guard cells are involved in stomatal movement by regulating structural changes and intracellular signaling. However, it is unclear whether actin dynamics regulate the adhesion of the PM to the CW. In this study, we investigated the relationship between actin dynamics and PM-CW adhesion by the hyperosmotic-induced plasmolysis of tobacco guard cells. We found that actin filaments in guard cells were depolymerized during mannitol-induced plasmolysis. The inhibition of actin dynamics by treatment with latrunculin B or jasplakinolide and the disruption of the adhesion between the PM and the CW by treatment with RGDS peptide (Arg-Gly-Asp-Ser) enhanced guard cell plasmolysis. However, treatment with latrunculin B alleviated the RGDS peptide-induced plasmolysis and endocytosis. Our results reveal that the actin depolymerization is involved in the regulation of the PW-CW adhesion during hyperosmotic-induced plasmolysis in tobacco guard cells.

  8. Interactions between lipids and proteins are critical for organization of plasma membrane-ordered domains in tobacco BY-2 cells.

    Science.gov (United States)

    Grosjean, Kevin; Der, Christophe; Robert, Franck; Thomas, Dominique; Mongrand, Sébastien; Simon-Plas, Françoise; Gerbeau-Pissot, Patricia

    2018-06-27

    The laterally heterogeneous plant plasma membrane (PM) is organized into finely controlled specialized areas that include membrane-ordered domains. Recently, the spatial distribution of such domains within the PM has been identified as playing a key role in cell responses to environmental challenges. To examine membrane order at a local level, BY-2 tobacco suspension cell PMs were labelled with an environment-sensitive probe (di-4-ANEPPDHQ). Four experimental models were compared to identify mechanisms and cell components involved in short-term (1 h) maintenance of the ordered domain organization in steady-state cell PMs: modulation of the cytoskeleton or the cell wall integrity of tobacco BY-2 cells; and formation of giant vesicles using either a lipid mixture of tobacco BY-2 cell PMs or the original lipid and protein combinations of the tobacco BY-2 cell PM. Whilst inhibiting phosphorylation or disrupting either the cytoskeleton or the cell wall had no observable effects, we found that lipids and proteins significantly modified both the abundance and spatial distribution of ordered domains. This indicates the involvement of intrinsic membrane components in the local physical state of the plant PM. Our findings support a major role for the 'lipid raft' model, defined as the sterol-dependent ordered assemblies of specific lipids and proteins in plant PM organization.

  9. Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform.

    Science.gov (United States)

    Mitchell, Claire A; Kalies, Stefan; Cizmár, Tomás; Heisterkamp, Alexander; Torrance, Lesley; Roberts, Alison G; Gunn-Moore, Frank J; Dholakia, Kishan

    2013-01-01

    A tightly-focused ultrashort pulsed laser beam incident upon a cell membrane has previously been shown to transiently increase cell membrane permeability while maintaining the viability of the cell, a technique known as photoporation. This permeability can be used to aid the passage of membrane-impermeable biologically-relevant substances such as dyes, proteins and nucleic acids into the cell. Ultrashort-pulsed lasers have proven to be indispensable for photoporating mammalian cells but they have rarely been applied to plant cells due to their larger sizes and rigid and thick cell walls, which significantly hinders the intracellular delivery of exogenous substances. Here we demonstrate and quantify femtosecond optical injection of membrane impermeable dyes into intact BY-2 tobacco plant cells growing in culture, investigating both optical and biological parameters. Specifically, we show that the long axial extent of a propagation invariant ("diffraction-free") Bessel beam, which relaxes the requirements for tight focusing on the cell membrane, outperforms a standard Gaussian photoporation beam, achieving up to 70% optoinjection efficiency. Studies on the osmotic effects of culture media show that a hypertonic extracellular medium was found to be necessary to reduce turgor pressure and facilitate molecular entry into the cells.

  10. Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform.

    Directory of Open Access Journals (Sweden)

    Claire A Mitchell

    Full Text Available A tightly-focused ultrashort pulsed laser beam incident upon a cell membrane has previously been shown to transiently increase cell membrane permeability while maintaining the viability of the cell, a technique known as photoporation. This permeability can be used to aid the passage of membrane-impermeable biologically-relevant substances such as dyes, proteins and nucleic acids into the cell. Ultrashort-pulsed lasers have proven to be indispensable for photoporating mammalian cells but they have rarely been applied to plant cells due to their larger sizes and rigid and thick cell walls, which significantly hinders the intracellular delivery of exogenous substances. Here we demonstrate and quantify femtosecond optical injection of membrane impermeable dyes into intact BY-2 tobacco plant cells growing in culture, investigating both optical and biological parameters. Specifically, we show that the long axial extent of a propagation invariant ("diffraction-free" Bessel beam, which relaxes the requirements for tight focusing on the cell membrane, outperforms a standard Gaussian photoporation beam, achieving up to 70% optoinjection efficiency. Studies on the osmotic effects of culture media show that a hypertonic extracellular medium was found to be necessary to reduce turgor pressure and facilitate molecular entry into the cells.

  11. Role of peroxynitrite in the responses induced by heat stress in tobacco BY-2 cultured cells.

    Science.gov (United States)

    Malerba, Massimo; Cerana, Raffaella

    2018-07-01

    Temperatures above the optimum are sensed as heat stress (HS) by all living organisms and represent one of the major environmental challenges for plants. Plants can cope with HS by activating specific defense mechanisms to minimize damage and ensure cellular functionality. One of the most common effects of HS is the overproduction of reactive oxygen and nitrogen species (ROS and RNS). The role of ROS and RNS in the regulation of many plant physiological processes is well established. On the contrary, in plants very little is known about the physiological role of peroxynitrite (ONOO - ), the RNS species generated by the interaction between NO and O 2 - . In this work, the role of ONOO - on some of the stress responses induced by HS in tobacco BY-2 cultured cells has been investigated by measuring these responses both in the presence and in the absence of 2,6,8-trihydroxypurine (urate), a specific scavenger of ONOO - . The obtained results suggest a potential role for ONOO - in some of the responses induced by HS in tobacco cultured cells. In particular, ONOO - seems implicated in a form of cell death showing apoptotic features and in the regulation of the levels of proteins involved in the response to stress.

  12. Cell damage of hepatoma-22 cells exposed to continuous wave ultrasound.

    Science.gov (United States)

    Wang, Pan; Wang, Xiaobing; Liu, Quanhong

    2012-01-01

    processes. Moreover the mechanical effect might also be involved in inducing cell damage because there was significant mitochondria membrane potential loss and no visible ROS detection when cells were exposed to ultrasound for 30 s.

  13. Mitochondrial oxidative stress in human hepatoma cells exposed to stavudine

    International Nuclear Information System (INIS)

    Velsor, Leonard W.; Kovacevic, Miro; Goldstein, Mark; Leitner, Heather M.; Lewis, William; Day, Brian J.

    2004-01-01

    The toxicity of nucleoside reverse transcriptase inhibitors (NRTIs) is linked to altered mitochondrial DNA (mtDNA) replication and subsequent disruption of cellular energetics. This manifests clinically as elevated concentrations of lactate in plasma. The mechanism(s) underlying how the changes in mtDNA replication lead to lactic acidosis remains unclear. It is hypothesized that mitochondrial oxidative stress links the changes in mtDNA replication to mitochondrial dysfunction and ensuing NRTIs toxicity. To test this hypothesis, changes in mitochondrial function, mtDNA amplification efficiency, and oxidative stress were assessed in HepG2-cultured human hepatoblasts treated with the NRTI stavudine (2',3'-didehydro-2',3'-deoxythymidine or d4T) for 48 h. d4T produced significant mitochondrial dysfunction with a 1.5-fold increase in cellular lactate to pyruvate ratios. In addition, d4T caused a dose-dependent decrease in mtDNA amplification and a correlative increase in abundance of markers of mitochondrial oxidative stress. Manganese (III) meso-tetrakis (4-benzoic acid) porphyrin, MnTBAP, a catalytic antioxidant, ameliorated or reversed d4T-induced changes in cell injury, energetics, mtDNA amplification, and mitochondrial oxidative stress. In conclusion, d4T treatment elevates mitochondrial reactive oxygen species (ROS), enhances mitochondrial oxidative stress, and contributes mechanistically to NRTI-induced toxicity. These deleterious events may be potentiated in acquired immunodeficiency syndrome (AIDS) by human immunodeficiency virus (HIV) infection itself, coinfection (e.g., viral hepatitis), aging, substance, and alcohol use

  14. Cerebrospinal fluid cells and proteins in patients occupationally exposed to organic solvents

    Energy Technology Data Exchange (ETDEWEB)

    Juntunen, J; Taskinen, E; Luisto, M; Iivanainen, M; Nurminen, M

    1982-06-01

    Cerebrospinal fluid (CSF) cells and proteins were determined for 33 patients exposed to industrial organic solvents. A lymphoid reaction, i.e., a pathologically elevated number or percentage of enlarged lymphoid cells was observed in one-third of the patients, more often in patients with chronic intoxication (40%) than in those currently exposed to organic solvents (32%). An almost significant decrease of small lymphocytes in the CSF was observed among patients who had a past history of chronic solvent intoxication but no recent exposure. No cytological evidence of tissue destruction was found. Signs of slight blood--CSF barrier damage occurred in 5 (23%) of the currently exposed patients, but intrathecal IgG synthesis was not observed. Increased cellular activity in the CSF was also accentuated in principal component analysis. The results suggest slight nonspecific immunoactivation in the central nervous system of subjects exposed to organic solvents.

  15. Hyaluronan synthesis in cultured tobacco cells (BY-2) expressing a chlorovirus enzyme: cytological studies.

    Science.gov (United States)

    Rakkhumkaew, Numfon; Shibatani, Shigeo; Kawasaki, Takeru; Fujie, Makoto; Yamada, Takashi

    2013-04-01

    Extraction of hyaluronan from animals or microbial fermentation has risks including contamination with pathogens and microbial toxins. In this work, tobacco cultured-cells (BY-2) were successfully transformed with a chloroviral hyaluronan synthase (cvHAS) gene to produce hyaluronan. Cytological studies revealed accumulation of HA on the cells, and also in subcellular fractions (protoplasts, miniplasts, vacuoplasts, and vacuoles). Transgenic BY-2 cells harboring a vSPO-cvHAS construct containing the vacuolar targeting signal of sporamin connected to the N-terminus of cvHAS accumulated significant amounts of HA in vacuoles. These results suggested that cvHAS successfully functions on the vacuolar membrane and synthesizes/transports HA into vacuoles. Efficient synthesis of HA using this system provides a new method for practical production of HA. Copyright © 2012 Wiley Periodicals, Inc.

  16. Evaluation of cytomorphometric changes in tobacco users and diagnosed oral squamous cell carcinoma individuals

    Directory of Open Access Journals (Sweden)

    Urmila Udayashankar

    2016-01-01

    Full Text Available Aims: To determine the cellular and nuclear area of keratinocytes in smears obtained from the oral mucosa of tobacco users, those with oral squamous cell carcinoma (OSCC, and from normal healthy persons and resolve if any significant difference exists in these three groups. Materials and Methods: The study group comprised 100 subjects 20 controls, (40 OSCC patients-20 from lesional sites and 20 from nonlesional sites, 20 tobacco smokers and 20 tobacco chewers in the age group of 25-75 years. Oral mucosal smears obtained by using a cytobrush were stained with Papanicolaou (PAP stain and using 20X objective in trinocular Olympus model BX53 with Jenoptik scientific grade-dedicated microphotographic camera images were taken. With ProgRes version 8.0 image analysis software, 20 cells with defined borders were evaluated from each slide. Finally, one-way analysis of variance (ANOVA was used to compare the above parameters in the studied groups. Statistical Analysis Used: Minitab and Excel software were used to analyze the data. One-way ANOVA was used to compare the above parameters in the studied groups. Results: The mean value of the cell area for groups I, II, III, IV, and V were 2838 ± 275.2, 2762.1 ± 511.4, 2861.9 ± 512.9, 2643.8 ± 333.3, and 3064.3 ± 362.7, respectively, the nuclear area (NA was 83.88 ± 9.86, 106.19 ± 13.45, 95.11 ± 14.24, 85.55 ± 21.11, and 80.83 ± 13.45, respectively, and nuclear-to-cellular (N:C ratio was 0.0297, 0.03924, 0.0337, 0.03257, and 0.02678, respectively. Conclusions: Thus, our study elucidates that cytomorphology gauges the effect of tobacco on the oral mucosa and possibly establishes a link between premalignant and malignant transformations even before a lesion is visibly noted.

  17. Comparing genotoxic signatures in cord blood cells from neonates exposed in utero to zidovudine or tenofovir.

    Science.gov (United States)

    Vivanti, Alexandre; Soheili, Tayebeh S; Cuccuini, Wendy; Luce, Sonia; Mandelbrot, Laurent; Lechenadec, Jerome; Cordier, Anne-Gael; Azria, Elie; Soulier, Jean; Cavazzana, Marina; Blanche, Stéphane; André-Schmutz, Isabelle

    2015-07-17

    Zidovudine and tenofovir are the two main nucleos(t)ide analogs used to prevent mother-to-child transmission of HIV. In vitro, both drugs bind to and integrate into human DNA and inhibit telomerase. The objective of the present study was to assess the genotoxic effects of either zidovudine or tenofovir-based combination therapies on cord blood cells in newborns exposed in utero. We compared the aneuploid rate and the gene expression profiles in cord blood samples from newborns exposed either to zidovudine or tenofovir-based combination therapies during pregnancy and from unexposed controls (n = 8, 9, and 8, respectively). The aneuploidy rate was measured on the cord blood T-cell karyotype. Gene expression profiles of cord blood T cells and hematopoietic stem and progenitor cells were determined with microarrays, analyzed in a gene set enrichment analysis and confirmed by real-time quantitative PCRs. Aneuploidy was more frequent in the zidovudine-exposed group (26.3%) than in the tenofovir-exposed group (14.2%) or in controls (13.3%; P < 0.05 for both). The transcription of genes involved in DNA repair, telomere maintenance, nucleotide metabolism, DNA/RNA synthesis, and the cell cycle was deregulated in samples from both the zidovudine and the tenofovir-exposed groups. Although tenofovir has a lower clastogenic impact than zidovudine, gene expression profiling showed that both drugs alter the transcription of DNA repair and telomere maintenance genes.

  18. B-cell infiltration in the respiratory mucosa of turkeys exposed to subtype C avian metapneumovirus.

    Science.gov (United States)

    Cha, Ra Mi; Khatri, Mahesh; Sharma, Jagdev M

    2007-09-01

    Turkeys exposed to avian metapneumovirus (aMPV) subtype C showed extensive lymphoid cell infiltrations in the nasal turbinates of the upper respiratory tract. The cellular infiltration occurred after the first virus exposure but not after re-exposure. Quantitation of the relative proportions of mucosal immunoglobulin (Ig)A+, IgG+, and IgM+ cells in controls and virus-exposed turkeys revealed that at 7 days after the first virus exposure, when mucosal infiltration was well pronounced, there was a significant increase (P < 0.05) in the numbers of infiltrating IgA+ but not of IgG+ and IgM+ cells. After the second virus exposure, although the overall numbers of mucosal lymphoid cells were similar in the virus-exposed and control turkeys, the relative proportions of IgA+ and IgG+ cells were significantly higher in the virus-exposed turkeys (P < 0.05) than in controls. Furthermore, elevated levels of aMPV-specific IgA were detected in the nasal secretions and the bile of virus-exposed birds after the second but not after the first virus exposure. These results suggest, for the first time, the possible involvement of local mucosal immunoglobulins in the pathogenesis of aMPV in turkeys.

  19. Estrogen enhanced cell-cell signalling in breast cancer cells exposed to targeted irradiation

    International Nuclear Information System (INIS)

    Shao, Chunlin; Folkard, Melvyn; Held, Kathryn D; Prise, Kevin M

    2008-01-01

    Radiation-induced bystander responses, where cells respond to their neighbours being irradiated are being extensively studied. Although evidence shows that bystander responses can be induced in many types of cells, it is not known whether there is a radiation-induced bystander effect in breast cancer cells, where the radiosensitivity may be dependent on the role of the cellular estrogen receptor (ER). This study investigated radiation-induced bystander responses in estrogen receptor-positive MCF-7 and estrogen receptor-negative MDA-MB-231 breast cancer cells. The influence of estrogen and anti-estrogen treatments on the bystander response was determined by individually irradiating a fraction of cells within the population with a precise number of helium-3 using a charged particle microbeam. Damage was scored as chromosomal damage measured as micronucleus formation. A bystander response measured as increased yield of micronucleated cells was triggered in both MCF-7 and MDA-MB-231 cells. The contribution of the bystander response to total cell damage in MCF-7 cells was higher than that in MDA-MB-231 cells although the radiosensitivity of MDA-MB-231 was higher than MCF-7. Treatment of cells with 17β-estradiol (E2) increased the radiosensitivity and the bystander response in MCF-7 cells, and the effect was diminished by anti-estrogen tamoxifen (TAM). E2 also increased the level of intracellular reactive oxygen species (ROS) in MCF-7 cells in the absence of radiation. In contrast, E2 and TAM had no influence on the bystander response and ROS levels in MDA-MB-231 cells. Moreover, the treatment of MCF-7 cells with antioxidants eliminated both the E2-induced ROS increase and E2-enhanced bystander response triggered by the microbeam irradiation, which indicates that ROS are involved in the E2-enhanced bystander micronuclei formation after microbeam irradiation. The observation of bystander responses in breast tumour cells may offer new potential targets for radiation

  20. In vitro cell-mediated immunity studies of plutonium-exposed beagle dogs

    International Nuclear Information System (INIS)

    Morris, J.E.; Graham, T.; Park, J.F.

    1980-01-01

    Mitogen-induced activation was measured in spleen and mesenteric lymph node cell preparations from dogs exposed to a single inhalation exposure of plutonium oxide ( 238 Pu or 239 Pu). Reduced stimulation indices of splenic lymphocytes from exposed animals suggest that a reduction in lymphocyte function has occurred in this tissue. No apparent reduction in mitogen stimulation indices was observed in mesenteric lymph node cultures

  1. Histone H3 is absent from organelle nucleoids in BY-2 cultured tobacco cells.

    Science.gov (United States)

    Takusagawa, Mari; Tamotsu, Satoshi; Sakai, Atsushi

    2013-07-01

    The core histone proteins (H2A, H2B, H3 and H4) are nuclear-localised proteins that play a central role in the formation of nucleosome structure. They have long been considered to be absent from extra-nuclear, DNA-containing organelles; that is plastids and mitochondria. Recently, however, the targeting of core histone H3 to mitochondria, and the presence of nucleosome-like structures in mitochondrial nucleoids, were proposed in cauliflower and tobacco respectively. Thus, we examined whether histone H3 was present in plant organelles and participated in the organisation of nucleoid structure, using highly purified organelles and organelle nucleoids isolated from BY-2 cultured tobacco cells. Immunofluorescence microscopic observations and Western blotting analyses demonstrated that histone H3 was absent from organelles and organelle nucleoids, consistent with the historical hypothesis. Thus, the organisation of organelle nucleoids, including putative nucleosome-like repetitive structures, should be constructed and maintained without participation of histone H3. © 2013 International Federation for Cell Biology.

  2. Environmental tobacco smoking, mutagen sensitivity, and head and neck squamous cell carcinoma.

    Science.gov (United States)

    Zhang, Z F; Morgenstern, H; Spitz, M R; Tashkin, D P; Yu, G P; Hsu, T C; Schantz, S P

    2000-10-01

    Although active tobacco smoking has been considered a major risk factor for head and neck cancer, few studies have evaluated environmental tobacco smoke (ETS) and its interaction with mutagen sensitivity on the risk of head and neck cancer. We investigated the relationship between ETS and head and neck cancer in a case-control study of 173 previously untreated cases with pathologically confirmed diagnoses of squamous cell carcinoma of the head and neck and 176 cancer-free controls at Memorial Sloan-Kettering Cancer Center between 1992 and 1994. A structured questionnaire was used to collect ETS exposure and other covariates including a history of active tobacco smoking and alcohol use. ETS measures include a history of ETS exposure at home and at workplace. The associations between passive smoking and head and neck cancer were analyzed by Mantel-Haenszel methods and logistic regression models. Additive and multiplicative models were used to evaluate effect modifications between ETS and mutagen sensitivity. The crude odds ratio (OR) for ETS exposure was 2.8 [95% confidence intervals (CI), 1.3-6.0]. Controlling for age, sex, race, education, alcohol consumption, pack-years of cigarette smoking, and marijuana use, the risk of squamous cell carcinoma of the head and neck was increased with ETS (adjusted OR, 2.4; 95% CI, 0.9-6.8). Dose-response relationships were observed for the degree of ETS exposure; the adjusted ORs were 2.1 (95% CI, 0.7-6.1) for those with moderate exposure and 3.6 (95% CI, 1.1-11.5) for individuals with heavy exposure (P for trend = 0.025), in comparison with those who never had ETS exposures. These associations and the dose-response relationships were still present when the analysis was restricted to nonactive smoking cases and controls (crude OR, 2.2; 95% CI, 0.6-8.4). Crude odds ratios were 1.8 for those with moderate ETS exposure and 4.3 for individuals with heavy ETS exposure among nonsmoking cases and controls (P for trend = 0.008). More

  3. Nitric Oxide- and Hydrogen Peroxide-Responsive Gene Regulation during Cell Death Induction in Tobacco1[W

    Science.gov (United States)

    Zago, Elisa; Morsa, Stijn; Dat, James F.; Alard, Philippe; Ferrarini, Alberto; Inzé, Dirk; Delledonne, Massimo; Van Breusegem, Frank

    2006-01-01

    Nitric oxide (NO) and hydrogen peroxide (H2O2) are regulatory molecules in various developmental processes and stress responses. Tobacco (Nicotiana tabacum) leaves exposed to moderate high light dramatically potentiated NO-mediated cell death in catalase-deficient (CAT1AS) but not in wild-type plants, providing genetic evidence for a partnership between NO and H2O2 during the induction of programmed cell death. With this experimental model system, the specific impact on gene expression was characterized by either NO or H2O2 alone or both molecules combined. By means of genome-wide cDNA-amplified fragment length polymorphism analysis, transcriptional changes were compared in high light-treated CAT1AS and wild-type leaves treated with or without the NO donor sodium nitroprusside. Differential gene expression was detected for 214 of the approximately 8,000 transcript fragments examined. For 108 fragments, sequence analysis revealed homology to genes with a role in signal transduction, defense response, hormone interplay, proteolysis, transport, and metabolism. Surprisingly, only 16 genes were specifically induced by the combined action of NO and H2O2, whereas the majority were regulated by either of them alone. At least seven transcription factors were mutually up-regulated, indicating significant overlap between NO and H2O2 signaling pathways. These results consolidate significant cross-talk between NO and H2O2, provide new insight into the early transcriptional response of plants to increased NO and H2O2 levels, and identify target genes of the combined action of NO and H2O2 during the induction of plant cell death. PMID:16603664

  4. Exogenous proline enhances the sensitivity of Tobacco BY-2 cells to arsenate.

    Science.gov (United States)

    Nahar, Mst Nur-E-Nazmun; Islam, Mohammad Muzahidul; Hoque, Md Anamul; Yonezawa, Anna; Prodhan, Md Yeasin; Nakamura, Toshiyuki; Nakamura, Yoshimasa; Munemasa, Shintaro; Murata, Yoshiyuki

    2017-09-01

    Arsenic causes physiological and structural disorders in plants. Proline is accumulated as a compatible solute in plants under various stress conditions and mitigates stresses. Here, we investigated the effects of exogenous proline on tobacco Bright Yellow-2 (BY-2) cultured cells under [Formula: see text] stress. Arsenate did not inhibit BY-2 cell growth at 40 and 50 μM but did it at 60 μM. Proline at 0.5 to 10 mM did not affect the cell growth but delayed it at 20 mM. At 40 μM [Formula: see text], neither 0.5 mM nor 1 mM proline affected the cell growth but 10 mM proline inhibited it. In the presence of [Formula: see text], 10 mM proline increased the number of Evans Blue-stained (dead) cells and decreased the number of total cells. Together, our results suggest that exogenous proline does not alleviate arsenate toxicity but enhances the sensitivity of BY-2 cells to arsenate.

  5. Tobacco Mosaic Virus with Peroxidase-Like Activity for Cancer Cell Detection through Colorimetric Assay.

    Science.gov (United States)

    Guo, Jiawang; Zhao, Xia; Hu, Jun; Lin, Yuan; Wang, Qian

    2018-01-22

    Cell-based ELISA (CELLISA) has been widely used in disease diagnosis due to its simplicity and low cost. Recently, peroxidase-like nanomaterials have emerged as promising systems for CELLISA applications. In this work, tobacco mosaic virus (TMV) was simultaneously tailored with peroxidase-like inorganic nanoparticles (platinum nanoparticles) and cancer cell target groups (folic acid, FA) to obtain TMV-FA-Pt nanoparticles for cancer cell detection. Induced by the uniformly distributed reactive groups and well-defined structure of the TMV particle, platinum nanoparticles could be grown in situ on the exterior surface of TMV with excellent monodispersity and uniform spatial distribution. Meanwhile, FA with a PEG 1000 linker was successfully conjugated to the coat proteins of TMV through the Cu(I)-catalyzed alkyne-azide cycloaddition reaction, an efficient "click" chemistry. Our study demonstrated that the resultant TMV-FA-Pt had specific affinity to cancer cells and was successfully used to detect cancer cells through CELLISA. Less than 1.0 × 10 4 cells/mL of cancer cells could be readily detected.

  6. Tobacco Products

    Science.gov (United States)

    ... Exposure is High in Multiunit Housing Smokeless Products Electronic Cigarettes Youth Tobacco Prevention Tobacco Products Tobacco Ingredient ... Tweet Share Compartir Find Fact Sheets on Products (Cigars, Bidis and Betel Quid with Tobacco (Gutka) and ...

  7. Melatonin Protects Cultured Tobacco Cells against Lead-Induced Cell Death via Inhibition of Cytochrome c Translocation

    Science.gov (United States)

    Kobylińska, Agnieszka; Reiter, Russel J.; Posmyk, Malgorzata M.

    2017-01-01

    Melatonin was discovered in plants more than two decades ago and, especially in the last decade, it has captured the interests of plant biologists. Beyond its possible participation in photoperiod processes and its role as a direct free radical scavenger as well as an indirect antioxidant, melatonin is also involved in plant defense strategies/reactions. However, the mechanisms that this indoleamine activates to improve plant stress tolerance still require identification and clarification. In the present report, the ability of exogenous melatonin to protect Nicotiana tabacum L. line Bright Yellow 2 (BY-2) suspension cells against the toxic exposure to lead was examined. Studies related to cell proliferation and viability, DNA fragmentation, possible translocation of cytochrome c from mitochondria to cytosol, cell morphology after fluorescence staining and also the in situ accumulation of superoxide radicals measured via the nitro blue tetrazolium reducing test, were conducted. This work establishes a novel finding by correcting the inhibition of release of mitochondrial ctytocrome c in to the cytoplasm with the high accumulation of superoxide radicals. The results show that pretreatment with 200 nm of melatonin protected tobacco cells from DNA damage caused by lead. Melatonin, as an efficacious antioxidant, limited superoxide radical accumulation as well as cytochrome c release thereby, it likely prevents the activation of the cascade of processes leading to cell death. Fluorescence staining with acridine orange and ethidium bromide documented that lead-stressed cells additionally treated with melatonin displayed intact nuclei. The results revealed that melatonin at proper dosage could significantly increase BY-2 cell proliferation and protected them against death. It was proved that melatonin could function as an effective priming agent to promote survival of tobacco cells under harmful lead-induced stress conditions. PMID:28959267

  8. Melatonin Protects Cultured Tobacco Cells against Lead-Induced Cell Death via Inhibition of Cytochrome c Translocation

    Directory of Open Access Journals (Sweden)

    Agnieszka Kobylińska

    2017-09-01

    Full Text Available Melatonin was discovered in plants more than two decades ago and, especially in the last decade, it has captured the interests of plant biologists. Beyond its possible participation in photoperiod processes and its role as a direct free radical scavenger as well as an indirect antioxidant, melatonin is also involved in plant defense strategies/reactions. However, the mechanisms that this indoleamine activates to improve plant stress tolerance still require identification and clarification. In the present report, the ability of exogenous melatonin to protect Nicotiana tabacum L. line Bright Yellow 2 (BY-2 suspension cells against the toxic exposure to lead was examined. Studies related to cell proliferation and viability, DNA fragmentation, possible translocation of cytochrome c from mitochondria to cytosol, cell morphology after fluorescence staining and also the in situ accumulation of superoxide radicals measured via the nitro blue tetrazolium reducing test, were conducted. This work establishes a novel finding by correcting the inhibition of release of mitochondrial ctytocrome c in to the cytoplasm with the high accumulation of superoxide radicals. The results show that pretreatment with 200 nm of melatonin protected tobacco cells from DNA damage caused by lead. Melatonin, as an efficacious antioxidant, limited superoxide radical accumulation as well as cytochrome c release thereby, it likely prevents the activation of the cascade of processes leading to cell death. Fluorescence staining with acridine orange and ethidium bromide documented that lead-stressed cells additionally treated with melatonin displayed intact nuclei. The results revealed that melatonin at proper dosage could significantly increase BY-2 cell proliferation and protected them against death. It was proved that melatonin could function as an effective priming agent to promote survival of tobacco cells under harmful lead-induced stress conditions.

  9. Coniferyl alcohol hinders the growth of tobacco BY-2 cells and Nicotiana benthamiana seedlings.

    Science.gov (United States)

    Väisänen, Enni E; Smeds, Annika I; Fagerstedt, Kurt V; Teeri, Teemu H; Willför, Stefan M; Kärkönen, Anna

    2015-09-01

    Externally added coniferyl alcohol at high concentrations reduces the growth of Nicotiana cells and seedlings. Coniferyl alcohol is metabolized by BY-2 cells to several compounds. Coniferyl alcohol (CA) is a common monolignol and a building block of lignin. The toxicity of monolignol alcohols has been stated in the literature, but there are only few studies suggesting that this is true. We investigated the physiological effects of CA on living plant cells in more detail. Tobacco (Nicotiana tabacum) Bright yellow-2 cells (BY-2) and Nicotiana benthamiana seedlings both showed concentration-dependent growth retardation in response to 0.5-5 mM CA treatment. In some cases, CA addition caused cell death in BY-2 cultures, but this response was dependent on the growth stage of the cells. Based on LC-MS/MS analysis, BY-2 cells did not accumulate the externally supplemented CA, but metabolized it to ferulic acid, ferulic acid glycoside, coniferin, and to some other phenolic compounds. In addition to growth inhibition, CA caused the formation of a lignin-like compound detected by phloroglucinol staining in N. benthamiana roots and occasionally in BY-2 cells. To prevent this, we added potassium iodide (KI, at 5 mM) to overcome the peroxidase-mediated CA polymerization to lignin. KI had, however, toxic effects on its own: in N. benthamiana seedlings, it caused reduction in growth; in BY-2 cells, reduction in growth and cell viability. Surprisingly, CA restored the growth of KI-treated BY-2 cells and N. benthamiana seedlings. Our results suggest that CA at high concentrations is toxic to plant cells.

  10. Zeatin is indispensable for the G2-M transition in tobacco BY-2 cells.

    Science.gov (United States)

    Laureys, F; Dewitte, W; Witters, E; Van Montagu, M; Inzé, D; Van Onckelen, H

    1998-04-10

    The importance of N6-isoprenoid cytokinins in the G2-M transition of Nicotiana tabacum BY-2 cells was investigated. Both cytokinin biosynthesis and entry in mitosis were partially blocked by application at early or late G2 of lovastatin (10 microM), an inhibitor of mevalonic acid synthesis. LC-MS/MS quantification of endogenous cytokinins proved that lovastatin affects cytokinin biosynthesis by inhibiting HMG-CoA reductase. Out of eight different aminopurines and a synthetic auxin tested for their ability to override lovastatin inhibition of mitosis, only zeatin was active. Our data point to a key role for a well-defined cytokinin (here, zeatin) in the G2-M transition of tobacco BY-2 cells.

  11. The radiosensitizing effect of doranidazole on human colorectal cancer cells exposed to high doses of irradiation

    International Nuclear Information System (INIS)

    Zhang, Li; Gong, Aimin; Ji, Jun; Wu, Yuanyuan; Zhu, Xiaoyu; Lv, Suqing; Lv, Hongzhu; Sun, Xizhuo

    2007-01-01

    This paper investigates the effects of a new radiosensitizer, doranidazole, and enhancing irradiation on colorectal cancer cells. The radiosensitizing effect of doranidazole was determined using colony formation and propidium iodide (PI) assays to measure cell growth inhibition and the cell killing effect of human colorectal cancer cell lines exposed to high doses of γ-ray irradiation under hypoxic conditions in vitro. Fluorescence staining and cell migration assays were also used to assess the radiosensitizing effect. Cell proliferation evaluated by clonogenic survival curves was significantly inhibited by 5 mmol/L doranidazole, particularly at doses ranging from 10 to 30 Gy of irradiation. The radiosensitizing effect of doranidazole on colorectal cancer cells occurs in a time- and dose-dependent manner. Doranidazole also inhibited the mobility of cell invasion and migration. Doranidazole can enhance the killing effect and the cell growth inhibition of colorectal cancer after high-dose irradiation in a time and dose-dependent manner

  12. Changes in cell wall properties coincide with overexpression of extensin fusion proteins in suspension cultured tobacco cells.

    Science.gov (United States)

    Tan, Li; Pu, Yunqiao; Pattathil, Sivakumar; Avci, Utku; Qian, Jin; Arter, Allison; Chen, Liwei; Hahn, Michael G; Ragauskas, Arthur J; Kieliszewski, Marcia J

    2014-01-01

    Extensins are one subfamily of the cell wall hydroxyproline-rich glycoproteins, containing characteristic SerHyp4 glycosylation motifs and intermolecular cross-linking motifs such as the TyrXaaTyr sequence. Extensins are believed to form a cross-linked network in the plant cell wall through the tyrosine-derivatives isodityrosine, pulcherosine, and di-isodityrosine. Overexpression of three synthetic genes encoding different elastin-arabinogalactan protein-extensin hybrids in tobacco suspension cultured cells yielded novel cross-linking glycoproteins that shared features of the extensins, arabinogalactan proteins and elastin. The cell wall properties of the three transgenic cell lines were all changed, but in different ways. One transgenic cell line showed decreased cellulose crystallinity and increased wall xyloglucan content; the second transgenic cell line contained dramatically increased hydration capacity and notably increased cell wall biomass, increased di-isodityrosine, and increased protein content; the third transgenic cell line displayed wall phenotypes similar to wild type cells, except changed xyloglucan epitope extractability. These data indicate that overexpression of modified extensins may be a route to engineer plants for bioenergy and biomaterial production.

  13. Micronucleus frequency in exfoliated buccal cells from hairdresser who expose to hair products

    Directory of Open Access Journals (Sweden)

    Koh Hui Yee

    2015-06-01

    Full Text Available Background: Hairdresser is one of the fastest growing occupations in today’s society. Hairdresser help styling, cutting, colouring, perming, curling, straightening hair and various treatment to customer. Somehow, hairdresser are constantly exposed to chemical substances such as aromatic amines, hydrogen peroxide, thioglycolic acid, formaldehyde in hair products which can cause damage to human’s genome. Micronucleus is one of the effective biomarker for processes associated with the induction of DNA damage. Purpose: The aim of this study was to determine the micronucleus frequencies in buccal mucosa epithelial cells of hairdresser who were exposed to chemical of hair products. Method: This study was conducted on twenty female subjects, who were divided into 2 groups: exposed and non-exposed (control group. All subjects recruited were working in the same beauty salon. Buccal cells were obtained from each individual by using cytobrush. The cells were stained with modified Feulgen-Ronssenback method and counting of micronucleus per 1000 cell was done under light microscope. The data were analyzed using independent t-test and one-way Anova (p<0.05. Result: The result showed a significant difference in micronucleus frequency between 2 groups. There were a significantly increase of micronucleus frequency in hairdressers and increase of  micronucleus frequency with the longer duration of exposure. Conclusion: It concluded that the chemical substances of hair products had affected the micronucleus frequency ofthe epithelial cells in buccal mucosa of hairdressers.

  14. Cardiac development in zebrafish and human embryonic stem cells is inhibited by exposure to tobacco cigarettes and e-cigarettes.

    Directory of Open Access Journals (Sweden)

    Nathan J Palpant

    Full Text Available Maternal smoking is a risk factor for low birth weight and other adverse developmental outcomes.We sought to determine the impact of standard tobacco cigarettes and e-cigarettes on heart development in vitro and in vivo.Zebrafish (Danio rerio were used to assess developmental effects in vivo and cardiac differentiation of human embryonic stem cells (hESCs was used as a model for in vitro cardiac development.In zebrafish, exposure to both types of cigarettes results in broad, dose-dependent developmental defects coupled with severe heart malformation, pericardial edema and reduced heart function. Tobacco cigarettes are more toxic than e-cigarettes at comparable nicotine concentrations. During cardiac differentiation of hESCs, tobacco smoke exposure results in a delayed transition through mesoderm. Both types of cigarettes decrease expression of cardiac transcription factors in cardiac progenitor cells, suggesting a persistent delay in differentiation. In definitive human cardiomyocytes, both e-cigarette- and tobacco cigarette-treated samples showed reduced expression of sarcomeric genes such as MLC2v and MYL6. Furthermore, tobacco cigarette-treated samples had delayed onset of beating and showed low levels and aberrant localization of N-cadherin, reduced myofilament content with significantly reduced sarcomere length, and increased expression of the immature cardiac marker smooth muscle alpha-actin.These data indicate a negative effect of both tobacco cigarettes and e-cigarettes on heart development in vitro and in vivo. Tobacco cigarettes are more toxic than E-cigarettes and exhibit a broader spectrum of cardiac developmental defects.

  15. Combustible and non-combustible tobacco product preparations differentially regulate human peripheral blood mononuclear cell functions.

    Science.gov (United States)

    Arimilli, Subhashini; Damratoski, Brad E; Prasad, G L

    2013-09-01

    Natural killer (NK) cells and T cells play essential roles in innate and adaptive immune responses in protecting against microbial infections and in tumor surveillance. Although evidence suggests that smoking causes immunosuppression, there is limited information whether the use of smokeless tobacco (ST) products affects immune responses. In this study, we assessed the effects of two preparations of cigarette smoke, ST extract and nicotine on T cell and NK cell responses using Toll-like receptor-ligand stimulated human peripheral blood mononuclear cells (PBMCs). The tobacco product preparations (TPPs) tested included whole smoke conditioned media (WS-CM), total particulate matter (TPM) and a ST product preparation in complete artificial saliva (ST/CAS). The PBMCs were stimulated with polyinosinic:polycytidylic acid (poly I:C) and lipopolysaccharide (LPS). A marked reduction of the expression of intracellular IFN-γ and TNF-α was evident in NK cells and T cells treated with WS-CM and TPM. Consistently, attenuation of ligand-induced secretion of cytokines (IL-1β, IL-10, IL-12 and TNF-α) from PBMCs treated with WS-CM and TPM were observed. While the treatment with TPPs did not alter the expression of the maturation marker CD69, WS-CM and TPM inhibited the cytolytic activity of human PBMCs. Suppression of perforin by WS-CM was also detected. Although interference from the vehicle confounded the interpretation of effects of ST/CAS, some effects were evident only at high concentrations. Nicotine treatment minimally impacted expression of cytokines and cytolytic activity. Data presented herein suggests that the function of NK cells and T cells is influenced by exposure to TPPs (based on equi-nicotine units) in the following order: WS-CM>TPM>ST/CAS. These findings are consistent with the hypothesis put forward by others that chronic smoking leads to immunosuppression, an effect that may contribute to increased microbial infections and cancer incidence among smokers

  16. Arabinogalactan Proteins Are Involved in Salt-Adaptation and Vesicle Trafficking in Tobacco by-2 Cell Cultures.

    Science.gov (United States)

    Olmos, Enrique; García De La Garma, Jesús; Gomez-Jimenez, Maria C; Fernandez-Garcia, Nieves

    2017-01-01

    Arabinogalactan proteins (AGPs) are a highly diverse family of glycoproteins that are commonly found in most plant species. However, little is known about the physiological and molecular mechanisms of their function. AGPs are involved in different biological processes such as cell differentiation, cell expansion, tissue development and somatic embryogenesis. AGPs are also involved in abiotic stress response such as salinity modulating cell wall expansion. In this study, we describe how salt-adaptation in tobacco BY-2 cell cultures induces important changes in arabinogalactan proteins distribution and contents. Using the immuno-dot blot technique with different anti-AGP antibodies (JIM13, JIM15, and others), we observed that AGPs were highly accumulated in the culture medium of salt-adapted tobacco cells, probably due to the action of phospholipases. We located these AGP epitopes using immunogold labeling in the cytoplasm associated to the endoplasmic reticulum, the golgi apparatus, and vesicles, plasma membrane and tonoplast. Our results show that salt-adaptation induced a significant reduction of the cytoplasm, plasma membrane and tonoplast content of these epitopes. Yariv reagent was added to the control and salt-adapted tobacco cell cultures, leading to cell death induction in control cells but not in salt-adapted cells. Ultrastructural and immunogold labeling revealed that cell death induced by Yariv reagent in control cells was due to the interaction of Yariv reagent with the AGPs linked to the plasma membranes. Finally, we propose a new function of AGPs as a possible sodium carrier through the mechanism of vesicle trafficking from the apoplast to the vacuoles in salt-adapted tobacco BY-2 cells. This mechanism may contribute to sodium homeostasis during salt-adaptation to high saline concentrations.

  17. Detection of genomic instability in normal human bronchial epithelial cells exposed to 238Pu

    International Nuclear Information System (INIS)

    Kennedy, C.H.; Fukushima, N.H.; Neft, R.E.; Lechner, J.F.

    1994-01-01

    Alpha particle-emitting radon daughters constitute a risk for development of lung cancer in humans. The development of this disease involves multiple genetic alterations. These changes and the time course they follow are not yet defined despite numerous in vitro endeavors to transform human lung cells with various physical or chemical agents. However, genomic instability, characterized both by structural and numerical chromosomal aberrations and by elevated rates of point mutations, is a common feature of tumor cells. Further, both types of genomic instability have been reported in the noncancerous progeny of normal murine hemopoietic cells exposed in vitro to α-particles. The purpose of this investigation was to determine if genomic instability is also a prominent feature of normal human bronchial epithelial cells exposed to α-particle irradiation from the decay of inhaled radon daughters

  18. Chromosome aberrations of bone marrow cells in heavily exposed atomic bomb survivors

    International Nuclear Information System (INIS)

    Tanaka, Kimio; Kamada, Nanao; Kuramoto, Atsushi; Ohkita, Takeshi

    1986-01-01

    Seven hundred and ten bone marrow cells from 13 A-bomb survivors, who were heavily exposed to atomic radiation, were examined using chromosome banding method. An average frequency of chromosome aberrations was 17 %. The most common structural abnormality was translocation (47 %), followed by complex aberrations involving three or more chromosomes (32 %). These abnormalities were frequently seen in A-bomb survivors exposed to estimated doses of 3.5 - 4.0 Gy. Eighty two percent of the structural aberrations were stable. Diploid cells were seen in 0.4 % and tetraploid cells were seen in 0.7 %. The frequency of breakpoint sites was high in chromosomes 1 and 17; while it was low in chromosomes 3, 6, 9, and 11. Abnormal clones were seen in one of the 13 survivors. Chromosome aberrations common to the bone marrow cells and peripheral lymphocytes were not seen in the same individual. (Namekawa, K.)

  19. Gene expression profile in bone marrow and hematopoietic stem cells in mice exposed to inhaled benzene

    International Nuclear Information System (INIS)

    Faiola, Brenda; Fuller, Elizabeth S.; Wong, Victoria A.; Recio, Leslie

    2004-01-01

    Acute myeloid leukemia and chronic lymphocytic leukemia are associated with benzene exposure. In mice, benzene induces chromosomal breaks as a primary mode of genotoxicity in the bone marrow (BM). Benzene-induced DNA lesions can lead to changes in hematopoietic stem cells (HSC) that give rise to leukemic clones. To gain insight into the mechanism of benzene-induced leukemia, we investigated the DNA damage repair and response pathways in total bone marrow and bone marrow fractions enriched for HSC from male 129/SvJ mice exposed to benzene by inhalation. Mice exposed to 100 ppm benzene for 6 h per day, 5 days per week for 2 week showed significant hematotoxicity and genotoxicity compared to air-exposed control mice. Benzene exposure did not alter the level of apoptosis in BM or the percentage of HSC in BM. RNA isolated from total BM cells and the enriched HSC fractions from benzene-exposed and air-exposed mice was used for microarray analysis and quantitative real-time RT-PCR. Interestingly, mRNA levels of DNA repair genes representing distinct repair pathways were largely unaffected by benzene exposure, whereas altered mRNA expression of various apoptosis, cell cycle, and growth control genes was observed in samples from benzene-exposed mice. Differences in gene expression profiles were observed between total BM and HSC. Notably, p21 mRNA was highly induced in BM but was not altered in HSC following benzene exposure. The gene expression pattern suggests that HSC isolated immediately following a 2 weeks exposure to 100 ppm benzene were not actively proliferating. Understanding the toxicogenomic profile of the specific target cell population involved in the development of benzene-associated diseases may lead to a better understanding of the mechanism of benzene-induced leukemia and may identify important interindividual and tissue susceptibility factors

  20. Differential cisplatin responses in human carcinoma cell lines pre-exposed to fractionated X-irradiation

    International Nuclear Information System (INIS)

    Dempke, W.C.M.; Hosking, L.K.; Shellard, S.A.; Hill, B.T.

    1991-01-01

    These results suggest that cells exposed to X-irradiation may respond differently to subsequent cisplatin (CDDP) treatment. Initial studies of possible mechanisms responsible for these differential sensitivities indicate that they may differ according to whether resistance or hypersensitivity is expressed. (author)

  1. Immune cells in Chernobyl radiation workers exposed to low-dose irradiation

    International Nuclear Information System (INIS)

    Bazyka, D.; Chumak, A.; Byelyaeva, N.; Gulaya, N.; Margytich, V.; Thevenon, C.; Guichardant, M.; Lagarde, M.

    2002-01-01

    the aim of this work was to study immune response parameters in Chernobyl emergency and recovery operation radiation workers and nuclear industry workers exposed under professional limits. The monohydroxylated fatty acid content in peripheral blood mononuclear cell of radiation workers compared to unexposed control at the 12-th year after Chernobyl NPP accident was studied too

  2. Determining adaptive and adverse oxidative stress responses in human bronical epithelial cells exposed to zinc

    Science.gov (United States)

    Determining adaptive and adverse oxidative stress responses in human bronchial epithelial cells exposed to zincJenna M. Currier1,2, Wan-Yun Cheng1, Rory Conolly1, Brian N. Chorley1Zinc is a ubiquitous contaminant of ambient air that presents an oxidant challenge to the human lung...

  3. Ultrastructure of cells of Ulmus americana cultured in vitro and exposed to the culture filtrate of Ceratocystis ulmi

    Science.gov (United States)

    Paula M. Pijut; R. Daniel Lineberger; Subhash C. Domir; Jann M. Ichida; Charles R. Krause

    1990-01-01

    Calli of American elm susceptible and resistant to Dutch elm disease were exposed to a culture filtrate of a pathogenic isolate of Ceratocystis ulmi. Cells from untreated tissue exhibited typical internal composition associated with healthy, actively growing cells. All cells exposed to culture filtrate showed appreciable ultrastructural changes....

  4. Involvement of DNA methylation in the control of cell growth during heat stress in tobacco BY-2 cells.

    Science.gov (United States)

    Centomani, Isabella; Sgobba, Alessandra; D'Addabbo, Pietro; Dipierro, Nunzio; Paradiso, Annalisa; De Gara, Laura; Dipierro, Silvio; Viggiano, Luigi; de Pinto, Maria Concetta

    2015-11-01

    The alteration of growth patterns, through the adjustment of cell division and expansion, is a characteristic response of plants to environmental stress. In order to study this response in more depth, the effect of heat stress on growth was investigated in tobacco BY-2 cells. The results indicate that heat stress inhibited cell division, by slowing cell cycle progression. Cells were stopped in the pre-mitotic phases, as shown by the increased expression of CycD3-1 and by the decrease in the NtCycA13, NtCyc29 and CDKB1-1 transcripts. The decrease in cell length and the reduced expression of Nt-EXPA5 indicated that cell expansion was also inhibited. Since DNA methylation plays a key role in controlling gene expression, the possibility that the altered expression of genes involved in the control of cell growth, observed during heat stress, could be due to changes in the methylation state of their promoters was investigated. The results show that the altered expression of CycD3-1 and Nt-EXPA5 was consistent with changes in the methylation state of the upstream region of these genes. These results suggest that DNA methylation, controlling the expression of genes involved in plant development, contributes to growth alteration occurring in response to environmental changes.

  5. Characterization of xylan in the early stages of secondary cell wall formation in tobacco bright yellow-2 cells.

    Science.gov (United States)

    Ishii, Tadashi; Matsuoka, Keita; Ono, Hiroshi; Ohnishi-Kameyama, Mayumi; Yaoi, Katsuro; Nakano, Yoshimi; Ohtani, Misato; Demura, Taku; Iwai, Hiroaki; Satoh, Shinobu

    2017-11-15

    The major polysaccharides present in the primary and secondary walls surrounding plant cells have been well characterized. However, our knowledge of the early stages of secondary wall formation is limited. To address this, cell walls were isolated from differentiating xylem vessel elements of tobacco bright yellow-2 (BY-2) cells induced by VASCULAR-RELATED NAC-DOMAIN7 (VND7). The walls of induced VND7-VP16-GR BY-2 cells consisted of cellulose, pectic polysaccharides, hemicelluloses, and lignin, and contained more xylan and cellulose compared with non-transformed BY-2 and uninduced VND7-VP16-GR BY-2 cells. A reducing end sequence of xylan containing rhamnose and galaturonic acid- residues is present in the walls of induced, uninduced, and non-transformed BY-2 cells. Glucuronic acid residues in xylan from walls of induced cells are O-methylated, while those of xylan in non-transformed BY-2 and uninduced cells are not. Our results show that xylan changes in chemical structure and amounts during the early stages of xylem differentiation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. TMBP200, a XMAP215 homologue of tobacco BY-2 cells, has an essential role in plant mitosis.

    Science.gov (United States)

    Yasuhara, Hiroki; Oe, Yuki

    2011-07-01

    TMBP200 from tobacco BY-2 cells is a member of the highly conserved family of microtubule-associated proteins that includes Xenopus XMAP215, human TOGp, and Arabidopsis MOR1/GEM1. XMAP215 homologues have an essential role in spindle assembly and function in animals and yeast, but their role in plant mitosis is not fully clarified. Here, we show by immunoblot analysis that TMBP200 levels in synchronously cultured BY-2 cells increased when the cells entered mitosis, thus indicating that TMBP200 plays an important role in mitosis in tobacco. To investigate the role of TMBP200 in mitosis, we employed inducible RNA interference to silence TMBP200 expression in BY-2 cells. The resulting depletion of TMBP200 caused severe defects in bipolar spindle formation and resulted in the appearance of multinucleated cells with variable-sized nuclei. This finding indicates that TMBP200 has an essential role in bipolar spindle formation and function.

  7. Metabolomic fingerprinting of primed tobacco cells provide the first evidence for the biological origin of cis-chlorogenic acid

    CSIR Research Space (South Africa)

    Mhlongo, MI

    2015-01-01

    Full Text Available . Very little, however, is known about the biological significance and origin of cis-isomers. Here we show for the first time the accumulation of cis-5-caffeoylquinic acid in cultured tobacco cells treated with different inducers of plant defence...

  8. Protein C Inhibitor (PCI) Binds to Phosphatidylserine Exposing Cells with Implications in the Phagocytosis of Apoptotic Cells and Activated Platelets

    Science.gov (United States)

    Rieger, Daniela; Assinger, Alice; Einfinger, Katrin; Sokolikova, Barbora; Geiger, Margarethe

    2014-01-01

    Protein C Inhibitor (PCI) is a secreted serine protease inhibitor, belonging to the family of serpins. In addition to activated protein C PCI inactivates several other proteases of the coagulation and fibrinolytic systems, suggesting a regulatory role in hemostasis. Glycosaminoglycans and certain negatively charged phospholipids, like phosphatidylserine, bind to PCI and modulate its activity. Phosphatidylerine (PS) is exposed on the surface of apoptotic cells and known as a phagocytosis marker. We hypothesized that PCI might bind to PS exposed on apoptotic cells and thereby influence their removal by phagocytosis. Using Jurkat T-lymphocytes and U937 myeloid cells, we show here that PCI binds to apoptotic cells to a similar extent at the same sites as Annexin V, but in a different manner as compared to live cells (defined spots on ∼10–30% of cells). PCI dose dependently decreased phagocytosis of apoptotic Jurkat cells by U937 macrophages. Moreover, the phagocytosis of PS exposing, activated platelets by human blood derived monocytes declined in the presence of PCI. In U937 cells the expression of PCI as well as the surface binding of PCI increased with time of phorbol ester treatment/macrophage differentiation. The results of this study suggest a role of PCI not only for the function and/or maturation of macrophages, but also as a negative regulator of apoptotic cell and activated platelets removal. PMID:25000564

  9. Zinc-Dependent Protection of Tobacco and Rice Cells From Aluminum-Induced Superoxide-Mediated Cytotoxicity

    Science.gov (United States)

    Lin, Cun; Hara, Ayaka; Comparini, Diego; Bouteau, François; Kawano, Tomonori

    2015-01-01

    Al3+ toxicity in growing plants is considered as one of the major factors limiting the production of crops on acidic soils worldwide. In the last 15 years, it has been proposed that Al3+ toxicity are mediated with distortion of the cellular signaling mechanisms such as calcium signaling pathways, and production of cytotoxic reactive oxygen species (ROS) causing oxidative damages. On the other hand, zinc is normally present in plants at high concentrations and its deficiency is one of the most widespread micronutrient deficiencies in plants. Earlier studies suggested that lack of zinc often results in ROS-mediated oxidative damage to plant cells. Previously, inhibitory action of Zn2+ against lanthanide-induced superoxide generation in tobacco cells have been reported, suggesting that Zn2+ interferes with the cation-induced ROS production via stimulation of NADPH oxidase. In the present study, the effect of Zn2+ on Al3+-induced superoxide generation in the cell suspension cultures of tobacco (Nicotiana tabacum L., cell-line, BY-2) and rice (Oryza sativa L., cv. Nipponbare), was examined. The Zn2+-dependent inhibition of the Al3+-induced oxidative burst was observed in both model cells selected from the monocots and dicots (rice and tobacco), suggesting that this phenomenon (Al3+/Zn2+ interaction) can be preserved in higher plants. Subsequently induced cell death in tobacco cells was analyzed by lethal cell staining with Evans blue. Obtained results indicated that presence of Zn2+ at physiological concentrations can protect the cells by preventing the Al3+-induced superoxide generation and cell death. Furthermore, the regulation of the Ca2+ signaling, i.e., change in the cytosolic Ca2+ ion concentration, and the cross-talks among the elements which participate in the pathway were further explored. PMID:26648960

  10. Heterologous expression of plant cell wall glycosyltransferases in Pichia, pea and tobacco

    DEFF Research Database (Denmark)

    Petersen, Bent Larsen; Damager, Iben; Faber, Kirsten

    Cell). In the present study, Flag-tagged (MDYKDDDD) RGXT2 was expressed in Pichia pastoris as secreted soluble protein, in pea (using the Pea early browning virus as expression vector) as soluble intra-cellular protein and in tobacco as full length membrane bound protein. The amount of expressed...... to participate in plant CW biosynthesis, has been achieved in only a few cases. We have previously reported the characterisation of two highly homologous plant-specific membrane-bound GTs, which when expressed as secreted tagged soluble proteins in the baculo virus system, catalysed the transfer of xylose from...... protein was estimated using anti Flag Ab and corresponding activity monitored. Pros and cons of using the various expression systems are discussed....

  11. Assessment of cultivation factors that affect biomass and geraniol production in transgenic tobacco cell suspension cultures.

    Directory of Open Access Journals (Sweden)

    Nikolay Vasilev

    Full Text Available A large-scale statistical experimental design was used to determine essential cultivation parameters that affect biomass accumulation and geraniol production in transgenic tobacco (Nicotiana tabacum cv. Samsun NN cell suspension cultures. The carbohydrate source played a major role in determining the geraniol yield and factors such as filling volume, inoculum size and light were less important. Sucrose, filling volume and inoculum size had a positive effect on geraniol yield by boosting growth of plant cell cultures whereas illumination of the cultures stimulated the geraniol biosynthesis. We also found that the carbohydrates sucrose and mannitol showed polarizing effects on biomass and geraniol accumulation. Factors such as shaking frequency, the presence of conditioned medium and solubilizers had minor influence on both plant cell growth and geraniol content. When cells were cultivated under the screened conditions for all the investigated factors, the cultures produced ∼ 5.2 mg/l geraniol after 12 days of cultivation in shaking flasks which is comparable to the yield obtained in microbial expression systems. Our data suggest that industrial experimental designs based on orthogonal arrays are suitable for the selection of initial cultivation parameters prior to the essential medium optimization steps. Such designs are particularly beneficial in the early optimization steps when many factors must be screened, increasing the statistical power of the experiments without increasing the demand on time and resources.

  12. Exposure to tobacco-derived materials induces overproduction of secreted proteinases in mast cells

    International Nuclear Information System (INIS)

    Small-Howard, Andrea; Turner, Helen

    2005-01-01

    Mast cells reside at interfaces with the environment, including the mucosa of the respiratory and gastrointestinal tracts. This localization exposes mast cells to inhaled, or ingested, environmental challenges. In the airways of smokers, resident immune cells will be in contact with the condensed components of cigarette smoke. Mast cells are of particular interest due to their ability to promote airway remodeling and mucus hypersecretion. Clinical data show increased levels of mast cell-secreted tryptase and increased numbers of degranulated mast cells in the lavage and bronchial tissue of smokers. Since mast cell-secreted proteinases (MCPTs), including tryptases, contribute to pathological airway remodeling, we investigated the relationship between mast cell proteinases and smoke exposure. We exposed a mast cell line to cigarette smoke condensate (CSC). We show that CSC exposure increases MCPT levels in mast cells using an assay for tryptase-type MCPT activity. We hypothesized that this increase in MCPT activity reflects a CSC-induced increase in the cytosolic pool of proteinase molecules, via stimulation of MCPT transcription. Transcript array data suggested that mRNA changes in response to CSC were limited in number and peaked after 3 h of CSC exposure. However, we noted marked transcriptional regulation of several MCPT genes. CSC-induced changes in the mRNA levels for MCPTs were confirmed using quantitative RT-PCR. Taken together, our data suggest that chronic exposure to cigarette smoke up-regulates MCPT levels in mast cells at both the protein and the mRNA level. We suggest that the pathological airway remodeling that has been described in clinical studies of smoke inhalation may be attributable to MCPT overproduction in vivo

  13. Cellular and exosome mediated molecular defense mechanism in bovine granulosa cells exposed to oxidative stress.

    Directory of Open Access Journals (Sweden)

    Mohammed Saeed-Zidane

    Full Text Available Various environmental insults including diseases, heat and oxidative stress could lead to abnormal growth, functions and apoptosis in granulosa cells during ovarian follicle growth and oocyte maturation. Despite the fact that cells exposed to oxidative stress are responding transcriptionally, the potential release of transcripts associated with oxidative stress response into extracellular space through exosomes is not yet determined. Therefore, here we aimed to investigate the effect of oxidative stress in bovine granulosa cells in vitro on the cellular and exosome mediated defense mechanisms. Bovine granulosa cells were aspirated from ovarian follicles and cultured in DMEM/F-12 Ham culture medium supplemented with 10% exosome-depleted fetal bovine serum. In the first experiment sub-confluent cells were treated with 5 μM H2O2 for 40 min to induce oxidative stress. Thereafter, cells were subjected to ROS and mitochondrial staining, cell proliferation and cell cycle assays. Furthermore, gene and protein expression analysis were performed in H2O2-challenged versus control group 24 hr post-treatment using qRT-PCR and immune blotting or immunocytochemistry assay, respectively. Moreover, exosomes were isolated from spent media using ultracentrifugation procedure, and subsequently used for RNA isolation and qRT-PCR. In the second experiment, exosomes released by granulosa cells under oxidative stress (StressExo or those released by granulosa cells without oxidative stress (NormalExo were co-incubated with bovine granulosa cells in vitro to proof the potential horizontal transfer of defense molecules from exosomes to granulosa cells and investigate any phenotype changes. Exposure of bovine granulosa cells to H2O2 induced the accumulation of ROS, reduced mitochondrial activity, increased expression of Nrf2 and its downstream antioxidant genes (both mRNA and protein, altered the cell cycle transitions and induced cellular apoptosis. Granulosa cells

  14. Cytotoxic mechanism related to dihydrolipoamide dehydrogenase in Leydig cells exposed to heavy metals

    International Nuclear Information System (INIS)

    Ji, Xunmin; Li, Zhiliang; Chen, Hongxia; Li, Junqi; Tian, Huajian; Li, Zengli; Gao, Xuejuan; Xiang, Qi; Su, Zhijian; Huang, Yadong; Zhang, Qihao

    2015-01-01

    Heavy metals are common environmental toxicants with adverse effects on steroid biosynthesis. The importance of mitochondria has been recognized in cytotoxic mechanism of heavy metals on Leydig cells these years. But it is still poorly known. Our previous study reported that dihydrolipoamide dehydrogenase (DLD) located on the mitochondria was significantly decreased in Leydig cells exposed to cadmium, which suggested that DLD might be involved in the cytotoxic effects. Therefore, the altered expression of DLD was validated in rats and R2C cells exposed to cadmium, manganese and lead, and the role of DLD in the steroid synthesis pathway cAMP/PKA-ERK1/2 was investigated in this study. With a low expression of DLD, heavy metals dramatically reduced the levels of steroid hormone by inhibiting the activation of cAMP/PKA, PKC signaling pathway and the steroidogenic enzymes StAR, CYP11A1 and 3β-HSD. After knockdown of DLD in R2C cells, progesterone synthesis was reduced by 40%, and the intracellular concentration of cAMP, protein expression of StAR, 3β-HSD, PKA, and the phosphorylation of ERK1/2 were also decreased. These results highlight that DLD is down-regulation and related to steroid biosynthesis in Leyig cells exposed to heavy metals; cAMP/PKA act as downstream effector molecules of DLD, which activate phosphorylation of ERK1/2 to initiate the steroidogenesis

  15. 'Rogue' cells observed in children exposed to radiation from the Chernobyl accident

    International Nuclear Information System (INIS)

    Sevan'kaev, A.V.; Tsyb, A.F.; Zhloba, A.A.; Moiseenko, V.V.; Skrjabin, A.M.; Climov, V.M.

    1993-01-01

    Eight 'rogue' lymphocyte metaphases containing a large number of aberrant chromosomes were noted during a survey of chromosomal damage in 328 Belarussian children. The study population comprised children of families living in territory contaminated by radiation from the Chernobyl accident. The majority of the sample had been evacuated within 1 week from very heavily polluted territory to areas that had received much less fallout. Two hundred cells were scored per subject and one rogue cell was found in a child exposed in utero; one in a child conceived after the accident and six in the postnatally exposed group. The possibility that the damage was due to exposure to radio-iodine concentrated in the thyroid gland, or to radiation from incorporated hot particles' of an alpha or beta/gamma emitter is discussed. It is concluded that the damage to these cells is unlikely to have been caused by radiation. (Author)

  16. Increased frequency of micronucleated exfoliated cells among humans exposed in vivo to mobile telephone radiations

    International Nuclear Information System (INIS)

    Manoj Kumar Sharma; Abhay Singh Yadav

    2007-01-01

    Complete text of publication follows. The health concerns have been raised following the enormous increase in the use of wireless mobile telephones through out the world. This investigation had been taken, with the motive to find out whether mobile phone radiations cause any in vivo effects on the frequency of micronucleated exfoliated cells in the exposed subjects. A total of 109 subjects including 85 regular mobile phone users (exposed) and 24 non-users (controls) had participated in this study. Exfoliated cells were obtained by swabbing the buccal-mucosa from exposed as well as sex-age-matched controls. One thousand exfoliated cells were screened from each individual for nuclear anomalies including micronuclei (MN), karyolysis (KL), karyorrhexis (KH), broken egg (BE) and bi-nucleated (BN) cells. The average daily duration of exposure to mobile phone radiations is 61.26 minutes with an overall average duration of exposure in term of years is 2.35 years in exposed subjects along with the 9.84±0.745 MNC (micronucleated cells) and 10.72±0.889 TMN (total micronuclei) as compared to zero duration of exposure along with average 3.75±0.774 MNC and 4.00±0.808 TMN in controls. The means are significantly different in case MNC and TMN at 0.01% level of significance. For all other nuclear anomalies (KL, KH, BE and BN cells) the means are found statistically nonsignificant. A positive correlation was found in the frequency of MNC and TMN with respect to duration of exposure time.

  17. Silver ions increase plasma membrane permeability through modulation of intracellular calcium levels in tobacco BY-2 cells

    Czech Academy of Sciences Publication Activity Database

    Klíma, Petr; Laňková, Martina; Vandenbussche, F.; Van Der Straeten, D.; Petrášek, Jan

    2018-01-01

    Roč. 37, č. 5 (2018), s. 809-818 ISSN 0721-7714 R&D Projects: GA ČR GA16-10948S Grant - others:OPPK(XE) CZ.2.16/3.1.00/21519 Institutional support: RVO:61389030 Keywords : Auxin * Calcium * Ethylene * Silver ions * Tobacco BY-2 cells * Transmembrane transport Subject RIV: ED - Physiology OBOR OECD: Cell biology Impact factor: 2.869, year: 2016

  18. Inhibition of Cycloartenol Synthase (CAS) Function in Tobacco BY-2 Cell Suspensions: A Proteomic Analysis.

    Science.gov (United States)

    Gas-Pascual, Elisabet; Simonovik, Biljana; Heintz, Dimitri; Bergdoll, Marc; Schaller, Hubert; Bach, Thomas J

    2015-08-01

    The effect of an inhibitor of cycloartenol synthase (CAS, EC 5.4.99.8) on the proteome of tobacco BY-2 cells has been examined. CAS catalyzes the first committed step in phytosterol synthesis in plants. BY-2 cells were treated with RO 48-8071, a potent inhibitor of oxidosqualene cyclization. Proteins were separated by two-dimensional electrophoresis and spots, that clearly looked differentially accumulated after visual inspection, were cut, in-gel trypsin digested, and peptides were analyzed by nano-HPLC-MS/MS. Distinct peptides were compared to sequences in the data banks and attributed to corresponding proteins and genes. Inhibition of CAS induced proteins that appear to mitigate the negative effects of the chemical exposure. However, as all enzymes that are directly involved in phytosterol biosynthesis are low-abundant proteins, significant changes in their levels could not be observed. Differences could be seen with enzymes involved in primary metabolism (glycolysis, pentose phosphate pathway etc.), in proteins of the chaperonin family, and those, like actin, that participate in formation and strengthening of the cytoskeleton and have some impact on cell growth and division.

  19. Inhibition of Cycloartenol Synthase (CAS) Function in Tobacco BY-2 Cells.

    Science.gov (United States)

    Gas-Pascual, Elisabet; Simonovik, Biljana; Schaller, Hubert; Bach, Thomas J

    2015-08-01

    Tobacco BY-2 cell suspensions are our preferred model for studying isoprenoid biosynthesis pathways, due to their easy genetic transformation and the efficient absorption of metabolic precursors, intermediates, and/or inhibitors. Using this model system, we have analyzed the effects of chemical and genetic blockage of cycloartenol synthase (CAS, EC 5.4.99.8), an oxidosqualene cyclase that catalyzes the first committed step in the sterol pathway of plants. BY-2 cells were treated with RO 48-8071, a potent inhibitor of oxidosqualene cyclization. Short-term treatments (24 h) resulted in accumulation of oxidosqualene with no changes in the final sterol products. Interestingly, long-term treatments (6 days) induced down-regulation in gene expression not only of CAS but also of the SMT2 gene coding sterol methyltransferase 2 (EC 2.1.1.41). This explains some of the increase in 24-methyl sterols at the expense of the 24-ethyl sterols stigmasterol and sitosterol. In our alternative strategy, CAS gene expression was partially blocked by using an inducible artificial microRNA. The limited effectiveness of this approach might be explained by some dependence of the machinery for RNAi formation on an operating MVA/sterol pathway. For comparison we checked the effect of RO 48-8071 on a green cell suspension of Arabidopsis and on seedlings, containing a small spectrum of triterpenes besides phytosterols. Triterpenes remained essentially unaffected, but phytosterol accumulation was clearly diminished.

  20. Effect of Magnetic Nanoparticles on Tobacco BY-2 Cell Suspension Culture

    Science.gov (United States)

    Krystofova, Olga; Sochor, Jiri; Zitka, Ondrej; Babula, Petr; Kudrle, Vit; Adam, Vojtech; Kizek, Rene

    2012-01-01

    Nanomaterials are structures whose exceptionality is based on their large surface, which is closely connected with reactivity and modification possibilities. Due to these properties nanomaterials are used in textile industry (antibacterial textiles with silver nanoparticles), electronics (high-resolution imaging, logical circuits on the molecular level) and medicine. Medicine represents one of the most important fields of application of nanomaterials. They are investigated in connection with targeted therapy (infectious diseases, malignant diseases) or imaging (contrast agents). Nanomaterials including nanoparticles have a great application potential in the targeted transport of pharmaceuticals. However, there are some negative properties of nanoparticles, which must be carefully solved, as hydrophobic properties leading to instability in aqueous environment, and especially their possible toxicity. Data about toxicity of nanomaterials are still scarce. Due to this fact, in this work we focused on studying of the effect of magnetic nanoparticles (NPs) and modified magnetic nanoparticles (MNPs) on tobacco BY-2 plant cell suspension culture. We aimed at examining the effect of NPs and MNPs on growth, proteosynthesis—total protein content, thiols—reduced (GSH) and oxidized (GSSG) glutathione, phytochelatins PC2-5, glutathione S-transferase (GST) activity and antioxidant activity of BY-2 cells. Whereas the effect of NPs and MNPs on growth of cell suspension culture was only moderate, significant changes were detected in all other biochemical parameters. Significant changes in protein content, phytochelatins levels and GST activity were observed in BY-2 cells treated with MNPs nanoparticles treatment. Changes were also clearly evident in the case of application of NPs. Our results demonstrate the ability of MNPs to negatively affect metabolism and induce biosynthesis of protective compounds in a plant cell model represented by BY-2 cell suspension culture. The

  1. In vivo imaging and quantitative monitoring of autophagic flux in tobacco BY-2 cells.

    Science.gov (United States)

    Hanamata, Shigeru; Kurusu, Takamitsu; Okada, Masaaki; Suda, Akiko; Kawamura, Koki; Tsukada, Emi; Kuchitsu, Kazuyuki

    2013-01-01

    Autophagy has been shown to play essential roles in the growth, development and survival of eukaryotic cells. However, simple methods for quantification and visualization of autophagic flux remain to be developed in living plant cells. Here, we analyzed the autophagic flux in transgenic tobacco BY-2 cell lines expressing fluorescence-tagged NtATG8a as a marker for autophagosome formation. Under sucrose-starved conditions, the number of punctate signals of YFP-NtATG8a increased, and the fluorescence intensity of the cytoplasm and nucleoplasm decreased. Conversely, these changes were not observed in BY-2 cells expressing a C-terminal glycine deletion mutant of the NtATG8a protein (NtATG8aΔG). To monitor the autophagic flux more easily, we generated a transgenic BY-2 cell line expressing NtATG8a fused to a pH-sensitive fluorescent tag, a tandem fusion of the acid-insensitive RFP and the acid-sensitive YFP. In sucrose-rich conditions, both fluorescent signals were detected in the cytoplasm and only weakly in the vacuole. In contrast, under sucrose-starved conditions, the fluorescence intensity of the cytoplasm decreased, and the RFP signal clearly increased in the vacuole, corresponding to the fusion of the autophagosome to the vacuole and translocation of ATG8 from the cytoplasm to the vacuole. Moreover, we introduce a novel simple easy way to monitor the autophagic flux non-invasively by only measuring the ratio of fluorescence of RFP and YFP in the cell suspension using a fluorescent image analyzer without microscopy. The present in vivo quantitative monitoring system for the autophagic flux offers a powerful tool for determining the physiological functions and molecular mechanisms of plant autophagy induced by environmental stimuli.

  2. Apoptosis and necroptosis are induced in rainbow trout cell lines exposed to cadmium

    Energy Technology Data Exchange (ETDEWEB)

    Krumschnabel, Gerhard, E-mail: Gerhard.Krumschnabel@i-med.ac.at [Division of Developmental Immunology, Biocenter, Medical University Innsbruck, Fritz-Preglstr. 3, Innsbruck (Austria); Ebner, Hannes L.; Hess, Michael W. [Division of Histology and Embryology, Medical University Innsbruck, Innsbruck (Austria); Villunger, Andreas [Division of Developmental Immunology, Biocenter, Medical University Innsbruck, Fritz-Preglstr. 3, Innsbruck (Austria)

    2010-08-01

    Cadmium is an important environmental toxicant that can kill cells. A number of studies have implicated apoptosis as well as necrosis and, most recently, a form of programmed necrosis termed necroptosis in the process of cadmium-mediated toxicity, but the exact mechanism remains ill-defined and may depend on the affected cell type. This study investigated which mode of cell death may be responsible for cell death induction in cadmium-exposed trout cell lines from gill and liver and if this cell death was sensitive to inhibitors of necroptosis or apoptosis, respectively. It was observed that intermediate levels of cadmium that killed approximately 50% of the cells over 96-120 h of exposure caused cell death that morphologically resembled apoptosis and was associated with an increase of apoptotic markers such as the number of cells with diminished DNA content (sub-G1 cells), condensed or fragmented nuclei, and elevation of caspase-3 activity. At the same time, however, cells also lost plasma membrane integrity, as indicated by uptake of propidium iodide, showed a decrease of ATP levels and mitochondrial membrane potential, and displayed cell swelling, signs associated with secondary necrosis, or equally possible, necroptotic cell death. Importantly, many of these alterations were at least partly inhibited by the necroptosis inhibitor necrostatin-1 and were to a lesser extent also sensitive to the pan-caspase inhibitor zVAD-fmk, indicating that multiple modes of cell death are concurrently induced in cadmium-exposed trout cells, including necroptosis and apoptosis. Cell death appeared to lack concurrent radical formation, consistent with genetically regulated necroptotic cell death, but was characterized by the rapid induction of DNA damage markers, and the early onset of disintegration of the Golgi complex. Comparative experiments evaluating copper-toxicity indicated that in comparison to cadmium much higher concentrations of this metal were required to induce cell

  3. Apoptosis and necroptosis are induced in rainbow trout cell lines exposed to cadmium

    International Nuclear Information System (INIS)

    Krumschnabel, Gerhard; Ebner, Hannes L.; Hess, Michael W.; Villunger, Andreas

    2010-01-01

    Cadmium is an important environmental toxicant that can kill cells. A number of studies have implicated apoptosis as well as necrosis and, most recently, a form of programmed necrosis termed necroptosis in the process of cadmium-mediated toxicity, but the exact mechanism remains ill-defined and may depend on the affected cell type. This study investigated which mode of cell death may be responsible for cell death induction in cadmium-exposed trout cell lines from gill and liver and if this cell death was sensitive to inhibitors of necroptosis or apoptosis, respectively. It was observed that intermediate levels of cadmium that killed approximately 50% of the cells over 96-120 h of exposure caused cell death that morphologically resembled apoptosis and was associated with an increase of apoptotic markers such as the number of cells with diminished DNA content (sub-G1 cells), condensed or fragmented nuclei, and elevation of caspase-3 activity. At the same time, however, cells also lost plasma membrane integrity, as indicated by uptake of propidium iodide, showed a decrease of ATP levels and mitochondrial membrane potential, and displayed cell swelling, signs associated with secondary necrosis, or equally possible, necroptotic cell death. Importantly, many of these alterations were at least partly inhibited by the necroptosis inhibitor necrostatin-1 and were to a lesser extent also sensitive to the pan-caspase inhibitor zVAD-fmk, indicating that multiple modes of cell death are concurrently induced in cadmium-exposed trout cells, including necroptosis and apoptosis. Cell death appeared to lack concurrent radical formation, consistent with genetically regulated necroptotic cell death, but was characterized by the rapid induction of DNA damage markers, and the early onset of disintegration of the Golgi complex. Comparative experiments evaluating copper-toxicity indicated that in comparison to cadmium much higher concentrations of this metal were required to induce cell

  4. Mechanisms of mutagenesis in human cells exposed to 55 MeV protons

    Science.gov (United States)

    Gauny, S.; Wiese, C.; Kronenberg, A.

    2001-01-01

    Protons represent the major type of charged particle radiation in spaceflight environments. The purpose of this study was to assess mutations arising in human lymphoid cells exposed to protons. Mutations were quantitated at the thymidine kinase (TK1) locus in cell lines derived from the same donor: TK6 cells (wt TP53) and WTK1 cells (mutant TP53). WTK1 cells were much more susceptible to mutagenesis following proton exposure than TK6 cells. Intragenic deletions were observed among early-arising TK1 mutants in TK6 cells, but not in WTK1 cells where all of the mutants arose by LOH. Deletion was the predominant mode of LOH in TK6 cells, while allelic recombination was the major mode of LOH in WTK1 cells. Deletions were of variable lengths, from <1 cM to 64 cM, while mutations that arose by allelic recombination often extended to the telomere. In summary, proton exposures elicited many types of mutations at an autosomal locus in human cells. Most involved large scale loss of genetic information, either through deletion or by recombination.

  5. Frequency patterns of T-cell exposed motifs in immunoglobulin heavy chain peptides presented by MHCs

    Directory of Open Access Journals (Sweden)

    Robert D. Bremel

    2014-10-01

    Full Text Available Immunoglobulins are highly diverse protein sequences that are processed and presented to T-cells by B-cells and other antigen presenting cells. We examined a large dataset of immunoglobulin heavy chain variable regions (IGHV to assess the diversity of T-cell exposed motifs (TCEM. TCEM comprise those amino acids in a MHC-bound peptide which face outwards, surrounded by the MHC histotope, and which engage the T-cell receptor. Within IGHV there is a distinct pattern of predicted MHC class II binding and a very high frequency of re-use of the TCEMs. The re-use frequency indicates that only a limited number of different cognate T-cells are required to engage many different clonal B-cells. The amino acids in each outward-facing TCEM are intercalated with the amino acids of inward-facing MHC groove-exposed motifs (GEM. Different GEM may have differing, allele-specific, MHC binding affinities. The intercalation of TCEM and GEM in a peptide allows for a vast combinatorial repertoire of epitopes, each eliciting a different response. Outcome of T-cell receptor binding is determined by overall signal strength, which is a function of the number of responding T-cells and the duration of engagement. Hence, the frequency of T-cell exposed motif re-use appears to be an important determinant of whether a T-cell response is stimulatory or suppressive. The frequency distribution of TCEMs implies that somatic hypermutation is followed by clonal expansion that develop along repeated pathways. The observations of TCEM and GEM derived from immunoglobulins suggest a relatively simple, yet powerful, mechanism to correlate T-cell polyspecificity, through re-use of TCEMs, with a very high degree of specificity achieved by combination with a diversity of GEMs. The frequency profile of TCEMs also points to an economical mechanism for maintaining T-cell memory, recall, and self-discrimination based on an endogenously generated profile of motifs.

  6. Cytokinin-induced cell death is associated with elevated expression of alternative oxidase in tobacco BY-2 cells.

    Science.gov (United States)

    Mlejnek, Petr

    2013-10-01

    N(6)-benzyladenine (BA) and N(6)-benzyladenosine ([9R]BA) induce massive production of reactive oxygen species (ROS) that is eventually followed by a loss of cell viability in tobacco BY-2 cells (Mlejnek et al. Plant Cell Environ 26:1723-1735, 2003, Plant Sci 168:389-395, 2005). Results presented in this work suggest that the main sources of ROS are likely mitochondria and that the maintenance of the mitochondrial transmembrane potential is crucial for ROS production in cytokinin-treaded BY-2 cells. Therefore, the possible involvement of alternative oxidase (AOX) in cell death process induced by BA and [9R]BA was studied. About three- to fourfold increase in mRNA levels of AOX1 was observed a few hours after the BA and [9R]BA addition into the growth medium. The elevated expression of AOX1 mRNA could be prevented by adding adenine and adenosine which simultaneously reduced the cytotoxic effects of BA and [9R]BA, respectively. N(6)-benzyladenine 7-β-D-glucoside ([7G]BA) which is a common non-toxic metabolite of BA and [9R]BA did not affect the AOX1 mRNA expression. Although AOX1 seemed to be involved in protection of BY-2 cells against the abiotic stress induced by BA and [9R]BA, the results do not support the idea that it protects cells from death exclusively by scavenging of reactive oxygen species. Indeed, N-propyl gallate, an inhibitor of AOX, decreased cell survival despite it concomitantly decreased the ROS production. This finding is in contrast to the effect of salicylhydroxamic acid, another well-known inhibitor of AOX, which also increased the number of dying cells while it increased the ROS production.

  7. Enhanced micronucleus formation in the descendants of {gamma}-ray-irradiated tobacco cells: Evidence for radiation-induced genomic instability in plant cells

    Energy Technology Data Exchange (ETDEWEB)

    Yokota, Yuichiro, E-mail: yokota.yuichiro@jaea.go.jp [Life Science and Biotechnology Division, Quantum Beam Science Directorate, Japan Atomic Energy Agency, 1233 Watanuki-machi, Takasaki, Gunma 370-1292 (Japan); Funayama, Tomoo; Hase, Yoshihiro [Life Science and Biotechnology Division, Quantum Beam Science Directorate, Japan Atomic Energy Agency, 1233 Watanuki-machi, Takasaki, Gunma 370-1292 (Japan); Hamada, Nobuyuki [Radiation Safety Research Center, Nuclear Technology Research Laboratory, Central Research Institute of Electric Power Industry, 2-11-1 Iwado-kita, Komae, Tokyo 201-8511 (Japan); Kobayashi, Yasuhiko; Tanaka, Atsushi; Narumi, Issay [Life Science and Biotechnology Division, Quantum Beam Science Directorate, Japan Atomic Energy Agency, 1233 Watanuki-machi, Takasaki, Gunma 370-1292 (Japan)

    2010-09-10

    Ionizing radiation-induced genomic instability has been documented in various end points such as chromosomal aberrations and mutations, which arises in the descendants of irradiated mammalian or yeast cells many generations after the initial insult. This study aimed at addressing radiation-induced genomic instability in higher plant tobacco cells. We thus investigated micronucleus (MN) formation and cell proliferation in tobacco cells irradiated with {gamma}-rays and their descendants. In {gamma}-irradiated cells, cell cycle was arrested at G{sub 2}/M phase at around 24 h post-irradiation but released afterward. In contrast, MN frequency peaked at 48 h post-irradiation. Almost half of 40 Gy-irradiated cells had MN at 48 h post-irradiation, but proliferated as actively as sham-irradiated cells up to 120 h post-irradiation. Moreover, the descendants that have undergone at least 22 generations after irradiation still showed a two-fold MN frequency compared to sham-irradiated cells. This is the direct evidence for radiation-induced genomic instability in tobacco cells.

  8. Mechanisms of Cancer Induction by Tobacco-Specific NNK and NNN

    Energy Technology Data Exchange (ETDEWEB)

    Xue, Jiaping [Department of Physiology and Biophysics, University of Illinois at Chicago, Chicago, IL 60612 (United States); Yang, Suping; Seng, Seyha, E-mail: sseng@bidmc.harvard.edu [Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215 (United States)

    2014-05-14

    Tobacco use is a major public health problem worldwide. Tobacco-related cancers cause millions of deaths annually. Although several tobacco agents play a role in the development of tumors, the potent effects of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N'-nitrosonornicotine (NNN) are unique. Metabolically activated NNK and NNN induce deleterious mutations in oncogenes and tumor suppression genes by forming DNA adducts, which could be considered as tumor initiation. Meanwhile, the binding of NNK and NNN to the nicotinic acetylcholine receptor promotes tumor growth by enhancing and deregulating cell proliferation, survival, migration, and invasion, thereby creating a microenvironment for tumor growth. These two unique aspects of NNK and NNN synergistically induce cancers in tobacco-exposed individuals. This review will discuss various types of tobacco products and tobacco-related cancers, as well as the molecular mechanisms by which nitrosamines, such as NNK and NNN, induce cancer.

  9. A versatile coupled cell-free transcription-translation system based on tobacco BY-2 cell lysates.

    Science.gov (United States)

    Buntru, Matthias; Vogel, Simon; Stoff, Katrin; Spiegel, Holger; Schillberg, Stefan

    2015-05-01

    Cell-free protein synthesis is a powerful method for the high-throughput production of recombinant proteins, especially proteins that are difficult to express in living cells. Here we describe a coupled cell-free transcription-translation system based on tobacco BY-2 cell lysates (BYLs). Using a combination of fractional factorial designs and response surface models, we developed a cap-independent system that produces more than 250 μg/mL of functional enhanced yellow fluorescent protein (eYFP) and about 270 μg/mL of firefly luciferase using plasmid templates, and up to 180 μg/mL eYFP using linear templates (PCR products) in 18 h batch reactions. The BYL contains actively-translocating microsomal vesicles derived from the endoplasmic reticulum, promoting the formation of disulfide bonds, glycosylation and the cotranslational integration of membrane proteins. This was demonstrated by expressing a functional full-size antibody (∼ 150 μg/mL), the model enzyme glucose oxidase (GOx) (∼ 7.3 U/mL), and a transmembrane growth factor (∼ 25 μg/mL). Subsequent in vitro treatment of GOx with peptide-N-glycosidase F confirmed the presence of N-glycans. Our results show that the BYL can be used as a high-throughput expression and screening platform that is particularly suitable for complex and cytotoxic proteins. © 2014 Wiley Periodicals, Inc.

  10. Affinity Purification and Characterization of Functional Tubulin from Cell Suspension Cultures of Arabidopsis and Tobacco1

    Science.gov (United States)

    Fujita, Satoshi; Uchimura, Seiichi; Noguchi, Masahiro; Demura, Taku

    2016-01-01

    Microtubules assemble into several distinct arrays that play important roles in cell division and cell morphogenesis. To decipher the mechanisms that regulate the dynamics and organization of this versatile cytoskeletal component, it is essential to establish in vitro assays that use functional tubulin. Although plant tubulin has been purified previously from protoplasts by reversible taxol-induced polymerization, a simple and efficient purification method has yet to be developed. Here, we used a Tumor Overexpressed Gene (TOG) column, in which the tubulin-binding domains of a yeast (Saccharomyces cerevisiae) TOG homolog are immobilized on resin, to isolate functional plant tubulin. We found that several hundred micrograms of pure tubulin can readily be purified from cell suspension cultures of tobacco (Nicotiana tabacum) and Arabidopsis (Arabidopsis thaliana). The tubulin purified by the TOG column showed high assembly competence, partly because of low levels of polymerization-inhibitory phosphorylation of α-tubulin. Compared with porcine brain tubulin, Arabidopsis tubulin is highly dynamic in vitro at both the plus and minus ends, exhibiting faster shrinkage rates and more frequent catastrophe events, and exhibits frequent spontaneous nucleation. Furthermore, our study shows that an internal histidine tag in α-tubulin can be used to prepare particular isotypes and specifically engineered versions of α-tubulin. In contrast to previous studies of plant tubulin, our mass spectrometry and immunoblot analyses failed to detect posttranslational modification of the isolated Arabidopsis tubulin or detected only low levels of posttranslational modification. This novel technology can be used to prepare assembly-competent, highly dynamic pure tubulin from plant cell cultures. PMID:26747285

  11. High-level production of human interleukin-10 fusions in tobacco cell suspension cultures

    Science.gov (United States)

    Kaldis, Angelo; Ahmad, Adil; Reid, Alexandra; McGarvey, Brian; Brandle, Jim; Ma, Shengwu; Jevnikar, Anthony; Kohalmi, Susanne E; Menassa, Rima

    2013-01-01

    The production of pharmaceutical proteins in plants has made much progress in recent years with the development of transient expression systems, transplastomic technology and humanizing glycosylation patterns in plants. However, the first therapeutic proteins approved for administration to humans and animals were made in plant cell suspensions for reasons of containment, rapid scale-up and lack of toxic contaminants. In this study, we have investigated the production of human interleukin-10 (IL-10) in tobacco BY-2 cell suspension and evaluated the effect of an elastin-like polypeptide tag (ELP) and a green fluorescent protein (GFP) tag on IL-10 accumulation. We report the highest accumulation levels of hIL-10 obtained with any stable plant expression system using the ELP fusion strategy. Although IL-10-ELP has cytokine activity, its activity is reduced compared to unfused IL-10, likely caused by interference of ELP with folding of IL-10. Green fluorescent protein has no effect on IL-10 accumulation, but examining the trafficking of IL-10-GFP over the cell culture cycle revealed fluorescence in the vacuole during the stationary phase of the culture growth cycle. Analysis of isolated vacuoles indicated that GFP alone is found in vacuoles, while the full-size fusion remains in the whole-cell extract. This indicates that GFP is cleaved off prior to its trafficking to the vacuole. On the other hand, IL-10-GFP-ELP remains mostly in the ER and accumulates to high levels. Protein bodies were observed at the end of the culture cycle and are thought to arise as a consequence of high levels of accumulation in the ER. PMID:23297698

  12. Production, secretion, and stability of human secreted alkaline phosphatase in tobacco NT1 cell suspension cultures.

    Science.gov (United States)

    Becerra-Arteaga, Alejandro; Mason, Hugh S; Shuler, Michael L

    2006-01-01

    Tobacco NT1 cell suspension cultures secreting active human secreted alkaline phosphatase (SEAP) were generated for the first time as a model system to study recombinant protein production, secretion, and stability in plant cell cultures. The SEAP gene encodes a secreted form of the human placental alkaline phosphatase (PLAP). During batch culture, the highest level of active SEAP in the culture medium (0.4 U/mL, corresponding to approximately 27 mg/L) was observed at the end of the exponential growth phase. Although the level of active SEAP decreased during the stationary phase, the activity loss did not appear to be due to SEAP degradation (based on Western blots) but due to SEAP denaturation. The protein-stabilizing agents polyvinylpirrolidone (PVP) and bacitracin were added extracellularly to test for their ability to reduce the loss of SEAP activity during the stationary phase. Bacitracin (100 mg/L) was the most effective treatment at sustaining activity levels for up to 17 days post-subculture. Commercially available human placental alkaline phosphatase (PLAP) was used to probe the mechanism of SEAP deactivation. Experiments with PLAP in sterile and conditioned medium corroborated the denaturation of SEAP by factors generated by cell growth and not due to simple proteolysis. We also show for the first time that the factors promoting activity loss are heat labile at 95 degrees C but not at 70 degrees C, and they are not inactivated after a 5 day incubation period under normal culture conditions (27 degrees C). In addition, there were no significant changes in pH or redox potential when comparing sterile and cell-free conditioned medium during PLAP incubation, indicating that these factors were unimportant.

  13. Immune cells in Chernobyl recovery operation workers exposed over 500 mSv

    International Nuclear Information System (INIS)

    Bazyka, D.; Byelyaeva, N.; Chumak, A.; Golyarnik, N.; Maznichenko, O.; Kovalenko, Ju.

    2004-01-01

    Immune response parameters were studied in Chernobyl radiation emergency workers exposed to radiation over 500 mSv during 1986. Initial response stage to the radiation exposure was characterised by immunological deficiency with T-cell subsets changes. In the reconstitution period inhibition of immune function was associated with lymphocyte subset changes such as decreased CD3 + and CD4 + cell counts and increased number of somatic mutations in TCR-locus. Late period after the acute radiation exposure in Chernobyl radiation emergency workers is characterized by decreased CD8 + suppressor cell function that could lead to poor proliferation control. Subset analysis of CD34 + cells showed in ARS survivors counts significantly higher than in control especially for the most primitive progenitors with CD34 + CD90 + CD45 -/l ow and CD34 + CD45 - CD38 - phenotypes. (author)

  14. Protective Effects of Hydroalcoholic Extract of Nasturtium officinale on Rat Blood Cells Exposed to Arsenic

    Directory of Open Access Journals (Sweden)

    Felor Zargari

    2015-06-01

    Full Text Available Background: Arsenic is one of the most toxic metalloids. Anemia and leukopenia are common results of poisoning with arsenic, which may happen due to a direct hemolytic or cytotoxic effect on blood cells. The aim of this study was to examine the effects of hydroalcoholic extract of Nasturtium officinale on blood cells and antioxidant enzymes in rats exposed to sodium (metaarsenite. Methods: 32 Male Sprague Dawley rats were randomly divided into four groups; Group I (normal healthy rats, Group II (treated with 5.5mg/kg of body weight of NaAsO2, Group III (treated with 500mg/kg of body weight of hydro-alcoholic extract of N. officinale, and Group IV (treated with group II and III supplementations. Blood samples were collected and red blood cell, white blood cell, hematocrit, hemoglobin, platelet, total protein and albumin levels and total antioxidant capacity were measured. Data was analyzed with Mann-Whitney U test. Results: WBC, RBC and Hct were decreased in the rats exposed to NaAsO2 (p<0.05. A significant increase was seen in RBC and Hct after treatment with the plant extract (p<0.05. There was no significant decrease in serum albumin and total protein in the groups exposed to NaAsO2 compared to the group I, but NaAsO2 decreased the total antioxidant capacity, significantly. Conclusion: The Nasturtium officinale extract have protective effect on arsenic-induced damage of blood cells.

  15. Chromosomal Aberrations in Normal and AT Cells Exposed to High Dose of Low Dose Rate Irradiation

    Science.gov (United States)

    Kawata, T.; Shigematsu, N.; Kawaguchi, O.; Liu, C.; Furusawa, Y.; Hirayama, R.; George, K.; Cucinotta, F.

    2011-01-01

    Ataxia telangiectasia (A-T) is a human autosomally recessive syndrome characterized by cerebellar ataxia, telangiectases, immune dysfunction, and genomic instability, and high rate of cancer incidence. A-T cell lines are abnormally sensitive to agents that induce DNA double strand breaks, including ionizing radiation. The diverse clinical features in individuals affected by A-T and the complex cellular phenotypes are all linked to the functional inactivation of a single gene (AT mutated). It is well known that cells deficient in ATM show increased yields of both simple and complex chromosomal aberrations after high-dose-rate irradiation, but, less is known on how cells respond to low-dose-rate irradiation. It has been shown that AT cells contain a large number of unrejoined breaks after both low-dose-rate irradiation and high-dose-rate irradiation, however sensitivity for chromosomal aberrations at low-dose-rate are less often studied. To study how AT cells respond to low-dose-rate irradiation, we exposed confluent normal and AT fibroblast cells to up to 3 Gy of gamma-irradiation at a dose rate of 0.5 Gy/day and analyzed chromosomal aberrations in G0 using fusion PCC (Premature Chromosomal Condensation) technique. Giemsa staining showed that 1 Gy induces around 0.36 unrejoined fragments per cell in normal cells and around 1.35 fragments in AT cells, whereas 3Gy induces around 0.65 fragments in normal cells and around 3.3 fragments in AT cells. This result indicates that AT cells can rejoin breaks less effectively in G0 phase of the cell cycle? compared to normal cells. We also analyzed chromosomal exchanges in normal and AT cells after exposure to 3 Gy of low-dose-rate rays using a combination of G0 PCC and FISH techniques. Misrejoining was detected in the AT cells only? When cells irradiated with 3 Gy were subcultured and G2 chromosomal aberrations were analyzed using calyculin-A induced PCC technique, the yield of unrejoined breaks decreased in both normal and AT

  16. Opium, tobacco, and alcohol use in relation to oesophageal squamous cell carcinoma in a high-risk area of Iran

    Science.gov (United States)

    Nasrollahzadeh, D; Kamangar, F; Aghcheli, K; Sotoudeh, M; Islami, F; Abnet, C C; Shakeri, R; Pourshams, A; Marjani, H A; Nouraie, M; Khatibian, M; Semnani, S; Ye, W; Boffetta, P; Dawsey, S M; Malekzadeh, R

    2008-01-01

    The very high incidence of oesophageal squamous cell carcinoma (ESCC) in Golestan Province in northeastern Iran was suggested by studies in the 1970s as partly due to opium use, which is not uncommon in this area, but based on limited numbers. From December 2003 to June 2007, we administered a validated structured questionnaire to 300 ESCC cases and 571 controls, matched on neighbourhood of residence, age (±2 years), and sex. We used conditional logistic regression models to calculate odds ratios (ORs) and 95% confidence intervals (95% CIs) adjusted for potential confounders. Compared with those who used neither tobacco nor opium, risk of ESCC was increased in those who used tobacco only (OR, 95% CI: 1.70, 1.05–2.73), in those who used opium only (2.12, 1.21–3.74), and in those who used both tobacco and opium (2.35, 1.50–3.67). All forms of tobacco use (cigarettes, hookah, and nass) were associated with higher ESCC risk. Similarly, use of both crude opium and other forms of opium were associated with higher risk. Alcohol consumption was seen in only 2% of the cases and 2% of the controls, and was not associated with ESCC risk. PMID:18475303

  17. Live-cell imaging study of mitochondrial morphology in mammalian cells exposed to X-rays

    International Nuclear Information System (INIS)

    Noguchi, M.; Yokoya, A.; Narita, A.; Fujii, K.; Kanari, Y.

    2015-01-01

    Morphological changes in mitochondria induced by X-irradiation in normal murine mammary gland cells were studied with a live-cell microscopic imaging technique. Mitochondria were visualised by staining with a specific fluorescent probe in the cells, which express fluorescent ubiquitination-based cell-cycle indicator 2 (Fucci2) probes to visualise cell cycle. In unirradiated cells, the number of cells with fragmented mitochondria was about 20 % of the total cells through observation period (96 h). In irradiated cells, the population with fragmented mitochondria significantly increased depending on the absorbed dose. Particularly, for 8 Gy irradiation, the accumulation of fragmentation persists even in the cells whose cell cycle came to a stand (80 % in G1 (G0-like) phase). The fraction reached to a maximum at 96 h after irradiation. The kinetics of the fraction with fragmented mitochondria was similar to that for cells in S/G2/M phase (20 %) through the observation period (120 h). The evidences show that, in irradiated cells, some signals are continually released from a nucleus or cytoplasm even in the G0-like cells to operate some sort of protein machineries involved in mitochondrial fission. It is inferred that this delayed mitochondrial fragmentation is strongly related to their dysfunction, and hence might modulate radiobiological effects such as mutation or cell death. (authors)

  18. The transcriptome of HIV-1 infected intestinal CD4+ T cells exposed to enteric bacteria.

    Directory of Open Access Journals (Sweden)

    Alyson C Yoder

    2017-02-01

    Full Text Available Global transcriptome studies can help pinpoint key cellular pathways exploited by viruses to replicate and cause pathogenesis. Previous data showed that laboratory-adapted HIV-1 triggers significant gene expression changes in CD4+ T cell lines and mitogen-activated CD4+ T cells from peripheral blood. However, HIV-1 primarily targets mucosal compartments during acute infection in vivo. Moreover, early HIV-1 infection causes extensive depletion of CD4+ T cells in the gastrointestinal tract that herald persistent inflammation due to the translocation of enteric microbes to the systemic circulation. Here, we profiled the transcriptome of primary intestinal CD4+ T cells infected ex vivo with transmitted/founder (TF HIV-1. Infections were performed in the presence or absence of Prevotella stercorea, a gut microbe enriched in the mucosa of HIV-1-infected individuals that enhanced both TF HIV-1 replication and CD4+ T cell death ex vivo. In the absence of bacteria, HIV-1 triggered a cellular shutdown response involving the downregulation of HIV-1 reactome genes, while perturbing genes linked to OX40, PPAR and FOXO3 signaling. However, in the presence of bacteria, HIV-1 did not perturb these gene sets or pathways. Instead, HIV-1 enhanced granzyme expression and Th17 cell function, inhibited G1/S cell cycle checkpoint genes and triggered downstream cell death pathways in microbe-exposed gut CD4+ T cells. To gain insights on these differential effects, we profiled the gene expression landscape of HIV-1-uninfected gut CD4+ T cells exposed to bacteria. Microbial exposure upregulated genes involved in cellular proliferation, MAPK activation, Th17 cell differentiation and type I interferon signaling. Our findings reveal that microbial exposure influenced how HIV-1 altered the gut CD4+ T cell transcriptome, with potential consequences for HIV-1 susceptibility, cell survival and inflammation. The HIV-1- and microbe-altered pathways unraveled here may serve as a

  19. Phosphatidylserine-exposing blood and endothelial cells contribute to the hypercoagulable state in essential thrombocythemia patients.

    Science.gov (United States)

    Tong, Dongxia; Yu, Muxin; Guo, Li; Li, Tao; Li, Jihe; Novakovic, Valerie A; Dong, Zengxiang; Tian, Ye; Kou, Junjie; Bi, Yayan; Wang, Jinghua; Zhou, Jin; Shi, Jialan

    2018-04-01

    The mechanisms of thrombogenicity in essential thrombocythemia (ET) are complex and not well defined. Our objective was to explore whether phosphatidylserine (PS) exposure on blood cells and endothelial cells (ECs) can account for the increased thrombosis and distinct thrombotic risks among mutational subtypes in ET. Using flow cytometry and confocal microscopy, we found that the levels of PS-exposing erythrocytes, platelets, leukocytes, and serum-cultured ECs were significantly higher in each ET group [JAK2, CALR, and triple-negative (TN) (all P cells and serum-cultured ECs led to markedly shortened coagulation time and dramatically increased levels of FXa, thrombin, and fibrin production. This procoagulant activity could be largely blocked by addition of lactadherin (approx. 70% inhibition). Confocal microscopy showed that the FVa/FXa complex and fibrin fibrils colocalized with PS on ET serum-cultured ECs. Additionally, we found a relationship between D-dimer, prothrombin fragment F1 + 2, and PS exposure. Our study reveals a previously unrecognized link between hypercoagulability and exposed PS on cells, which might also be associated with distinct thrombotic risks among mutational subtypes in ET. Thus, blocking PS-binding sites may represent a new therapeutic target for preventing thrombosis in ET.

  20. Deciphering early events involved in hyperosmotic stress-induced programmed cell death in tobacco BY-2 cells.

    Science.gov (United States)

    Monetti, Emanuela; Kadono, Takashi; Tran, Daniel; Azzarello, Elisa; Arbelet-Bonnin, Delphine; Biligui, Bernadette; Briand, Joël; Kawano, Tomonori; Mancuso, Stefano; Bouteau, François

    2014-03-01

    Hyperosmotic stresses represent one of the major constraints that adversely affect plants growth, development, and productivity. In this study, the focus was on early responses to hyperosmotic stress- (NaCl and sorbitol) induced reactive oxygen species (ROS) generation, cytosolic Ca(2+) concentration ([Ca(2+)]cyt) increase, ion fluxes, and mitochondrial potential variations, and on their links in pathways leading to programmed cell death (PCD). By using BY-2 tobacco cells, it was shown that both NaCl- and sorbitol-induced PCD seemed to be dependent on superoxide anion (O2·(-)) generation by NADPH-oxidase. In the case of NaCl, an early influx of sodium through non-selective cation channels participates in the development of PCD through mitochondrial dysfunction and NADPH-oxidase-dependent O2·(-) generation. This supports the hypothesis of different pathways in NaCl- and sorbitol-induced cell death. Surprisingly, other shared early responses, such as [Ca(2+)]cyt increase and singlet oxygen production, do not seem to be involved in PCD.

  1. Colon stem cell and crypt dynamics exposed by cell lineage reconstruction.

    Directory of Open Access Journals (Sweden)

    Yitzhak Reizel

    2011-07-01

    Full Text Available Stem cell dynamics in vivo are often being studied by lineage tracing methods. Our laboratory has previously developed a retrospective method for reconstructing cell lineage trees from somatic mutations accumulated in microsatellites. This method was applied here to explore different aspects of stem cell dynamics in the mouse colon without the use of stem cell markers. We first demonstrated the reliability of our method for the study of stem cells by confirming previously established facts, and then we addressed open questions. Our findings confirmed that colon crypts are monoclonal and that, throughout adulthood, the process of monoclonal conversion plays a major role in the maintenance of crypts. The absence of immortal strand mechanism in crypts stem cells was validated by the age-dependent accumulation of microsatellite mutations. In addition, we confirmed the positive correlation between physical and lineage proximity of crypts, by showing that the colon is separated into small domains that share a common ancestor. We gained new data demonstrating that colon epithelium is clustered separately from hematopoietic and other cell types, indicating that the colon is constituted of few progenitors and ruling out significant renewal of colonic epithelium from hematopoietic cells during adulthood. Overall, our study demonstrates the reliability of cell lineage reconstruction for the study of stem cell dynamics, and it further addresses open questions in colon stem cells. In addition, this method can be applied to study stem cell dynamics in other systems.

  2. Molecular pathways in the bystander response of cells exposed to very low fluences of alpha particles

    International Nuclear Information System (INIS)

    Little, J.B.

    2000-01-01

    Full text: We have examined biological effects in cell populations exposed to very low mean doses of alpha radiation by which only a small fraction of the cells are actually traversed by an alpha particle. We showed earlier that an enhanced frequency of sister chromatid exchanges and HPRT mutations occur in the non-irradiated, 'bystander' cells. The frequency of mutations induced by a single alpha particle traversing the nucleus of a cell was increased nearly fivefold at the lowest fluence studied, a result of mutations occurring in bystander cells. This was associated with a similar increase in the induction of micronuclei, indicating the induction of DNA damage in bystander cells. In order to gain information concerning molecular pathways, we studied changes in gene expression in bystander cells in confluent cultures of human diploid fibroblasts or mouse embryo-derived fibroblasts (MEFs) by western analysis and in-situ immunofluorescence. The expression levels of p53, p21 Waf1 and p34 cdc2 were significantly modulated in bystander cells. The upregulation of p53 and p21 Waf1 did not occur in cultures irradiated at low density, and was markedly reduced in the presence of the gap junction inhibitor lindane. The importance of gap-junction mediated intercellular communication was confirmed in connexin-43 knockout MEFs. Western blot analyses and electrophoretic mobility shift assays indicate that the bystander response is suppressed by incubation with superoxide dismutase as well as an inhibitor of NADPH oxidase, and is associated with the induction of NFKB, suggesting the effect is mediated by oxidative stress. The stress-activated protein kinase p38 and its downstream effector ATF2 are also induced in bystander cells independent of oxidative stress. These results will be discussed in terms of whether activation of the p53 damage response pathway is the direct result of signaling from irradiated cells, or rather is a consequence of DNA induced damage in the bystander

  3. Fate of D3 mouse embryonic stem cells exposed to X-rays or carbon ions.

    Science.gov (United States)

    Luft, S; Pignalosa, D; Nasonova, E; Arrizabalaga, O; Helm, A; Durante, M; Ritter, S

    2014-01-15

    The risk of radiation exposure during embryonic development is still a major problem in radiotoxicology. In this study we investigated the response of the murine embryonic stem cell (mESC) line D3 to two radiation qualities: sparsely ionizing X-rays and densely ionizing carbon ions. We analyzed clonogenic cell survival, proliferation, induction of chromosome aberrations as well as the capability of cells to differentiate to beating cardiomyocytes up to 3 days after exposure. Our results show that, for all endpoints investigated, carbon ions are more effective than X-rays at the same radiation dose. Additionally, in long term studies (≥8 days post-irradiation) chromosomal damage and the pluripotency state were investigated. These studies reveal that pluripotency markers are present in the progeny of cells surviving the exposure to both radiation types. However, only in the progeny of X-ray exposed cells the aberration frequency was comparable to that of the control population, while the progeny of carbon ion irradiated cells harbored significantly more aberrations than the control, generally translocations. We conclude that cells surviving the radiation exposure maintain pluripotency but may carry stable chromosomal rearrangements after densely ionizing radiation. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Human papillomavirus-exposed Langerhans cells are activated by stabilized Poly-I:C

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    Diane M. Da Silva

    2015-12-01

    Full Text Available Human papillomaviruses (HPV establish persistent infections because of evolved immune evasion mechanisms, particularly HPV-mediated suppression of the immune functions of Langerhans cells (LC, the antigen presenting cells of the epithelium. Polyinosinic-polycytidilic acid (Poly-I:C is broadly immunostimulatory with the ability to enhance APC expression of costimulatory molecules and inflammatory cytokines resulting in T cell activation. Here we investigated the activation of primary human LC derived from peripheral blood monocytes after exposure to HPV16 virus like particles followed by treatment with stabilized Poly-I:C compounds (s-Poly-I:C, and their subsequent induction of HPV16-specific T cells. Our results indicate that HPV16 particles alone were incapable of inducing LC activation as demonstrated by the lack of costimulatory molecules, inflammatory cytokines, chemokine-directed migration, and HPV16-specific CD8+ T cells in vitro. Conversely, s-Poly-I:C caused significant upregulation of costimulatory molecules and induction of chemokine-directed migration of LC that were pre-exposed to HPV16. In HLA-A*0201-positive donors, s-Poly-I:C treatment was able to induce CD8+ T cell immune responses against HPV16-derived peptides. Thus, s-Poly-I:C compounds are attractive for translation into therapeutics in which they could potentially mediate clearance of persistent HPV infection. Keywords: Papillomavirus, HPV16, Langerhans cells, Immune escape

  5. Altered global gene expression profiles in human gastrointestinal epithelial Caco2 cells exposed to nanosilver

    Directory of Open Access Journals (Sweden)

    Saura C. Sahu

    Full Text Available Extensive consumer exposure to food- and cosmetics-related consumer products containing nanosilver is of public safety concern. Therefore, there is a need for suitable in vitro models and sensitive predictive rapid screening methods to assess their toxicity. Toxicogenomic profile showing subtle changes in gene expressions following nanosilver exposure is a sensitive toxicological endpoint for this purpose. We evaluated the Caco2 cells and global gene expression profiles as tools for predictive rapid toxicity screening of nanosilver. We evaluated and compared the gene expression profiles of Caco-2 cells exposed to 20 nm and 50 nm nanosilver at a concentration 2.5 μg/ml. The global gene expression analysis of Caco2 cells exposed to 20 nm nanosilver showed that a total of 93 genes were altered at 4 h exposure, out of which 90 genes were up-regulated and 3 genes were down-regulated. The 24 h exposure of 20 nm silver altered 15 genes in Caco2 cells, out of which 14 were up-regulated and one was down-regulated. The most pronounced changes in gene expression were detected at 4 h. The greater size (50 nm nanosilver at 4 h exposure altered more genes by more different pathways than the smaller (20 nm one. Metallothioneins and heat shock proteins were highly up-regulated as a result of exposure to both the nanosilvers. The cellular pathways affected by the nanosilver exposure is likely to lead to increased toxicity. The results of our study presented here suggest that the toxicogenomic characterization of Caco2 cells is a valuable in vitro tool for assessing toxicity of nanomaterials such as nanosilver. Keywords: Nanosilver, Silver nanoparticles, Nanoparticles, Toxicogenomics, DNA microarray, Global gene expression profiles, Caco2 cells

  6. Altered Natural Killer Cell Function in HIV-Exposed Uninfected Infants

    Directory of Open Access Journals (Sweden)

    Christiana Smith

    2017-04-01

    Full Text Available ObjectivesHIV-exposed uninfected (HEU infants have higher rates of severe and fatal infections compared with HIV-unexposed (HUU infants, likely due to immune perturbations. We hypothesized that alterations in natural killer (NK cell activity might occur in HEU infants and predispose them to severe infections.DesignCase–control study using cryopreserved peripheral blood mononuclear cells (PBMCs at birth and 6 months from HEU infants enrolled from 2002 to 2009 and HUU infants enrolled from 2011 to 2013.MethodsNK cell phenotype and function were assessed by flow cytometry after 20-h incubation with and without K562 cells.ResultsThe proportion of NK cells among PBMCs was lower at birth in 12 HEU vs. 22 HUU (1.68 vs. 10.30%, p < 0.0001 and at 6 months in 52 HEU vs. 72 HUU (3.09 vs. 4.65%, p = 0.0005. At birth, HEU NK cells demonstrated increased killing of K562 target cells (p < 0.0001 and increased expression of CD107a (21.65 vs. 12.70%, p = 0.047, but these differences resolved by 6 months. Stimulated HEU NK cells produced less interferon (IFNγ at birth (0.77 vs. 2.64%, p = 0.008 and at 6 months (4.12 vs. 8.39%, p = 0.001, and showed reduced perforin staining at 6 months (66.95 vs. 77.30%, p = 0.0008. Analysis of cell culture supernatants indicated that lower NK cell activity in HEU was associated with reduced interleukin (IL-12, IL-15, and IL-18. Addition of recombinant human IL-12 to stimulated HEU PBMCs restored IFNγ production to that seen in stimulated HUU cultures.ConclusionNK cell proportion, phenotype, and function are altered in HEU infants. NK cell cytotoxicity and degranulation are increased in HEU at birth, but HEU NK cells have reduced IFNγ and perforin production, suggesting an adequate initial response, but decreased functional reserve. NK cell function improved with addition of exogenous IL-12, implicating impaired production of IL-12 by accessory cells. Alterations in NK cell and accessory

  7. Polarized localization and borate-dependent degradation of the Arabidopsis borate transporter BOR1 in tobacco BY-2 cells.

    Science.gov (United States)

    Yamauchi, Noboru; Gosho, Tadashi; Asatuma, Satoru; Toyooka, Kiminori; Fujiwara, Toru; Matsuoka, Ken

    2013-01-01

    In Arabidopsis the borate transporter BOR1, which is located in the plasma membrane, is degraded in the presence of excess boron by an endocytosis-mediated mechanism. A similar mechanism was suggested in rice as excess boron decreased rice borate transporter levels, although in this case whether the decrease was dependent on an increase in degradation or a decrease in protein synthesis was not elucidated. To address whether the borate-dependent degradation mechanism is conserved among plant cells, we analyzed the fate of GFP-tagged BOR1 (BOR1-GFP) in transformed tobacco BY-2 cells. Cells expressing BOR1-GFP displayed GFP fluorescence at the plasma membrane, especially at the membrane between two attached cells. The plasma membrane signal was abolished when cells were incubated in medium with a high concentration of borate (3 to 5 mM). This decrease in BOR1-GFP signal was mediated by a specific degradation of the protein after internalization by endocytosis from the plasma membrane. Pharmacological analysis indicated that the decrease in BOR1-GFP largely depends on the increase in degradation rate and that the degradation was mediated by a tyrosine-motif and the actin cytoskeleton. Tyr mutants of BOR1-GFP, which has been shown to inhibit borate-dependent degradation in Arabidopsis root cells, did not show borate-dependent endocytosis in tobacco BY-2 cells. These findings indicate that the borate-dependent degradation machinery of the borate transporter is conserved among plant species.

  8. Programmed cell death of tobacco BY-2 cells induced by still culture conditions is affected by the age of the culture under agitation.

    Science.gov (United States)

    Hiraga, Asahi; Kaneta, Tsuyoshi; Sato, Yasushi; Sato, Seiichi

    2010-01-25

    Evans Blue staining indicated that actively growing tobacco BY-2 cells in the exponential phase died more rapidly than quiescent cells in the stationary phase when the cells cultured under agitation were placed under still conditions. Fifty percent cell death was induced at about 18, 26, 80 and 140 h for early, mid, late exponential- and stationary-phase cells, respectively. Actively growing cells became TUNEL (transferase-mediated dUTP nick end labelling)-positive more rapidly than quiescent cells, suggesting that the cell death evaluated by Evans Blue is accompanied by DNA cleavages. Electrophoresis of genomic DNA showed a typical 'DNA laddering' pattern formed by multiples of about 200 bp internucleosomal units. Chromatin condensation was first detected at least within 24 h by light microscopy, and then cell shrinkage followed. These findings suggest that the death of BY-2 cells induced by still conditions is PCD (programmed cell death).

  9. Proteinase activity in cell nuclei of rats exposed to γ-radiation and methyl nitrosourea

    International Nuclear Information System (INIS)

    Malakhova, L.V.; Surkenova, G.N.; Gaziev, A.I.

    1990-01-01

    Activity of nuclear proteinases in blood and liver cells of rats exposed to whole-body γ-irradiation (10 Gy) has been comparatively studied by the capacity of splitting the caseic substrate. Proteinase activity in nuclei of irradiated rat leukocytes was shown to increase by 2.5 times and to gradually decrease after 48 h reaching 150-160% as compared to the control. Two hours following a single injection of methyl nitrosourea the alteration in the activity of proteinases in nuclei of rat hepatocytes and leukocytes was different from the alteration of this index after γ-irradiation

  10. Evidence that proliferation of golgi apparatus depends on both de novo generation from the endoplasmic reticulum and formation from pre-existing stacks during the growth of tobacco BY-2 cells.

    Science.gov (United States)

    Abiodun, Moses Olabiyi; Matsuoka, Ken

    2013-04-01

    In higher plants, the numbers of cytoplasmic-distributed Golgi stacks differ based on function, age and cell type. It has not been clarified how the numbers are controlled, whether all the Golgi apparatus in a cell function equally and whether the increase in Golgi number is a result of the de novo formation from the endoplasmic reticulum (ER) or fission of pre-existing stacks. A tobacco prolyl 4-hydroxylase (NtP4H1.1), which is a cis-Golgi-localizing type II membrane protein, was tagged with a photoconvertible fluorescent protein, mKikGR (monomeric Kikume green red), and expressed in tobacco bright yellow 2 (BY-2) cells. Transformed cells were exposed to purple light to convert the fluorescence from green to red. A time-course analysis after the conversion revealed a progressive increase in green puncta and a decrease in the red puncta. From 3 to 6 h, we observed red, yellow and green fluorescent puncta corresponding to pre-existing Golgi; Golgi containing both pre-existing and newly synthesized protein; and newly synthesized Golgi. Analysis of the number and fluorescence of Golgi at different phases of the cell cycle suggested that an increase in Golgi number with both division and de novo synthesis occurred concomitantly with DNA replication. Investigation with different inhibitors suggested that the formation of new Golgi and the generation of Golgi containing both pre-existing and newly synthesized protein are mediated by different machineries. These results and modeling based on quantified results indicate that the Golgi apparatuses in tobacco BY-2 cells are not uniform and suggest that both de novo synthesis from the ER and Golgi division contribute almost equally to the increase in proliferating cells.

  11. Innate Lymphoid Cells Mediate Pulmonary Eosinophilic Inflammation, Airway Mucous Cell Metaplasia, and Type 2 Immunity in Mice Exposed to Ozone.

    Science.gov (United States)

    Kumagai, Kazuyoshi; Lewandowski, Ryan P; Jackson-Humbles, Daven N; Buglak, Nicholas; Li, Ning; White, Kaylin; Van Dyken, Steven J; Wagner, James G; Harkema, Jack R

    2017-08-01

    Exposure to elevated levels of ambient ozone in photochemical smog is associated with eosinophilic airway inflammation and nonatopic asthma in children. In the present study, we determined the role of innate lymphoid cells (ILCs) in the pathogenesis of ozone-induced nonatopic asthma by using lymphoid cell-sufficient C57BL/6 mice, ILC-sufficient Rag2 -/- mice (devoid of T and B cells), and ILC-deficient Rag2 -/- Il2rg -/- mice (depleted of all lymphoid cells including ILCs). Mice were exposed to 0 or 0.8 parts per million ozone for 1 day or 9 consecutive weekdays (4 hr/day). A single exposure to ozone caused neutrophilic inflammation, airway epithelial injury, and reparative DNA synthesis in all strains of mice, irrespective of the presence or absence of ILCs. In contrast, 9-day exposures induced eosinophilic inflammation and mucous cell metaplasia only in the lungs of ILC-sufficient mice. Repeated ozone exposures also elicited increased messenger RNA expression of transcripts associated with type 2 immunity and airway mucus production in ILC-sufficient mice. ILC-deficient mice repeatedly exposed to ozone had no pulmonary pathology or increased gene expression related to type 2 immunity. These results suggest a new paradigm for the biologic mechanisms underlying the development of a phenotype of childhood nonatopic asthma that has been linked to ambient ozone exposures.

  12. Human papillomavirus-exposed Langerhans cells are activated by stabilized Poly-I:C.

    Science.gov (United States)

    Da Silva, Diane M; Woodham, Andrew W; Rijkee, Laurie K; Skeate, Joseph G; Taylor, Julia R; Koopman, Maaike E; Brand, Heike E; Wong, Michael K; McKee, Greg M; Salazar, Andres M; Kast, W Martin

    2015-12-01

    Human papillomaviruses (HPV) establish persistent infections because of evolved immune evasion mechanisms, particularly HPV-mediated suppression of the immune functions of Langerhans cells (LC), the antigen presenting cells of the epithelium. Polyinosinic-polycytidilic acid (Poly-I:C) is broadly immunostimulatory with the ability to enhance APC expression of costimulatory molecules and inflammatory cytokines resulting in T cell activation. Here we investigated the activation of primary human LC derived from peripheral blood monocytes after exposure to HPV16 virus like particles followed by treatment with stabilized Poly-I:C compounds (s-Poly-I:C), and their subsequent induction of HPV16-specific T cells. Our results indicate that HPV16 particles alone were incapable of inducing LC activation as demonstrated by the lack of costimulatory molecules, inflammatory cytokines, chemokine-directed migration, and HPV16-specific CD8 + T cells in vitro . Conversely, s-Poly-I:C caused significant upregulation of costimulatory molecules and induction of chemokine-directed migration of LC that were pre-exposed to HPV16. In HLA-A*0201-positive donors, s-Poly-I:C treatment was able to induce CD8 + T cell immune responses against HPV16-derived peptides. Thus, s-Poly-I:C compounds are attractive for translation into therapeutics in which they could potentially mediate clearance of persistent HPV infection.

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

    Science.gov (United States)

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

    2015-07-01

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

  14. Gene expression signatures in peripheral blood cells from Japanese women exposed to environmental cadmium

    International Nuclear Information System (INIS)

    Dakeshita, Satoru; Kawai, Tomoko; Uemura, Hirokazu; Hiyoshi, Mineyoshi; Oguma, Etsuko; Horiguchi, Hyogo; Kayama, Fujio; Aoshima, Keiko; Shirahama, Satoshi; Rokutan, Kazuhito; Arisawa, Kokichi

    2009-01-01

    The objective of this study was to examine the effects of environmental cadmium (Cd) exposure on the gene expression profile of peripheral blood cells, using an original oligoDNA microarray. The study population consisted of 20 female residents in a Cd-polluted area (Cd-exposed group) and 20 female residents in a non-Cd-polluted area individually matched for age (control group). The mRNA levels in Cd-exposed subjects were compared with those in respective controls, using a microarray containing oligoDNA probes for 1867 genes. Median Cd concentrations in blood (3.55 μg/l) and urine (8.25 μg/g creatinine) from the Cd-exposed group were 2.4- and 1.9-times higher than those of the control group, respectively. Microarray analysis revealed that the Cd-exposed group significantly up-regulated 137 genes and down-regulated 80 genes, compared with the control group. The Ingenuity Pathway Analysis Application (IPA) revealed that differentially expressed genes were likely to modify oxidative stress and mitochondria-dependent apoptosis pathways. Among differentially expressed genes, the expression of five genes was positively correlated with Cd concentrations in blood or urine. Quantitative real-time PCR (RT-PCR) analysis validated the significant up-regulation of CASP9, TNFRSF1B, GPX3, HYOU1, SLC3A2, SLC19A1, SLC35A4 and ITGAL, and down-regulation of BCL2A1 and COX7B. After adjustment for differences in the background characteristics of the two groups, we finally identified seven Cd-responsive genes (CASP9, TNFRSF1B, GPX3, SLC3A2, ITGAL, BCL2A1, and COX7B), all of which constituted a network that controls oxidative stress response by IPA. These seven genes may be marker genes useful for the health risk assessment of chronic low level exposure to Cd

  15. Thioredoxin (Trxo1) interacts with proliferating cell nuclear antigen (PCNA) and its overexpression affects the growth of tobacco cell culture.

    Science.gov (United States)

    Calderón, Aingeru; Ortiz-Espín, Ana; Iglesias-Fernández, Raquel; Carbonero, Pilar; Pallardó, Federico Vicente; Sevilla, Francisca; Jiménez, Ana

    2017-04-01

    Thioredoxins (Trxs), key components of cellular redox regulation, act by controlling the redox status of many target proteins, and have been shown to play an essential role in cell survival and growth. The presence of a Trx system in the nucleus has received little attention in plants, and the nuclear targets of plant Trxs have not been conclusively identified. Thus, very little is known about the function of Trxs in this cellular compartment. Previously, we studied the intracellular localization of PsTrxo1 and confirmed its presence in mitochondria and, interestingly, in the nucleus under standard growth conditions. In investigating the nuclear function of PsTrxo1 we identified proliferating cellular nuclear antigen (PCNA) as a PsTrxo1 target by means of affinity chromatography techniques using purified nuclei from pea leaves. Such protein-protein interaction was corroborated by dot-blot and bimolecular fluorescence complementation (BiFC) assays, which showed that both proteins interact in the nucleus. Moreover, PsTrxo1 showed disulfide reductase activity on previously oxidized recombinant PCNA protein. In parallel, we studied the effects of PsTrxo1 overexpression on Tobacco Bright Yellow-2 (TBY-2) cell cultures. Microscopy and flow-cytometry analysis showed that PsTrxo1 overexpression increases the rate of cell proliferation in the transformed lines, with a higher percentage of the S phase of the cell cycle at the beginning of the cell culture (days 1 and 3) and at the G2/M phase after longer times of culture (day 9), coinciding with an upregulation of PCNA protein. Furthermore, in PsTrxo1 overexpressed cells there is a decrease in the total cellular glutathione content but maintained nuclear GSH accumulation, especially at the end of the culture, which is accompanied by a higher mitotic index, unlike non-overexpressing cells. These results suggest that Trxo1 is involved in the cell cycle progression of TBY-2 cultures, possibly through its link with cellular PCNA

  16. Membrane topology of Golgi-localized probable S-adenosylmethionine-dependent methyltransferase in tobacco (Nicotiana tabacum) BY-2 cells.

    Science.gov (United States)

    Liu, Jianping; Hayashi, Kyoko; Matsuoka, Ken

    2015-01-01

    S-adenosylmethionine (SAM)-dependent methyltransferases (MTases) transfer methyl groups to substrates. In this study, a novel putative tobacco SAM-MTase termed Golgi-localized methyl transferase 1 (GLMT1) has been characterized. GLMT1 is comprised of 611 amino acids with short N-terminal region, putative transmembrane region, and C-terminal SAM-MTase domain. Expression of monomeric red fluorescence protein (mRFP)-tagged protein in tobacco BY-2 cell indicated that GLMT1 is a Golgi-localized protein. Analysis of the membrane topology by protease digestion suggested that both C-terminal catalytic region and N-terminal region seem to be located to the cytosolic side of the Golgi apparatus. Therefore, GLMT1 might have a different function than the previously studied SAM-MTases in plants.

  17. Enhanced poly(3-hydroxybutyrate) production in transgenic tobacco BY-2 cells using engineered acetoacetyl-CoA reductase.

    Science.gov (United States)

    Yokoo, Toshinori; Matsumoto, Ken'ichiro; Ooba, Takashi; Morimoto, Kenjiro; Taguchi, Seiichi

    2015-01-01

    Highly active mutant of NADPH-dependent acetoacetyl-CoA reductase (PhaB) was expressed in Nicotiana tabacum cv. Bright Yellow-2 cultured cells to produce poly(3-hydroxybutyrate) [P(3HB)]. The mutated PhaB increased P(3HB) content by three-fold over the control, indicating that the mutant was a versatile tool for P(3HB) production. Additionally, the PhaB-catalyzed reaction was suggested to be a rate-limiting step of P(3HB) biosynthesis in tobacco BY-2 cells.

  18. Polyamine metabolism during the cell cycle of synchronized tobacco BY-2 cell line

    Czech Academy of Sciences Publication Activity Database

    Gemperlová, Lenka; Cvikrová, Milena; Fischerová, Lucie; Binarová, Pavla; Fischer, L.; Eder, Josef

    2009-01-01

    Roč. 47, č. 7 (2009), s. 584-591 ISSN 0981-9428 R&D Projects: GA AV ČR IAA500200719 Institutional research plan: CEZ:AV0Z50380511; CEZ:AV0Z50200510 Keywords : ADC * Cell cycle * DAO Subject RIV: EF - Botanics Impact factor: 2.485, year: 2009

  19. Cells exposed to nanosecond electrical pulses exhibit biomarkers of mechanical stress

    Science.gov (United States)

    Roth, Caleb C.; Barnes, Ronald A.; Ibey, Bennett L.; Beier, Hope T.; Moen, Erick K.; Glickman, Randolph D.

    2015-03-01

    Exposure of cells to very short (stressors on a cell, including electrical, electro-chemical, and mechanical stress. Thus, nsEP exposure is not a "clean" insult, making determination of the mechanism of nanoporation quite difficult. We hypothesize that nsEP exposure creates acoustic shock waves capable of causing nanoporation. Microarray analysis of primary adult human dermal fibroblasts (HDFa) exposed to nsEP, indicated several genes associated with mechanical stress were selectively upregulated 4 h post exposure. The idea that nanoporation is caused by external mechanical force from acoustic shock waves has, to our knowledge, not been investigated. This work will critically challenge the existing paradigm that nanoporation is caused solely by an electric-field driven event and could provide the basis for a plausible explanation for electroporation.

  20. Microarray analysis of gene expression in peripheral blood mononuclear cells from dioxin-exposed human subjects

    International Nuclear Information System (INIS)

    McHale, Cliona M.; Zhang, Luoping; Hubbard, Alan E.; Zhao, Xin; Baccarelli, Andrea; Pesatori, Angela C.; Smith, Martyn T.; Landi, Maria Teresa

    2007-01-01

    Tetrachlorodibenzo-p-dioxin (TCDD) is classified as a human carcinogen and exerts toxic effects on the skin (chloracne). Effects on reproductive, immunological, and endocrine systems have also been observed in animal models. TCDD acts through the aryl hydrocarbon receptor (AhR) pathway influencing largely unknown gene networks. An industrial accident in Seveso, Italy in 1976 exposed thousands of people to substantial quantities of TCDD. Twenty years after the exposure, this study examines global gene expression in the mononuclear cells of 26 Seveso female never smokers, with similar age, alcohol consumption, use of medications, and background plasma levels of 22 dioxin congeners unrelated to the Seveso accident. Plasma dioxin levels were still elevated in the exposed subjects. We performed analyses in two different comparison groups. The first included high-exposed study subjects compared with individuals with background TCDD levels (average plasma levels 99.4 and 6.7 ppt, respectively); the second compared subjects who developed chloracne after the accident, and those who did not develop this disease. Overall, we observed a modest alteration of gene expression based on dioxin levels or on chloracne status. In the comparison between high levels and background levels of TCDD, four histone genes were up-regulated and modified expression of HIST1H3H was confirmed by real-time PCR. In the comparison between chloracne case-control subjects, five hemoglobin genes were up-regulated. Pathway analysis revealed two major networks for each comparison, involving cell proliferation, apoptosis, immunological and hematological disease, and other pathways. Further examination of the role of these genes in dioxin induced-toxicity is warranted

  1. Proliferation of the Golgi apparatus in tobacco BY-2 cells during cell proliferation after release from the stationary phase of growth.

    Science.gov (United States)

    Abiodun, Moses; Matsuoka, Ken

    2013-08-01

    We have recently developed a new method aimed at mass photo-conversion of photo-convertible fluorescence protein (PFP) fluorescence in transformed tobacco BY-2 cells. Using this method we reported recently that the Golgi apparatus is generated by the de novo formation from ER and the division of pre-existing Golgi stacks with similar extents In this work we report that the proliferation of the Golgi apparatus in tobacco cells that enter the growing cycle from the non-dividing cycle is quite similar to that in rapidly growing cells and that de novo formation from the ER and division of pre-existing stacks seems to contribute almost equally to the proliferation.

  2. Genotoxic Changes to Rodent Cells Exposed in Vitro to Tungsten, Nickel, Cobalt and Iron

    Directory of Open Access Journals (Sweden)

    Stephanie Bardack

    2014-03-01

    Full Text Available Tungsten-based materials have been proposed as replacements for depleted uranium in armor-penetrating munitions and for lead in small-arms ammunition. A recent report demonstrated that a military-grade composition of tungsten, nickel, and cobalt induced a highly-aggressive, metastatic rhabdomyosarcoma when implanted into the leg muscle of laboratory rats to simulate a shrapnel wound. The early genetic changes occurring in response to embedded metal fragments are not known. In this study, we utilized two cultured rodent myoblast cell lines, exposed to soluble tungsten alloys and the individual metals comprising the alloys, to study the genotoxic effects. By profiling cell transcriptomes using microarray, we found slight, yet distinct and unique, gene expression changes in rat myoblast cells after 24 h metal exposure, and several genes were identified that correlate with impending adverse consequences of ongoing exposure to weapons-grade tungsten alloy. These changes were not as apparent in the mouse myoblast cell line. This indicates a potential species difference in the cellular response to tungsten alloy, a hypothesis supported by current findings with in vivo model systems. Studies examining genotoxic-associated gene expression changes in cells from longer exposure times are warranted.

  3. The effect of electronic cigarette and tobacco smoke exposure on COPD bronchial epithelial cell inflammatory responses

    Directory of Open Access Journals (Sweden)

    Higham A

    2018-03-01

    Full Text Available Andrew Higham,1,2 Declan Bostock,1 George Booth,2 Josiah V Dungwa,2 Dave Singh1,2 1Division of Infection, Immunity and Respiratory Medicine, School of Biological Sciences, Faculty of Biology, Medicine and Health, Manchester Academic Health Science Centre, The University of Manchester and University Hospital of South Manchester, NHS Foundation Trust, Manchester, UK; 2Medicines Evaluation Unit, University Hospital of South Manchester, Manchester, UK Background: Electronic cigarettes (e-cigs are used to help smoking cessation. However, these devices contain harmful chemicals, and there are safety concerns. We have investigated the effects of e-cigs on the inflammatory response and viability of COPD bronchial epithelial cells (BECs.Methods: BECs from COPD patients and controls were exposed to e-cig vapor extract (ECVE and the levels of interleukin (IL-6, C-X-C motif ligand 8 (CXCL8, and lactate dehydrogenase release were measured. We also examined the effect of ECVE pretreatment on polyinosinic:polycytidylic acid (poly I:C-stimulated cytokine release from BECs. Parallel experiments using Calu-3 cells were performed. Comparisons were made with cigarette smoke extract (CSE.Results: ECVE and CSE caused an increase in the release of IL-6 and CXCL8 from Calu-3 cells. ECVE only caused toxicity in BECs and Calu-3 cells. Furthermore, ECVE and CSE dampened poly I:C-stimulated C-X-C motif ligand 10 release from both cell culture models, reaching statistical significance for CSE at an optical density of 0.3.Conclusion: ECVE caused toxicity and reduced the antiviral response to poly I:C. This raises concerns over the safety of e-cig use. Keywords: e-cigs, epithelial cells, COPD, air–liquid interface, cigarette smoke

  4. Up-regulation of phosphoinositide metabolism in tobacco cells constitutively expressing the human type I inositol polyphosphate 5-phosphatase

    Science.gov (United States)

    Perera, Imara Y.; Love, John; Heilmann, Ingo; Thompson, William F.; Boss, Wendy F.; Brown, C. S. (Principal Investigator)

    2002-01-01

    To evaluate the impact of suppressing inositol 1,4,5-trisphosphate (InsP(3)) in plants, tobacco (Nicotiana tabacum) cells were transformed with the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), an enzyme which specifically hydrolyzes InsP(3). The transgenic cell lines showed a 12- to 25-fold increase in InsP 5-ptase activity in vitro and a 60% to 80% reduction in basal InsP(3) compared with wild-type cells. Stimulation with Mas-7, a synthetic analog of the wasp venom peptide mastoparan, resulted in an approximately 2-fold increase in InsP(3) in both wild-type and transgenic cells. However, even with stimulation, InsP(3) levels in the transgenic cells did not reach wild-type basal values, suggesting that InsP(3) signaling is compromised. Analysis of whole-cell lipids indicated that phosphatidylinositol 4,5-bisphosphate (PtdInsP(2)), the lipid precursor of InsP(3), was greatly reduced in the transgenic cells. In vitro assays of enzymes involved in PtdInsP(2) metabolism showed that the activity of the PtdInsP(2)-hydrolyzing enzyme phospholipase C was not significantly altered in the transgenic cells. In contrast, the activity of the plasma membrane PtdInsP 5 kinase was increased by approximately 3-fold in the transgenic cells. In vivo labeling studies revealed a greater incorporation of (32)P into PtdInsP(2) in the transgenic cells compared with the wild type, indicating that the rate of PtdInsP(2) synthesis was increased. These studies show that the constitutive expression of the human type I InsP 5-ptase in tobacco cells leads to an up-regulation of the phosphoinositide pathway and highlight the importance of PtdInsP(2) synthesis as a regulatory step in this system.

  5. In tobacco BY-2 cells xyloglucan oligosaccharides alter the expression of genes involved in cell wall metabolism, signalling, stress responses, cell division and transcriptional control.

    Science.gov (United States)

    González-Pérez, Lien; Perrotta, Lara; Acosta, Alexis; Orellana, Esteban; Spadafora, Natasha; Bruno, Leonardo; Bitonti, Beatrice M; Albani, Diego; Cabrera, Juan Carlos; Francis, Dennis; Rogers, Hilary J

    2014-10-01

    Xyloglucan oligosaccharides (XGOs) are breakdown products of XGs, the most abundant hemicelluloses of the primary cell walls of non-Poalean species. Treatment of cell cultures or whole plants with XGOs results in accelerated cell elongation and cell division, changes in primary root growth, and a stimulation of defence responses. They may therefore act as signalling molecules regulating plant growth and development. Previous work suggests an interaction with auxins and effects on cell wall loosening, however their mode of action is not fully understood. The effect of an XGO extract from tamarind (Tamarindus indica) on global gene expression was therefore investigated in tobacco BY-2 cells using microarrays. Over 500 genes were differentially regulated with similar numbers and functional classes of genes up- and down-regulated, indicating a complex interaction with the cellular machinery. Up-regulation of a putative XG endotransglycosylase/hydrolase-related (XTH) gene supports the mechanism of XGO action through cell wall loosening. Differential expression of defence-related genes supports a role for XGOs as elicitors. Changes in the expression of genes related to mitotic control and differentiation also support previous work showing that XGOs are mitotic inducers. XGOs also affected expression of several receptor-like kinase genes and transcription factors. Hence, XGOs have significant effects on expression of genes related to cell wall metabolism, signalling, stress responses, cell division and transcriptional control.

  6. [Effect of inhibitors serine/threonine protein kinases and protein phosphatases on mitosis progression of synchronized tobacco by-2 cells].

    Science.gov (United States)

    Sheremet, Ia A; Emets, A I; Azmi, A; Vissenberg, K; Verbelen, J-P; Blium, Ia B

    2012-01-01

    In order to investigate the role of various serine/ threonine protein kinases and protein phosphatases in the regulation of mitosis progression in plant cells the influence of cyclin-dependent (olomoucine) and Ca2+ -calmodulin-dependent (W7) protein kinases inhibitors, as well as protein kinase C inhibitors (H7 and staurosporine) and protein phosphatases inhibitor (okadaic acid) on mitosis progression in synchronized tobacco BY-2 cells has been studied. It was found that BY-2 culture treatment with inhibitors of cyclin dependent protein kinases and protein kinase C causes prophase delay, reduces the mitotic index and displaces of mitotic peak as compare with control cells. Inhibition of Ca2+ -calmodulin dependent protein kinases enhances the cell entry into prophase and delays their exit from mitosis. Meanwhile inhibition of serine/threonine protein phosphatases insignificantly enhances of synchronized BY-2 cells entering into all phases of mitosis.

  7. Incorporation of uridine-H3 into healthy and tobacco necrosis virus-infected mesophyll cells of Chenopodium amaranticolor

    International Nuclear Information System (INIS)

    Faccioli, G.; Rubies-Autonel, C.

    1975-01-01

    Tritiated uridine was selectively incorporated into the nucleus, nucleolus and cytoplasm of actinomycin D-treated Chenopodium amaranticolor cells locally infected with a strain of tobacco necrosis virus (TNV), 3 days after inoculation. Healthy cells did not show such an incorporation. Chloroplasts, in both types of cells, were free of label. Treatment with pancreatic ribonuclease removed the label completely in the majority of nuclei and nucleoli of infected cells. Since infectivity tests showed that AMD treatment increased virus multiplication by 10-12%, it is conceivable to think that the incorporation observed was due to virus synthesis. Preliminary infectivity experiments also showed that treatment of the cells with cycloheximide inhibited virus multiplication up to 80%, while chloramphenicol increased such multiplication. Our results lead to the conclusion that nucleus, nucleolus and cytoplasm but not chloroplasts are the sites involved in the synthesis of TNV. (orig.) [de

  8. EVALUATION OF CELL CYCLE OF Aspergillus nidulans EXPOSED TO THE EXTRACT OF Copaifera officinalis L PLANT

    Directory of Open Access Journals (Sweden)

    Simone Jurema Ruggeri Chiuchetta, Uériton Dias de Oliveira e Josy Fraccaro de Marins

    2006-12-01

    Full Text Available The oil extracted from the Copaifera officinalis L plant has been used in popular medicine to the treatment of several diseases, like cancer. In eukaryotic cells, the process of cellular proliferation follows a standard cycle, named cellular cycle. The transformation of a normal cell in a malignant one requires several steps, in which genes that control normal cellular division or cellular death are modified. Aspergillus nidulans fungus is an excellent system for the study of the cellular differentiation. Its asexual cycle results in the formation of conidia, which are disposed like chains, constituting a structure named conidiophore. This structure consists in an aerial hifae, multinucleate vesicle and uninucleate cells. Current research evaluated the capacity of the C. officinalis L plant extract in promoting alterations in the cellular cycle of A. nidulans diploid strains, by observing macroscopic and microscopic alterations in cellular growth of this fungus. Results shown that no macroscopic alterations were observed in cellular growth of strains exposed to the extract, however, microscopic alterations of conidiophore have been observed in the different extract concentrations analyzed. In this way, the study of the action of C. officinalis L plant extract becomes important considering the fact that this substance is capable to promote alterations in cellular cycle of eukaryotic cells.

  9. Global gene expression profiling in human lung cells exposed to cobalt

    Energy Technology Data Exchange (ETDEWEB)

    Malard, V.; Berenguer, F.; Prat, O.; Ruat, S.; Steinmetz, G.; Quemeneur, E. [CEA VALRHO, Serv Biochim and Toxicol Nucl, DSV, iBEB, F-30207 Bagnols Sur Ceze (France)

    2007-06-06

    It has been estimated that more than 1 million workers in the United States are exposed to cobalt. Occupational exposure to {sup 59}Co occurs mainly via inhalation and leads to various lung diseases. Cobalt is classified by the IARC as a possible human carcinogen (group 2B). Although there is evidence for in vivo and in vitro toxicity, the mechanisms of cobalt-induced lung toxicity are not fully known. The purpose of this work was to identify potential signatures of acute cobalt exposure using a toxico-genomic approach. Data analysis focused on some cellular processes and protein targets that are thought to be relevant for carcinogenesis, transport and bio-marker research. Results: A time course transcriptome analysis was performed on A549 human pulmonary cells, leading to the identification of 85 genes which are repressed or induced in response to soluble 59 Co. A group of 29 of these genes, representing the main biological functions, was assessed by quantitative RT-PCR. The expression profiles of six of them were then tested by quantitative RT-PCR in a time-dependent manner and three modulations were confirmed by Western blotting. The 85 modulated genes include potential cobalt carriers (FBXL2, ZNT1, SLC12A5), tumor suppressors or transcription factors (MAZ, DLG1, MYC, AXL) and genes linked to the stress response (UBC, HSPCB, BN1P3L). We also identified nine genes coding for secreted proteins as candidates for bio-marker research. Of those, T1MP2 was found to be down-regulated and this modulation was confirmed, in a dose-dependent manner, at protein level in the supernatant of exposed cells. Conclusion: Most of these genes have never been described as related to cobalt stress and provide original hypotheses for further study of the effects of this metal ion on human lung epithelial cells. A putative bio-marker of cobalt toxicity was identified. (authors)

  10. Dose and temporal effects on gene expression profiles of urothelial cells from rats exposed to diuron

    International Nuclear Information System (INIS)

    Ihlaseh-Catalano, Shadia M.; Bailey, Kathryn A.; Cardoso, Ana Paula F.; Ren, Hongzu; Fry, Rebecca C.; Camargo, João Lauro V.de; Wolf, Douglas C.

    2014-01-01

    Diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea) is a substituted urea herbicide that at high dietary levels (2500 ppm) induces rat urinary bladder hyperplasia after 20 weeks of exposure and neoplasia after 2 years. The effects on the urothelium after short-term exposure have not been described. The present 7-day study evaluated the dose-dependency of urothelial alterations in the urinary bladder using light microscopy, scanning electron microscopy, and genome-wide transcriptional profiling. Male Wistar rats were fed 0, 125, 500, 2500 ppm diuron for 7 days. The urinary bladder and isolated urothelial cells of these animals were processed for microscopic examination and gene expression profiling, respectively. No significant treatment-related morphologic effects were observed. The number of differentially expressed genes (DEGs) in the exposed groups increased with diuron levels. Diuron-altered genes involved in cell-to-cell interactions and tissue organization were identified in all treatment groups. After 7 days of diuron exposure, transcriptional responses were observed in the urothelium in the absence of clear morphologic changes. These morphological findings are different from those observed in a previous study in which 20 weeks of diuron exposure was associated with simple hyperplasia secondary to the persistent cytotoxicity and necrosis associated with continuous cellular regeneration. Comparison of the gene expression profiles of rats exposed to the 2500 ppm carcinogenic diuron dose for 7 days versus 20 weeks revealed few similarities between these two time points at the gene or pathway level. Taken together, these data provide insight into the dose- and temporal-dependent morphological and transcriptional changes associated with diuron exposure that may lead to the development of tumors in the rat urinary bladder

  11. Dose and temporal effects on gene expression profiles of urothelial cells from rats exposed to diuron.

    Science.gov (United States)

    Ihlaseh-Catalano, Shadia M; Bailey, Kathryn A; Cardoso, Ana Paula F; Ren, Hongzu; Fry, Rebecca C; de Camargo, João Lauro V; Wolf, Douglas C

    2014-11-05

    Diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea) is a substituted urea herbicide that at high dietary levels (2500 ppm) induces rat urinary bladder hyperplasia after 20 weeks of exposure and neoplasia after 2 years. The effects on the urothelium after short-term exposure have not been described. The present 7-day study evaluated the dose-dependency of urothelial alterations in the urinary bladder using light microscopy, scanning electron microscopy, and genome-wide transcriptional profiling. Male Wistar rats were fed 0, 125, 500, 2500 ppm diuron for 7 days. The urinary bladder and isolated urothelial cells of these animals were processed for microscopic examination and gene expression profiling, respectively. No significant treatment-related morphologic effects were observed. The number of differentially expressed genes (DEGs) in the exposed groups increased with diuron levels. Diuron-altered genes involved in cell-to-cell interactions and tissue organization were identified in all treatment groups. After 7 days of diuron exposure, transcriptional responses were observed in the urothelium in the absence of clear morphologic changes. These morphological findings are different from those observed in a previous study in which 20 weeks of diuron exposure was associated with simple hyperplasia secondary to the persistent cytotoxicity and necrosis associated with continuous cellular regeneration. Comparison of the gene expression profiles of rats exposed to the 2500 ppm carcinogenic diuron dose for 7 days versus 20 weeks revealed few similarities between these two time points at the gene or pathway level. Taken together, these data provide insight into the dose- and temporal-dependent morphological and transcriptional changes associated with diuron exposure that may lead to the development of tumors in the rat urinary bladder. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  12. The ADMA/DDAH/NO pathway in human vein endothelial cells exposed to arsenite.

    Science.gov (United States)

    Osorio-Yáñez, Citlalli; Chin-Chan, Miguel; Sánchez-Peña, Luz C; Atzatzi-Aguilar, Octavio G; Olivares-Reyes, Jesus A; Segovia, José; Del Razo, Luz M

    2017-08-01

    Inorganic arsenic (iAs) exposure is related to cardiovascular disease, which is characterized by endothelial dysfunction and nitric oxide (NO) depletion. The mechanisms underlying NO depletion as related to iAs exposure are not fully understood. The endogenous inhibitor of nitric oxide synthase, asymmetric dimethylarginine (ADMA), might be a molecular target of iAs. ADMA concentrations are regulated by proteins involved in its synthesis (arginine methyl transferase 1 [PRMT-1]) and degradation (dimethylarginine dimethylaminohydrolase [DDAH]). Both, ADMA and NO are susceptible to oxidative stress. We aimed to determine the ADMA/DDAH/NO pathway in human vein endothelial cells (HUVEC-CS) exposed to arsenite. We exposed HUVEC-CS cells to 1, 2.5 and 5μM of arsenite for 24h. We proved that arsenite at 5μM was able to decrease NO levels with an associated increase in ADMA and depletion of l-arginine in HUVEC-CS cells. We also found a decrease in DDAH-1 protein expression with 5μM of arsenite compared to the control group. However, we did not observe significant differences in PRMT-1 protein expression at any of the concentrations of arsenite employed. Finally, arsenite (2.5 and 5μM) increased NADPH oxidase 4 protein levels compared with the control group. We conclude that ADMA, l-arginine and DDAH are involved in NO depletion produced by arsenite, and that the mechanism is related to oxidative stress. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Oral tongue squamous cell carcinoma (OTSCC): alcohol and tobacco consumption versus non-consumption. A study in a Portuguese population.

    Science.gov (United States)

    Albuquerque, Rui; López-López, José; Marí-Roig, Antonio; Jané-Salas, Enric; Roselló-Llabrés, Xavier; Santos, Jorge Rosa

    2011-01-01

    There has been an increase in the incidence of carcinoma of the tongue, particularly among alcohol and tobacco non-users. However, the number of studies that would allow a better understanding of etiological factors and clinical features, particularly in the Portuguese population, is very limited. This study was based on patients with squamous cell carcinoma of the anterior two thirds of the tongue that were treated at the Department of Head and Neck Surgery of the "Instituto Portugues de Oncologia de Lisboa - Francisco Gentil" (IPOLFG) in Lisbon, Portugal, between January 1, 2001 and December 31, 2009. The patients were divided in alcohol and tobacco users and non-users in order to evaluate the differences between these 2 groups based on gender, age, tumor location, denture use, and tumor size, metastasis and stage. Of the 354 cases, 208 were users and 146 were non-users. The main location in both groups was the lateral border of the tongue. Denture use showed no significant effect in both study groups. It was possible to conclude that patients who did not drink or smoke were older and presented with smaller tumor size, lower incidence of ganglion metastasis and lower tumor stage compared with alcohol and tobacco users.

  14. Ultrastructural analysis of oral exfoliated epithelial cells of tobacco smokers and betel nut chewers: A scanning electron microscopy study.

    Science.gov (United States)

    Khan, Sameera Shamim; Shreedhar, Balasundari; Kamboj, Mala

    2016-01-01

    The study was undertaken to correlate epithelial surface pattern changes of oral exfoliated cells of tobacco smokers and betel nut chewers and also to compare them with patients of oral squamous cell carcinoma (OSCC) and healthy individuals. In this cross-sectional study, a total of fifty persons were included in the study, out of which thirty formed the study group (15 each tobacco smokers and betel nut chewers) and twenty formed the control group (ten each of OSCC patients - positive control and ten normal buccal mucosa - negative control). Their oral exfoliated cells were scraped, fixed, and studied under scanning electron microscope (SEM). The statistical analysis was determined using ANOVA, Tukey honestly significant difference, Chi-square test, and statistical SPASS software, P betel nut chewers compared to normal oral mucosa have been tabulated. In normal oral mucosa, cell surface morphology depends on the state of keratinization of the tissue. Thus, it could prove helpful in detecting any carcinomatous change at its incipient stage and also give an insight into the ultra-structural details of cellular differentiations in epithelial tissues.

  15. The Bioeffects Resulting from Prokaryotic Cells and Yeast Being Exposed to an 18 GHz Electromagnetic Field.

    Directory of Open Access Journals (Sweden)

    The Hong Phong Nguyen

    Full Text Available The mechanisms by which various biological effects are triggered by exposure to an electromagnetic field are not fully understood and have been the subject of debate. Here, the effects of exposing typical representatives of the major microbial taxa to an 18 GHz microwave electromagnetic field (EMFwere studied. It appeared that the EMF exposure induced cell permeabilisation in all of the bacteria and yeast studied, while the cells remained viable (94% throughout the exposure, independent of the differences in cell membrane fatty acid and phospholipid composition. The resulting cell permeabilisation was confirmed by detection of the uptake of propidium iodine and 23 nm fluorescent silica nanospheres using transmission electron microscopy (TEM and confocal laser scanning microscopy (CLSM. Upon EMF exposure, the bacterial cell membranes are believed to become permeable through quasi-endocytosis processes. The dosimetry analysis revealed that the EMF threshold level required to induce the uptake of the large (46 nm nanopsheres was between three and six EMF doses, with a specific absorption rate (SAR of 3 kW/kg and 5 kW/kg per exposure, respectively, depending on the bacterial taxa being studied. It is suggested that the taxonomic affiliation and lipid composition (e.g. the presence of phosphatidyl-glycerol and/or pentadecanoic fatty acid may affect the extent of uptake of the large nanospheres (46 nm. Multiple 18 GHz EMF exposures over a one-hour period induced periodic anomalous increases in the cell growth behavior of two Staphylococcus aureus strains, namely ATCC 25923 and CIP 65.8T.

  16. Tobacco Addiction

    Science.gov (United States)

    ... and lighters—anything that you connect with your smoking habit. Get rid of all old chewing tobacco containers ... nicotine addiction and more to do with the habit of smoking or using chewing tobacco. Some people gain weight ...

  17. Cytogenetic dose-response and adaptive response in cells of ungulate species exposed to ionizing radiation

    International Nuclear Information System (INIS)

    Ulsh, B.A.; Miller, S.M.; Mallory, F.F.; Mitchel, R.E.J.; Morrison, D.P.; Boreham, D.R.

    2004-01-01

    In the studies reported here, the micronucleus assay, a common cytogenetic technique, was used to examine the dose-responses in fibroblasts from three ungulate species (white-tailed deer, woodland caribou, and Indian muntjac) exposed to high doses of ionizing radiation (1-4 Gy of 60 Co gamma radiation). This assay was also used to examine the effects of exposure to low doses (1-100 mGy) typical of what these species experience in a year from natural and anthropogenic environmental sources. An adaptive response, defined as the induction of resistance to a stressor by a prior exposure to a small 'adapting' stress, was observed after exposure to low doses. This work indicates that very small doses are protective for the endpoint examined. The same level of protection was seen at all adapting doses, including 1 radiation track per cell, the lowest possible cellular dose. These results are consistent with other studies in a wide variety of organisms that demonstrate a protective effect of low doses at both cellular and whole-organism levels. This implies that environmental regulations predicated on the idea that even the smallest dose of radiation carries a quantifiable risk of direct adverse consequences to the exposed organism require further examination. Cytogenetic assays provide affordable and feasible biological effects-based alternatives that are more biologically relevant than traditional contaminant concentration-based radioecological risk assessment

  18. Tobacco Industry Interference in the WHO European Region

    DEFF Research Database (Denmark)

    Olsen, Céline E J L Brassart

    2012-01-01

    WHO selected tobacco industry interference as the theme of the 2012 World No Tobacco Day, recognizing the serious danger the tobacco industry poses to public health and the need to expose and counter the industry’s increasing attempts to undermine the WHO Framework Convention on Tobacco Control...

  19. Cellular processes involved in human epidermal cells exposed to extremely low frequency electric fields.

    Science.gov (United States)

    Collard, J-F; Hinsenkamp, M

    2015-05-01

    We observed on different tissues and organisms a biological response after exposure to pulsed low frequency and low amplitude electric or electromagnetic fields but the precise mechanism of cell response remains unknown. The aim of this publication is to understand, using bioinformatics, the biological relevance of processes involved in the modification of gene expression. The list of genes analyzed was obtained after microarray protocol realized on cultures of human epidermal explants growing on deepidermized human skin exposed to a pulsed low frequency electric field. The directed acyclic graph on a WebGestalt Gene Ontology module shows six categories under the biological process root: "biological regulation", "cellular process", "cell proliferation", "death", "metabolic process" and "response to stimulus". Enriched derived categories are coherent with the type of in vitro culture, the stimulation protocol or with the previous results showing a decrease of cell proliferation and an increase of differentiation. The Kegg module on WebGestalt has highlighted "cell cycle" and "p53 signaling pathway" as significantly involved. The Kegg website brings out interactions between FoxO, MAPK, JNK, p53, p38, PI3K/Akt, Wnt, mTor or NF-KappaB. Some genes expressed by the stimulation are known to have an exclusive function on these pathways. Analyses performed with Pathway Studio linked cell proliferation, cell differentiation, apoptosis, cell cycle, mitosis, cell death etc. with our microarrays results. Medline citation generated by the software and the fold change variation confirms a diminution of the proliferation, activation of the differentiation and a less well-defined role of apoptosis or wound healing. Wnt and DKK functional classes, DKK1, MACF1, ATF3, MME, TXNRD1, and BMP-2 genes proposed in previous publications after a manual analysis are also highlighted with other genes after Pathway Studio automatic procedure. Finally, an analysis conducted on a list of genes

  20. Immunotropic potency of microwave fields: preliminary studies on immunocompetent cells exposed in vitro

    International Nuclear Information System (INIS)

    Stankiewicz, W.; Dabrowski, M.P.; Sobiczewska, E.; Kubacki, R.; Szmigielski, S.

    2006-01-01

    Exposure in radiofrequency (RF) and microwave (MW) fields can influence the function of the immune system, but the data available on the immunotropic potency of RF/MW radiation are still full of uncertainties and controversies. In the available literature there exist no reports on complex assessment of function and responsiveness of the immune system. All investigations have been aimed to evaluate selected, fragmentary reaction of the system and/or functional response of immunocompetent cells in RF/MW-exposed subjects. However, at the present state of knowledge it is not possible to conclude about the possible immunotropic potencies of RF/MW radiation. The undisturbed defensive, tolerogenic, and proregenerative activities of the immune system are commonly recognised as one of the most important homeostatic functions of the organism. Thus, basic immunoregulatory activities which can be observed and precisely quantified in microcultures of immune cells separated from the human blood, represent a unique and objective model for the investigation of possible immunotropic effects of electromagnetic fields (EMFs). To determine the potential immunomodulatory influences of EMFs, the immunotropic effects of pulse modulated microwave (1300 MHz) were investigated in the cultures of blood mononuclear cells from sixteen healthy donors

  1. Oxidative stress response in neural stem cells exposed to different superparamagnetic iron oxide nanoparticles.

    Science.gov (United States)

    Pongrac, Igor M; Pavičić, Ivan; Milić, Mirta; Brkić Ahmed, Lada; Babič, Michal; Horák, Daniel; Vinković Vrček, Ivana; Gajović, Srećko

    2016-01-01

    Biocompatibility, safety, and risk assessments of superparamagnetic iron oxide nanoparticles (SPIONs) are of the highest priority in researching their application in biomedicine. One improvement in the biological properties of SPIONs may be achieved by different functionalization and surface modifications. This study aims to investigate how a different surface functionalization of SPIONs - uncoated, coated with d-mannose, or coated with poly-l-lysine - affects biocompatibility. We sought to investigate murine neural stem cells (NSCs) as important model system for regenerative medicine. To reveal the possible mechanism of toxicity of SPIONs on NSCs, levels of reactive oxygen species, intracellular glutathione, mitochondrial membrane potential, cell-membrane potential, DNA damage, and activities of SOD and GPx were examined. Even in cases where reactive oxygen species levels were significantly lowered in NSCs exposed to SPIONs, we found depleted intracellular glutathione levels, altered activities of SOD and GPx, hyperpolarization of the mitochondrial membrane, dissipated cell-membrane potential, and increased DNA damage, irrespective of the surface coating applied for SPION stabilization. Although surface coating should prevent the toxic effects of SPIONs, our results showed that all of the tested SPION types affected the NSCs similarly, indicating that mitochondrial homeostasis is their major cellular target. Despite the claimed biomedical benefits of SPIONs, the refined determination of their effects on various cellular functions presented in this work highlights the need for further safety evaluations. This investigation helps to fill the knowledge gaps on the criteria that should be considered in evaluating the biocompatibility and safety of novel nanoparticles.

  2. Selective decreases of nicotinic acetylcholine receptors in PC12 cells exposed to fluoride

    International Nuclear Information System (INIS)

    Chen Jia; Shan, K.-R.; Long, Y.-G.; Wang, Y.-N.; Nordberg, Agneta; Guan, Z.-Z.

    2003-01-01

    In an attempt to elucidate the mechanism by which excessive fluoride damages the central nervous system, the effects of exposure of PC12 cells to different concentrations of fluoride for 48 h on nicotinic acetylcholine receptors (nAChRs) were characterized here. Significant reductions in the number of binding sites for both [ 3 H]epibatidine and [ 125 I]α-bungarotoxin, as well as a significant decrease in the B max value for the high-affinity of epibatidine binding site were observed in PC12 cells subjected to high levels of fluoride. On the protein level, the α3 and α7 subunits of nAChRs were also significantly decreased in the cells exposed to high concentrations of fluoride. In contrast, such exposure had no significant effect on the level of the β2 subunit. These findings suggest that selective decreases in the number of nAChRs may play an important role in the mechanism(s) by which fluoride causes dysfunction of the central nervous system

  3. Sildenafil Prevents Apoptosis of Human First-Trimester Trophoblast Cells Exposed to Oxidative Stress

    Science.gov (United States)

    Bolnick, Jay M.; Kilburn, Brian A.; Bolnick, Alan D.; Diamond, Michael P.; Singh, Manvinder; Hertz, Michael; Dai, Jing

    2015-01-01

    Human first-trimester trophoblast cells proliferate at low O2, but survival is compromised by oxidative stress, leading to uteroplacental insufficiency. The vasoactive drug, sildenafil citrate (Viagra, Sigma, St Louis, Missouri), has proven useful in reducing adverse pregnancy outcomes. An important biological function of this pharmaceutical is its action as an inhibitor of cyclic guanosine monophosphate (cGMP) phosphodiesterase type 5 activity, which suggests that it could have beneficial effects on trophoblast survival. To investigate whether sildenafil can prevent trophoblast cell death, human first-trimester villous explants and the HTR-8/SVneo cytotrophoblast cell line were exposed to hypoxia and reoxygenation (H/R) to generate oxidative stress, which induces apoptosis. Apoptosis was optimally inhibited during H/R by 350 ng/mL sildenafil. Sildenafil-mediated survival was reversed by l-NG-nitro-l-arginine methyl ester hydrochloride or cGMP antagonist, indicating a dependence on both nitric oxide (NO) and cGMP. Indeed, either a cGMP agonist or an NO generator was cytoprotective independent of sildenafil. These findings suggest a novel intervention route for patients with recurrent pregnancy loss or obstetrical placental disorders. PMID:25431453

  4. DNA damage and the bystander response in tumor and normal cells exposed to X-rays.

    Science.gov (United States)

    Subhashree, M; Venkateswarlu, R; Karthik, K; Shangamithra, V; Venkatachalam, P

    2017-09-01

    Monolayer and suspension cultures of tumor (BMG-1, CCRF-CEM), normal (AG1522, HADF, lymphocytes) and ATM-mutant (GM4405) human cells were exposed to X-rays at doses used in radiotherapy (high dose and high dose-rate) or radiological imaging (low dose and low dose-rate). Radiation-induced DNA damage, its persistence, and possible bystander effects were evaluated, based on DNA damage markers (γ-H2AX, p53 ser15 ) and cell-cycle-specific cyclins (cyclin B1 and cyclin D1). Dose-dependent DNA damage and a dose-independent bystander response were seen after exposure to high dose and high dose-rate radiation. The level of induced damage (expression of p53 ser15 , γ-H2AX) depended on ATM status. However, low dose and dose-rate exposures neither increased expression of marker proteins nor induced a bystander response, except in the CCRF-CEM cells. Bystander effects after high-dose irradiation may contribute to stochastic and deterministic effects. Precautions to protect unexposed regions or to inhibit transmission of DNA damage signaling might reduce radiation risks. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Expression profile of cell cycle genes in the fish CATLA CATLA (Ham.) exposed to gamma radiation

    International Nuclear Information System (INIS)

    Anbumani, S.; Mohankumar Mary, N.

    2012-01-01

    The International Commission on Radiological Protection (ICRP) emphasized the need to protect non-human biota from the potential effects of ionizing radiation and proposed to include molecular effects such as DNA damage as endpoints. Molecular effects of ionizing radiation exposure in representative non-humans are largely unexplored and sufficient data is not available in fishes. Gene expression is a fast and sensitive end point in detecting the molecular cues as a result of ionizing radiation exposure in a wide variety of aquatic organisms under suspected environmental contamination. Exposure to ionizing radiation transiently alters gene expression profiles as cells regulate certain genes to protect cellular structures and repair damage. The present study focused on genes like Gadd45á, Cdk1 and Bcl-2 in DNA damage repair and cell cycle machinery and its implication as molecular markers of radiation exposure. This study is first of its kind showing the in vivo expression profile of cell cycle genes in fish exposed to gamma radiation. Although this preliminary investigation points to certain molecular markers of ionizing radiation, elaborate studies with various doses and dose-rates are required before these markers find application as prospective molecular markers in aquatic radiation biodosimetry

  6. Displacement of the mitotic apparatuses by centrifugation reveals cortical actin organization during cytokinesis in cultured tobacco BY-2 cells.

    Science.gov (United States)

    Arima, Kengo; Tamaoki, Daisuke; Mineyuki, Yoshinobu; Yasuhara, Hiroki; Nakai, Tomonori; Shimmen, Teruo; Yoshihisa, Tohru; Sonobe, Seiji

    2018-06-19

    In plant cytokinesis, actin is thought to be crucial in cell plate guidance to the cortical division zone (CDZ), but its organization and function are not fully understood. To elucidate actin organization during cytokinesis, we employed an experimental system, in which the mitotic apparatus is displaced and separated from the CDZ by centrifugation and observed using a global-local live imaging microscope that enabled us to record behavior of actin filaments in the CDZ and the whole cell division process in parallel. In this system, returning movement of the cytokinetic apparatus in cultured-tobacco BY-2 cells occurs, and there is an advantage to observe actin organization clearly during the cytokinetic phase because more space was available between the CDZ and the distantly formed phragmoplast. Actin cables were clearly observed between the CDZ and the phragmoplast in BY-2 cells expressing GFP-fimbrin after centrifugation. Both the CDZ and the edge of the expanding phragmoplast had actin bulges. Using live-cell imaging including the global-local live imaging microscopy, we found actin filaments started to accumulate at the actin-depleted zone when cell plate expansion started even in the cell whose cell plate failed to reach the CDZ. These results suggest that specific accumulation of actin filaments at the CDZ and the appearance of actin cables between the CDZ and the phragmoplast during cell plate formation play important roles in the guidance of cell plate edges to the CDZ.

  7. Anti-inflammatory activity of the basolateral fraction of Caco-2 cells exposed to a rosemary supercritical extract

    NARCIS (Netherlands)

    Arranz, E.; Mes, J.J.; Wichers, H.J.; Jaime, L.; Reglero, G.; Santoyo, S.

    2015-01-01

    The anti-inflammatory activity of the basolateral fraction of Caco-2 cells exposed to a rosemary supercritical extract was examined. Uptake of rosemary extract fractions was tested on Caco-2 cell monolayers (2–12 h incubation times) and the quantification of carnosic acid and carnosol was performed

  8. Differential Signaling and Sugar Exchanges in Response to Avirulent Pathogen- and Symbiont-Derived Molecules in Tobacco Cells

    Directory of Open Access Journals (Sweden)

    Carole Pfister

    2017-11-01

    Full Text Available Plants interact with microbes whose ultimate aim is to exploit plant carbohydrates for their reproduction. Plant–microbe interactions (PMIs are classified according to the nature of their trophic exchanges: while mutualistic microbes trade nutrients with plants, pathogens unilaterally divert carbohydrates. The early responses following microbe recognition and the subsequent control of plant sugar distribution are still poorly understood. To further decipher PMI functionality, we used tobacco cells treated with microbial molecules mimicking pathogenic or mutualistic PMIs, namely cryptogein, a defense elicitor, and chitotetrasaccharide (CO4, which is secreted by mycorrhizal fungi. CO4 was perceived by tobacco cells and triggered widespread transient signaling components such as a sharp cytosolic Ca2+ elevation, NtrbohD-dependent H2O2 production, and MAP kinase activation. These CO4-induced events differed from those induced by cryptogein, i.e., sustained events leading to cell death. Furthermore, cryptogein treatment inhibited glucose and sucrose uptake but not fructose uptake, and promoted the expression of NtSUT and NtSWEET sugar transporters, whereas CO4 had no effect on sugar uptake and only a slight effect on NtSWEET2B expression. Our results suggest that microbial molecules induce different signaling responses that reflect microbial lifestyle and the subsequent outcome of the interaction.

  9. Human Invariant Natural Killer T Cells Respond to Antigen-Presenting Cells Exposed to Lipids from Olea europaea Pollen.

    Science.gov (United States)

    Abos Gracia, Beatriz; López Relaño, Juan; Revilla, Ana; Castro, Lourdes; Villalba, Mayte; Martín Adrados, Beatriz; Regueiro, Jose Ramon; Fernández-Malavé, Edgar; Martínez Naves, Eduardo; Gómez Del Moral, Manuel

    2017-01-01

    Allergic sensitization might be influenced by the lipids present in allergens, which can be recognized by natural killer T (NKT) cells on antigen-presenting cells (APCs). The aim of this study was to analyze the effect of olive pollen lipids in human APCs, including monocytes as well as monocyte-derived macrophages (Mϕ) and dendritic cells (DCs). Lipids were extracted from olive (Olea europaea) pollen grains. Invariant (i)NKT cells, monocytes, Mϕ, and DCs were obtained from buffy coats of healthy blood donors, and their cell phenotype was determined by flow cytometry. iNKT cytotoxicity was measured using a lactate dehydrogenase assay. Gene expression of CD1A and CD1D was performed by RT-PCR, and the production of IL-6, IL-10, IL-12, and TNF-α cytokines by monocytes, Mϕ, and DCs was measured by ELISA. Our results showed that monocytes and monocyte-derived Mϕ treated with olive pollen lipids strongly activate iNKT cells. We observed several phenotypic modifications in the APCs upon exposure to pollen-derived lipids. Both Mϕ and monocytes treated with olive pollen lipids showed an increase in CD1D gene expression, whereas upregulation of cell surface CD1d protein occurred only in Mϕ. Furthermore, DCs differentiated in the presence of human serum enhance their surface CD1d expression when exposed to olive pollen lipids. Finally, olive pollen lipids were able to stimulate the production of IL-6 but downregulated the production of lipopolysaccharide- induced IL-10 by Mϕ. Olive pollen lipids alter the phenotype of monocytes, Mϕ, and DCs, resulting in the activation of NKT cells, which have the potential to influence allergic immune responses. © 2017 S. Karger AG, Basel.

  10. Phosphorylation of mitogen-activated protein kinase (MAPK) is required for cytokinesis and progression of cell cycle in tobacco BY-2 cells.

    Science.gov (United States)

    Ma, Zhaowu; Yu, Guanghui

    2010-02-15

    The role of mitogen-activated protein kinase (MAPK) in plant cytokinesis remains largely uncharacterized. To elucidate its role, tobacco Bright Yellow-2 (BY-2) cells have been synchronized using a two-step procedure, and the different phases of the cell cycle identified by Histone 4 gene expression and the mitotic index. MAPK expression was analyzed by semi-quantitative (SQ) RT-PCR and protein gel blot analysis for phosphorylated MAPK during cell cycle progression. The SQ RT-PCR analysis indicated that MAPK expression is lower in mitosis than in interphase (G1, G2 and S). However, the amount of phosphorylated MAPK remained stable throughout the cell cycle, indicating that MAPK activity is predominantly regulated at the post-translational level and that phosphorylation of MAPK plays an important role in mitosis. Application of the specific MAPK phosphorylation inhibitor U0126 revealed that while U0126 treatment decreases the phosphorylation of MAPK and the progression from telophase to early cytokinesis is significantly inhibited. The formation of the phragmoplast is also negatively affected at this stage. These results demonstrate that MAPK phosphorylation is involved in the formation of the cell plate within the phragmoplast during cytokinesis and that MAPK predominantly functions during the cytokinesis stage of the cell cycle in tobacco BY-2 cells. Copyright 2009 Elsevier GmbH. All rights reserved.

  11. Tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) induces cell proliferation in normal human bronchial epithelial cells through NFκB activation and cyclin D1 up-regulation

    International Nuclear Information System (INIS)

    Ho, Y.-S.; Chen, Chien-Ho; Wang, Y.-J.; Pestell, Richard G.; Albanese, Chris; Chen, R.-J.; Chang, M.-C.; Jeng, J.-H.; Lin, S.-Y.; Liang, Y.-C.; Tseng, H.; Lee, W.-S.; Lin, J.-K.; Chu, J.-S.; Chen, L.-C.; Lee, C.-H.; Tso, W.-L.; Lai, Y.-C.; Wu, C.-H.

    2005-01-01

    Cigarette smoke contains several carcinogens known to initiate and promote tumorigenesis as well as metastasis. Nicotine is one of the major components of the cigarette smoke and the 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a tobacco-specific carcinogen. Here, we demonstrated that NNK stimulated cell proliferation in normal human bronchial epithelial cells (NHBE) and small airway epithelial cells (SAEC). Cells exposed to NNK resulted in an increase in the level of cyclin D1 protein (as early as 3-6 h). Increased phosphorylation of the Rb Ser 795 was detected at 6-15 h after NNK treatment and thereby promoted cells entering into the S phase (at 15-21 h). The increased cyclin D1 protein level was induced through activation of the transcription factor, nuclear factor kB (NFκB), in the NHBE cells. Treatment of the NHBE cells with PD98059, an ERK1/2 (extracellular signal-regulated protein kinase)-specific inhibitor, specifically suppressed the NNK-induced IκBα phosphorylation at position 32 of the serine residue, suggesting that the ERK1/2 kinase was involved in the IκBα phosphorylation induced by NFκB activation. To determine whether the NNK-induced NFκB activation and cyclin D1 induction were also observed in vivo, A/J mice were treated with NNK (9.1 mg) for 20 weeks and the results showed a significant induction of cyclin D1 and NFκB translocation determined by immunoblotting analyses. We further demonstrated that the nicotine acetylcholine receptor (nAchR), which contains the α3-subunit, was the major target mediating NNK-induced cyclin D1 expression in the NHBE cells. In summary, our findings demonstrate for the first time that NNK could stimulate normal human bronchial cell proliferation through activation of the NFκB, which in turn up-regulated the cyclin D1 expression

  12. High-yield secretion of recombinant proteins expressed in tobacco cell culture with a designer glycopeptide tag: Process development.

    Science.gov (United States)

    Zhang, Ningning; Gonzalez, Maria; Savary, Brett; Xu, Jianfeng

    2016-03-01

    Low-yield protein production remains the most significant economic hurdle with plant cell culture technology. Fusions of recombinant proteins with hydroxyproline-O-glycosylated designer glycopeptide tags have consistently boosted secreted protein yields. This prompted us to study the process development of this technology aiming to achieve productivity levels necessary for commercial viability. We used a tobacco BY-2 cell culture expressing EGFP as fusion with a glycopeptide tag comprised of 32 repeat of "Ser-Pro" dipeptide, or (SP)32 , to study cell growth and protein secretion, culture scale-up, and establishment of perfusion cultures for continuous production. The BY-2 cells accumulated low levels of cell biomass (~7.5 g DW/L) in Schenk & Hildebrandt medium, but secreted high yields of (SP)32 -tagged EGFP (125 mg/L). Protein productivity of the cell culture has been stable for 6.0 years. The BY-2 cells cultured in a 5-L bioreactor similarly produced high secreted protein yield at 131 mg/L. Successful operation of a cell perfusion culture for 30 days was achieved under the perfusion rate of 0.25 and 0.5 day(-1) , generating a protein volumetric productivity of 17.6 and 28.9 mg/day/L, respectively. This research demonstrates the great potential of the designer glycopeptide technology for use in commercial production of valuable proteins with plant cell cultures. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Increased sesquiterpenoid biosynthesis and an apparent decrease in sterol biosynthesis in elicitor-treated tobacco cell suspension cultures

    International Nuclear Information System (INIS)

    Voegeli, U.; Bhatt, P.N.; Chappell, J.

    1987-01-01

    Addition of fungel elicitor prepared from Phytophthora parasitica to tobacco cell suspension cultures leads to an increased production of the phytoalexin capsidiol. Capsidiol is a sesquiterpenoid which is most likely synthesized from farnesylpyrophosphat (FPP) by a bicyclic cyclase reaction. Because FPP is also a substrate for squalene synthetase and therefore a precursor of sterol biosynthesis, the question arises whether or not the accumulation of capsidiol in elicitor-treated cells occurs at the expense of sterol biosynthesis. ( 14 C]-acetate was given to elicitor-treated and control (no treatment) cell cultures and incorporation into sterols and capsidiol determined. No labeled capsidiol was detected in control cells. In elicitor-treated cells about 12-15% of the radioactivity taken up by the cells was incorporated into capsidiol. In contrast, control cells incorporated 4 times more radioactivity into sterols than elicitor-treated cells. Similar results were obtained using ( 3 H)-mevalonate as a precursor of capsidiol and sterol biosynthesis. Likely explanations for the apparently decline in sterol biosynthesis in elicitor-treated cells include: (1) inhibition of squalene synthetase; (2) induction of capsidiol synthesizing enzymes; and (3) metabolic channeling of FPP into capsidiol versus sterols. These possibilities will be discussed further together with other results

  14. Alterations in body weight and blood glucose level of female hamsters exposed to electromagnetic fields of cell phones

    Directory of Open Access Journals (Sweden)

    A.R Lotfi

    2010-02-01

    Group 2 was exposed to electromagnetic field emitted by cell phones for 10 days (short term and group 3 for 50 day (long term. In the latter groups, the exposure was 1 hour per day. At the end of the experimental period, the animals were weighed and blood glucose concentrations were determined by obtaining blood samples from 8 randomly selected hamsters in each group.  The blood glucose level was significantly higher in long-term exposed group in comparison with the control and short-term exposed groups (175, 11.6 and 107 mg/dl, respectively (p

  15. The protective effect of curcumin in Olfactory Ensheathing Cells exposed to hypoxia.

    Science.gov (United States)

    Bonfanti, Roberta; Musumeci, Teresa; Russo, Cristina; Pellitteri, Rosalia

    2017-02-05

    Curcumin, a phytochemical component derived from the rhizomes of Curcuma longa, has shown a great variety of pharmacological activities, such as anti-inflammatory, anti-tumor, anti-depression and anti-oxidant activity. Therefore, in the last years it has been used as a therapeutic agent since it confers protection in different neurodegenerative diseases, cerebral ischemia and excitotoxicity. Olfactory Ensheathing Cells (OECs) are glial cells of the olfactory system. They are able to secrete several neurotrophic growth factors, promote axonal growth and support the remyelination of damaged axons. OEC transplantation has emerged as a possible experimental therapy to induce repair of spinal cord injury, even if the functional recovery is still limited. Since hypoxia is a secondary effect in spinal cord injury, this in vitro study investigates the protective effect of curcumin in OECs exposed to hypoxia. Primary OECs were obtained from neonatal rat olfactory bulbs and placed both in normal and hypoxic conditions. Furthermore, some cells were grown with basic Fibroblast Growth Factor (bFGF) and/or curcumin at different concentration and times. The results obtained through immunocytochemical procedures and MTT test show that curcumin stimulates cell viability in OECs grown in normal and hypoxic conditions. Furthermore, the synergistic effect of curcumin and bFGF is the most effective exerting protection on OECs. Since spinal cord injury is often accompanied by secondary insults, such as ischemia or hypoxia, our results suggest that curcumin in combination with bFGF might be considered a possible approach for restoration in injuries. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Construction and identification of differential expression genes of peripheral blood cells in radon-exposed mice

    International Nuclear Information System (INIS)

    Chen Rui; Shi Minhua; Hu Huacheng; Li Jianxiang; Nie Jihua; Tong Jian

    2009-01-01

    Objective: To screen and identify the differential expression genes on peripheral blood cells of mice based on the experimental animal model of radon exposure. Methods: BALB/c mice were exposed in a type HD-3 multifunctional radon-room, with the accumulative doses of radon-exposure group at 105 WLM and control group at 1 WLM. Total RNA was extracted from peripheral blood cells and the methods of SMART for dscDNA synthesis and SSH for gene screening was applied. With the construction of the cDNA library enriched with differentially expressed genes, the pMD 18-T plasmid containing LacZ operator at the multiple cloning site was used to allow a blue-white screening. The TA clones were amplified by nested PCR and the reverse Northern blot was used to identify up and down regulation of the clones. The differently expressed cDNA was then sequenced and analyzed. Results: The subtracted cDNA libraries were successfully constructed. A total of 390 recombinant white colonies were randomly selected. Among the 312 cDNA monoclones selected from both forward- and reverse-subtracted libraries, 41 clones were chosen to sequence for their differential expressions based on reverse Northern blot. Among the 41 sequenced clones, 10 clones with known function/annotation and 3 new ESTs with the GenBank accession numbers were obtained. Most of the known function/annotation genes were revealed to be related with cell proliferation, metabolism, cellular apoptosis and carcinogenesis. Conclusions: The animal model of radon exposure was established and the cDNA library of peripheral blood cells was successfully constructed. Radon exposure could up- and down-regulate a series of genes. Differentially expressed genes could be identified by using SSH technique and the results may help exploring mechanisms of random exposure. (authors)

  17. Radiosensitization of mammalian cells by misonidazole and oxygen: DNA damage exposed by Micrococcus luteus enzymes

    International Nuclear Information System (INIS)

    Skov, K.A.; Palcic, B.; Skarsgard, L.D.

    1979-01-01

    When misonidazole is present during irradiation of hypoxic mammalian cells, an enhancement of single-strand breaks (SSB) in DNA is observed. Oxygen also enhances SSB, presumably in a manner similar to that of misonidazole. The dose-modifying factor (DMF) for 15 mM misonidazole was found to be 3.4, compared to an oxygen enhancement ratio (OER) of 3.5. Another class of DNA damage, namely, sites exposed by an extract of Micrococcus luteus, was examined. Radiation-induced M. luteus extract-sensitive sites (MLS) were also found to be enhanced by the presence of misonidazole or molecular oxygen. The DMF for this damage by 15 mM misonidazole was 1.6 while the OER was 2.5. The ratio of MLS to SSB is approximately 1.25 under hypoxia, 0.9 in the presence of oxygen, and 0.6 in the presence of 15 mM misonidazole under hypoxic conditions. Incubation with misonidazole under conditions which are toxic to mammalian cells (37 0 C, hypoxia), and which result in many SSB, produces no detectable lesions sensitive to the M. luteus extract

  18. Enhanced inhibition of parvovirus B19 replication by cidofovir in extendedly exposed erythroid progenitor cells.

    Science.gov (United States)

    Bonvicini, Francesca; Bua, Gloria; Manaresi, Elisabetta; Gallinella, Giorgio

    2016-07-15

    Human parvovirus B19 (B19V) commonly induces self-limiting infections but can also cause severe clinical manifestations in patients with underlying haematological disorders or with immune system deficits. Currently, therapeutic options for B19V entirely rely on symptomatic and supportive treatments since a specific antiviral therapy is not yet available. Recently a first step in the research for active compounds inhibiting B19V replication has allowed identifying the acyclic nucleoside phosphonate cidofovir (CDV). Herein, the effect of CDV against B19V replication was characterized in human erythroid progenitor cells (EPCs) cultured and infected following different experimental approaches to replicate in vitro the infection of an expanding erythroid cell population in the bone marrow. B19V replication was selectively inhibited both in infected EPCs extendedly exposed to CDV 500μM (viral inhibition 82%) and in serially infected EPCs cultures with passage of the virus progeny, constantly under drug exposure (viral inhibition 99%). In addition, a potent inhibitory effect against B19V (viral inhibition 92%) was assessed in a short-term infection of EPCs treated with CDV 500μM 1day before viral infection. In the evaluated experimental conditions, the enhanced effect of CDV against B19V might be ascribed both to the increased intracellular drug concentration achieved by extended exposure, and to a progressive reduction in efficiency of the replicative process within treated EPCs population. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. DIFFERENTIAL SENSITIVITY OF MALE GERM CELLS TO MAINSTREAM AND SIDESTREAM TOBACCO SMOKE IN THE MOUSE

    Energy Technology Data Exchange (ETDEWEB)

    Polyzos, Aris; Schmid, Thomas Ernst; Pina-Guzman, Belem; Quintanilla-Vega, Betzabet; Marchetti, Francesco

    2009-03-13

    Cigarette smoking in men has been associated with increased chromosomal abnormalities in sperm and with increased risks for spontaneous abortions, birth defects and neonatal death. Little is known, however, about the reproductive consequences of paternal exposure to second-hand smoke. We used a mouse model to investigate the effects of paternal exposure to sidestream (SS) smoke, the main constituent of second-hand smoke, on the genetic integrity and function of sperm, and to determine whether male germ cells were equally sensitive to mainstream (MS) and SS smoke. A series of sperm DNA quality and reproductive endpoints were investigated after exposing male mice for two weeks to MS or SS smoke. Our results indicated that: (i) only SS smoke significantly affected sperm motility; (ii) only MS smoke induced DNA strand breaks in sperm; (iii) both MS and SS smoke increased sperm chromatin structure abnormalities; and (iv) MS smoke affected both fertilization and the rate of early embryonic development, while SS smoke affected fertilization only. These results show that MS and SS smoke have differential effects on the genetic integrity and function of sperm and provide further evidence that male exposure to second-hand smoke, as well as direct cigarette smoke, may diminish a couple's chance for a successful pregnancy and the birth of a healthy baby.

  20. Proliferation assay of mouse embryonic stem (ES) cells exposed to atmospheric-pressure plasmas at room temperature

    International Nuclear Information System (INIS)

    Miura, Taichi; Hirano, Kazumi; Ogura, Chika; Ikeguchi, Masamichi; Seki, Atsushi; Nishihara, Shoko; Ando, Ayumi; Kanazawa, Tatsuya; Hamaguchi, Satoshi

    2014-01-01

    Proliferation assays of mouse embryonic stem (ES) cells have been performed with cell culture media exposed to atmospheric-pressure plasmas (APPs), which generate reactive species in the media at room temperature. It is found that serum in cell culture media functions as a scavenger of highly reactive species and tends to protect cells in the media against cellular damage. On the other hand, if serum is not present in a cell culture medium when it is exposed to APP, the medium becomes cytotoxic and cannot be detoxified by serum added afterwards. Plasma-induced cytotoxic media hinder proliferation of mouse ES cells and may even cause cell death. It is also shown by nuclear magnetic resonance spectroscopy that organic compounds in cell culture media are in general not significantly modified by plasma exposure. These results indicate that if there is no serum in media when they are exposed to APPs, highly reactive species (such as OH radicals) generated in the media by the APP exposure are immediately converted to less reactive species (such as H 2 O 2 ), which can no longer readily react with serum that is added to the medium after plasma exposure. This study has clearly shown that it is these less reactive species, rather than highly reactive species, that make the medium cytotoxic to mouse ES cells. (paper)

  1. Acute respiratory bronchiolitis: an ultrastructural and autoradiographic study of epithelial cell injury and renewal in Rhesus monkeys exposed to ozone

    International Nuclear Information System (INIS)

    Castleman, W.L.; Dungworth, D.L.; Schwartz, L.W.; Tyler, W.S.

    1980-01-01

    The pathogenesis of acute respiratory bronchiolitis was examined in Rhesus monkeys exposed to 0.8 ppM ozone for 4 to 50 hours. Epithelial injury and renewal were qualitatively and quantitatively characterized by correlated techniques of scanning and transmission electron microscopy as well as by light-microscopic autoradiography following labeling with tritiated thymidine. Extensive degeneration and necrosis of Type 1 epithelial cells occurred on the respiratory bronchiolar wall during the initial 4 to 12 hours of exposure. Increased numbers of labeled epithelial cells were present in this region after 18 hours of exposure, and the highest labeling index (18%) was measured after 50 hours of exposure. Most (67 to 80%) of the labeled cells and all the mitotic epithelial cells (22) observed ultrastructurally were cuboidal bronchiolar epithelial cells. Of the labeled epithelial cells, 20 to 33% were Type 2 epithelial cells. After 50 hours of exposure the respiratory bronchiolar epithelium was hyperplastic. The predominant inflammatory cell in respiratory bronchiolar exudate was the alveolar macrophage. Monkeys that were exposed for 50 hours and allowed to recover in unozonized air for 7 days had incomplete resolution of respiratory bronchiolar epithelial hyperplasia. The results indicate that Type 1 epithelial cells lining respiratory bronchioles are the cell types most sensitive to injury and that both cuboidal bronchiolar epithelial cells and Type 2 epithelial cells function as stem cells in epithelial renewal

  2. Comet assay in gill cells of Prochilodus lineatus exposed in vivo to cypermethrin.

    Science.gov (United States)

    Poletta, G L; Gigena, F; Loteste, A; Parma, M J; Kleinsorge, E C; Simoniello, M F

    2013-11-01

    Agricultural chemicals can induce genetic alterations on aquatic organisms that have been associated with effects on growth, reproduction and population dynamics. The evaluation of DNA damage in fish using the comet assay (CA) frequently involves the utilization of erythrocytes. However, epithelial gill cells (EGC) can be more sensitive, as they are constantly dividing and in direct contact with potentially stressing compounds from the aquatic environment. The aim of the present study was to evaluate (1) the sensitivity and suitability of epithelial gill cells of Prochilodus lineatus in response to different genotoxic agents through the application of the CA, (2) the induction of DNA damage in this cell population after in vivo exposure to cypermethrin. Baseline value of the CA damage index (DI) for EGC of juvenile P. lineatus was 144.68±5.69. Damage increased in a dose-dependent manner after in vitro exposure of EGC to methyl methanesulfonate (MMS) and H2O2, two known genotoxic agents. In vivo exposure of fish to cypermethrin induced a significant increase in DNA DI of EGC at 0.150μg/l (DI: 239.62±6.21) and 0.300μg/l (270.63±2.09) compared to control (150.25±4.38) but no effect was observed at 0.075μg/l (168.50±10.77). This study shows that EGC of this species are sensitive for the application of the CA, demonstrating DNA damage in response to alkylation (MMS), oxidative damage (H2O2), and to the insecticide cypermethryn. These data, together with our previous study on DNA damage induction on erythrocytes of this species, provides useful information for future work involving biomonitoring in regions where P. lineatus is naturally exposed to pesticides and other genotoxic agents. Copyright © 2013 Elsevier Inc. All rights reserved.

  3. Gene expression profiles in asbestos-exposed epithelial and mesothelial lung cell lines

    Directory of Open Access Journals (Sweden)

    Kaski Samuel

    2007-03-01

    Full Text Available Abstract Background Asbestos has been shown to cause chromosomal damage and DNA aberrations. Exposure to asbestos causes many lung diseases e.g. asbestosis, malignant mesothelioma, and lung cancer, but the disease-related processes are still largely unknown. We exposed the human cell lines A549, Beas-2B and Met5A to crocidolite asbestos and determined time-dependent gene expression profiles by using Affymetrix arrays. The hybridization data was analyzed by using an algorithm specifically designed for clustering of short time series expression data. A canonical correlation analysis was applied to identify correlations between the cell lines, and a Gene Ontology analysis method for the identification of enriched, differentially expressed biological processes. Results We recognized a large number of previously known as well as new potential asbestos-associated genes and biological processes, and identified chromosomal regions enriched with genes potentially contributing to common responses to asbestos in these cell lines. These include genes such as the thioredoxin domain containing gene (TXNDC and the potential tumor suppressor, BCL2/adenovirus E1B 19kD-interacting protein gene (BNIP3L, GO-terms such as "positive regulation of I-kappaB kinase/NF-kappaB cascade" and "positive regulation of transcription, DNA-dependent", and chromosomal regions such as 2p22, 9p13, and 14q21. We present the complete data sets as Additional files. Conclusion This study identifies several interesting targets for further investigation in relation to asbestos-associated diseases.

  4. Long-chain bases and their phosphorylated derivatives differentially regulate cryptogein-induced production of reactive oxygen species in tobacco (Nicotiana tabacum) BY-2 cells.

    Science.gov (United States)

    Coursol, Sylvie; Fromentin, Jérôme; Noirot, Elodie; Brière, Christian; Robert, Franck; Morel, Johanne; Liang, Yun-Kuan; Lherminier, Jeannine; Simon-Plas, Françoise

    2015-02-01

    The proteinaceous elicitor cryptogein triggers defence reactions in Nicotiana tabacum (tobacco) through a signalling cascade, including the early production of reactive oxygen species (ROS) by the plasma membrane (PM)-located tobacco respiratory burst oxidase homologue D (NtRbohD). Sphingolipid long-chain bases (LCBs) are emerging as potent positive regulators of plant defence-related mechanisms. This led us to question whether both LCBs and their phosphorylated derivatives (LCB-Ps) are involved in the early signalling process triggered by cryptogein in tobacco BY-2 cells. Here, we showed that cryptogein-induced ROS production was inhibited by LCB kinase (LCBK) inhibitors. Additionally, Arabidopsis thaliana sphingosine kinase 1 and exogenously supplied LCB-Ps increased cryptogein-induced ROS production, whereas exogenously supplied LCBs had a strong opposite effect, which was not driven by a reduction in cellular viability. Immunogold-electron microscopy assay also revealed that LCB-Ps are present in the PM, which fits well with the presence of a high LCBK activity associated with this fraction. Our data demonstrate that LCBs and LCB-Ps differentially regulate cryptogein-induced ROS production in tobacco BY-2 cells, and support a model in which a cooperative synergism between LCBK/LCB-Ps and NtRbohD/ROS in the cryptogein signalling pathway is likely at the PM in tobacco BY-2 cells. © 2014 INRA New Phytologist © 2014 New Phytologist Trust.

  5. The aux1 gene of the Ri plasmid is sufficient to confer auxin autotrophy in tobacco BY-2 cells.

    Science.gov (United States)

    Nemoto, Keiichirou; Hara, Masamitsu; Goto, Shingo; Kasai, Kouji; Seki, Hikaru; Suzuki, Masashi; Oka, Atsuhiro; Muranaka, Toshiya; Mano, Yoshihiro

    2009-05-01

    Tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) cells are rapidly proliferating meristematic cells that require auxin for culture in vitro. We have established several transgenic BY-2 cell lines that carry the T-DNA of Agrobacterium rhizogenes 15834, which harbors an agropine-type root-inducing (Ri) plasmid. Two of these lines, BYHR-3 and BYHR-7, were used to test the role of auxin in the proliferation of plant cells. The lines grew rapidly in Linsmaier-Skoog (LS) medium lacking auxin and other phytohormones. The TR-DNA, containing the aux1 (tryptophan monooxygenase) and aux2 (indoleacetamide hydrolase) genes, was present in the genomes of both transgenic lines, whereas the TL-DNA, containing the rolA, B, C and D genes, was present in the genome of BYHR-7 but not BYHR-3. Since the introduction of the rolABCD genes alone did not affect the auxin requirement of BY-2 cells, the aux1 and aux2 genes, but not the rolABCD genes, appear to be relevant to the auxin autotrophy of these transgenic lines. Furthermore, the overexpression of aux1 allowed BY-2 cells to grow rapidly in the absence of auxin, suggesting the existence in plant cells of an unidentified gene whose product is functionally equivalent or similar to that of aux2 of the Ri plasmid.

  6. Comparative assessment of HPV, alcohol and tobacco etiological fractions in Algerian patients with laryngeal squamous cell carcinoma.

    Science.gov (United States)

    Kariche, Nora; Hortal, Montserrat Torres; Benyahia, Samir; Alemany, Laia; Moulaï, Nabila; Clavero, Omar; Muñoz, Marleny; Ouahioune, Wahiba; Djennaoui, Djamel; Touil-Boukoffa, Chafia; de Sanjosé, Silvia; Bourouba, Mehdi

    2018-01-01

    Despite the increasing incidence of laryngeal squamous cell carcinoma (LSCC) in Algeria, scarce information is available on the importance of the preventable etiological factors which may drive the disease. Remarkably, a significant number of cases occur in nonsmoker and nondrinker patients; hence, suggesting that alternative risk factors, like Human papillomavirus (HPV), might be etiologically involved. To gain more insight on the risk factors associated with the disease in the country, we evaluated the etiological fraction of HPV in comparison to tobacco and alcohol intake in LSCC patients. To evaluate the etiopathologic fraction (EF) for HPV compared to history of tobacco and alcohol in LSCC, HPV DNA presence in 46 invasive and 3 non-invasive formalin-fixed paraffin-embedded laryngeal tumors was screened using the SPF10-DEIA-LiPA25 Assay. Demographic data and information related to exposure to the risk factors were gathered through interviewer-assisted questionnaires. We observed that 40.8% of all LSCC cases were associated with smoking, 40.8% had combined tobacco and alcohol exposure history, and 14.3% did not show prior exposure to either risk factor. 1 out of 3 in-situ carcinoma cases was positive for HPV-6. HPV prevalence was null in the invasive tumors. HPV DNA was detected in 2.38% for all studied cases. 10.2% of LSCC patients did not associate with any of the studied risk factors. Here we show that HPV etiological fraction in LSCC Algerian patients is low and smoking and alcohol remain the principal etiopathologic risk for LSCC burden in Algeria.

  7. Dynamic trafficking of wheat γ-gliadin and of its structural domains in tobacco cells, studied with fluorescent protein fusions

    Science.gov (United States)

    Francin-Allami, Mathilde; Saumonneau, Amélie; Lavenant, Laurence; Bouder, Axelle; Sparkes, Imogen; Hawes, Chris; Popineau, Yves

    2011-01-01

    Prolamins, the main storage proteins of wheat seeds, are synthesized and retained in the endoplasmic reticulum (ER) of the endosperm cells, where they accumulate in protein bodies (PBs) and are then exported to the storage vacuole. The mechanisms leading to these events are unresolved. To investigate this unconventional trafficking pathway, wheat γ-gliadin and its isolated repeated N-terminal and cysteine-rich C-terminal domains were fused to fluorescent proteins and expressed in tobacco leaf epidermal cells. The results indicated that γ-gliadin and both isolated domains were able to be retained and accumulated as protein body-like structures (PBLS) in the ER, suggesting that tandem repeats are not the only sequence involved in γ-gliadin ER retention and PBLS formation. The high actin-dependent mobility of γ-gliadin PBLS is also reported, and it is demonstrated that most of them do not co-localize with Golgi body or pre-vacuolar compartment markers. Both γ-gliadin domains are found in the same PBLS when co-expressed, which is most probably due to their ability to interact with each other, as indicated by the yeast two-hybrid and FRET-FLIM experiments. Moreover, when stably expressed in BY-2 cells, green fluorescent protein (GFP) fusions to γ-gliadin and its isolated domains were retained in the ER for several days before being exported to the vacuole in a Golgi-dependent manner, and degraded, leading to the release of the GFP ‘core’. Taken together, the results show that tobacco cells are a convenient model to study the atypical wheat prolamin trafficking with fluorescent protein fusions. PMID:21617248

  8. DNA damage induced by hydrogen peroxide in cultured tobacco cells is dependent on the cell growth stage

    Czech Academy of Sciences Publication Activity Database

    Stavreva, D.; Gichner, Tomáš

    2002-01-01

    Roč. 514, - (2002), s. 147-152 ISSN 1383-5718 R&D Projects: GA ČR GA521/02/0400 Institutional research plan: CEZ:AV0Z5038910 Keywords : DNA * peroxide * tobacco Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.636, year: 2002

  9. Oxidative stress response in neural stem cells exposed to different superparamagnetic iron oxide nanoparticles

    Directory of Open Access Journals (Sweden)

    Pongrac IM

    2016-04-01

    Full Text Available Igor M Pongrac,1 Ivan Pavičić,2 Mirta Milić,2 Lada Brkič Ahmed,1 Michal Babič,3 Daniel Horák,3 Ivana Vinković Vrček,2 Srećko Gajović1 1School of Medicine, Croatian Institute for Brain Research, University of Zagreb, 2Institute for Medical Research and Occupational Health, Zagreb, Croatia; 3Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic Abstract: Biocompatibility, safety, and risk assessments of superparamagnetic iron oxide nanoparticles (SPIONs are of the highest priority in researching their application in biomedicine. One improvement in the biological properties of SPIONs may be achieved by different functionalization and surface modifications. This study aims to investigate how a different surface functionalization of SPIONs – uncoated, coated with D-mannose, or coated with poly-L-lysine – affects biocompatibility. We sought to investigate murine neural stem cells (NSCs as important model system for regenerative medicine. To reveal the possible mechanism of toxicity of SPIONs on NSCs, levels of reactive oxygen species, intracellular glutathione, mitochondrial membrane potential, cell-membrane potential, DNA damage, and activities of SOD and GPx were examined. Even in cases where reactive oxygen species levels were significantly lowered in NSCs exposed to SPIONs, we found depleted intracellular glutathione levels, altered activities of SOD and GPx, hyperpolarization of the mitochondrial membrane, dissipated cell-membrane potential, and increased DNA damage, irrespective of the surface coating applied for SPION stabilization. Although surface coating should prevent the toxic effects of SPIONs, our results showed that all of the tested SPION types affected the NSCs similarly, indicating that mitochondrial homeostasis is their major cellular target. Despite the claimed biomedical benefits of SPIONs, the refined determination of their effects on various cellular functions

  10. DNA repair studies in mouse germ cells exposed to two carcinogens and two non-carcinogens

    International Nuclear Information System (INIS)

    Sega, G.A.; Owens, J.G.

    1987-01-01

    An in vivo test was used to measure induced unscheduled DNA synthesis (UDS) in the germ cells of male mice exposed to the carcinogens benzo(a)pyrene [B(a)P] and 2-acetylaminofluorene (2AAF), and to the noncarcinogens pyrene (PYR) and 4-acetylaminofluorene (4AAF). Early spermatids, a DNA-repair competent stage, were used to test the effects of all chemicals. After chemical treatment and testicular injection of [ 3 H]dThd, sperm were recovered 16 days later from the caudal epididymides (these sperm were in early spermatid stages at the time of treatment) and assayed for the unscheduled incorporation of [ 3 H]dThd using liquid scintillation counting (LSC). Exposures of 2AAF ranged from 125 to 1600 mg/kg, 4AAF from 125 to 2000 mg/kg, PYR from 100 to 600 mg/kg, B(a)P from 100 to 400 mg/kg. Chemicals were administered both by intraperitoneal (i.p.) injection and by gavage. Methyl methanesulfonate (MMS) was used as a positive control

  11. Fibrogenic response of hepatic stellate cells in ovariectomised rats exposed to ketogenic diet.

    Science.gov (United States)

    Bobowiec, R; Wojcik, M; Jaworska-Adamu, J; Tusinska, E

    2013-02-01

    The discrepancy about the role of estrogens in hepatic fibrogenesis and lack of studies addressed of ketogenic diet (KD) on hepatic stellate cells (HSC), prompted us to investigate the activity of HSC in control, KD- and thioacetamide (TAA)-administrated rats with different plasma concentration of estradiol (E2). HSC were isolated by the collagenase perfusion methods and separated by the Percoll gradient centrifugation. After the 4(th) and 8(th) day of incubation, lysates of HSC and the media were collected for further analysis. The HSC derived from KD-rats released remarkably more transforming growth factor (TGF)-β1 than cells obtained from animals fed with a standard diet. The ovariectomy of KD-rats markedly intensified the secretion of this fibrogenic cytokine on the 8(th) day of incubation (201.33 ±1 7.15 pg/ml). In HSC of rats exposed to E2, the TGF-β1 concentration did not exceed 157 ± 34.39 pg/ml. In respect to the collagen type I, the HSC obtained from ovariectomised KD-rats released an augmented amount of this ECM protein after the 8(th) day of culture (1.83 ± 0.14 U/ml). In the same time, higher quantities of ASMA appeared in the KD rats (1.41 ± 0.3 pg/mg protein). Exposition of rats to E2 did not markedly decrease the amount of ASMA. In summary, KD was able to induce morphological and functional changes in HSC, especially derived from rats deprived of ovarian estrogens. However, the preservation of E2 in ovariectomised rats didn't substantially alter the activation of HSC.

  12. Cytoskeletal dynamics in interphase, mitosis and cytokinesis analysed through Agrobacterium-mediated transient transformation of tobacco BY-2 cells.

    Science.gov (United States)

    Buschmann, H; Green, P; Sambade, A; Doonan, J H; Lloyd, C W

    2011-04-01

    Transient transformation with Agrobacterium is a widespread tool allowing rapid expression analyses in plants. However, the available methods generate expression in interphase and do not allow the routine analysis of dividing cells. Here, we present a transient transformation method (termed 'TAMBY2') to enable cell biological studies in interphase and cell division. Agrobacterium-mediated transient gene expression in tobacco BY-2 was analysed by Western blotting and quantitative fluorescence microscopy. Time-lapse microscopy of cytoskeletal markers was employed to monitor cell division. Double-labelling in interphase and mitosis enabled localization studies. We found that the transient transformation efficiency was highest when BY-2/Agrobacterium co-cultivation was performed on solid medium. Transformants produced in this way divided at high frequency. We demonstrated the utility of the method by defining the behaviour of a previously uncharacterized microtubule motor, KinG, throughout the cell cycle. Our analyses demonstrated that TAMBY2 provides a flexible tool for the transient transformation of BY-2 with Agrobacterium. Fluorescence double-labelling showed that KinG localizes to microtubules and to F-actin. In interphase, KinG accumulates on microtubule lagging ends, suggesting a minus-end-directed function in vivo. Time-lapse studies of cell division showed that GFP-KinG strongly labels preprophase band and phragmoplast, but not the metaphase spindle. © 2010 The Authors. New Phytologist © 2010 New Phytologist Trust.

  13. Unirradiated cells rescue cells exposed to ionizing radiation: Activation of NF-κB pathway in irradiated cells

    Energy Technology Data Exchange (ETDEWEB)

    Lam, R.K.K. [Department of Physics and Materials Science, City University of Hong Kong (Hong Kong); Han, Wei [Center of Medical Physics and Technology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Yu, K.N., E-mail: peter.yu@cityu.edu.hk [Department of Physics and Materials Science, City University of Hong Kong (Hong Kong); State Key Laboratory in Marine Pollution, City University of Hong Kong (Hong Kong)

    2015-12-15

    Highlights: • Rescue effect was observed in both irradiated and HeLa and NIH/3T3 cells. • Novel setup and procedures to separate the rescue signals and the bystander signals. • Confirmed activation of NF-κB pathway in rescue effect using activation inhibitor. • Confirmed activation of NF-κB pathway in rescue effect using anti-NF-κB p65 antibody. - Abstract: We studied the involvement of NF-κB pathway activation in the rescue effect in HeLa and NIH/3T3 cells irradiated by α particles. Firstly, upon irradiation by 5 cGy of α particles, for both cell lines, the numbers of 53BP1 foci/cell at 12 h post-irradiation were significantly smaller when only 2.5% of the cell population was irradiated as compared to 100% irradiation, which demonstrated the rescue effect. Secondly, we studied the effect of NF-κB on the rescue effect through the use of the NF-κB activation inhibitor BAY-11-7082. Novel experimental setup and procedures were designed to prepare the medium (CM) which had conditioned the bystander cells previously partnered with irradiated cells, to ensure physical separation between rescue and bystander signals. BAY-11-7082 itself did not inflict DNA damages in the cells or have effects on activation of the NF-κB response pathway in the irradiated cells through direct irradiation. The rescue effect was induced in both cell lines by the CM, which was abrogated if BAY-11-7082 was added to the CM. Thirdly, we studied the effect of NF-κB on the rescue effect through staining for phosphorylated NF-κB (p-NF-κB) expression using the anti-NF-κB p65 (phospho S536) antibody. When the fraction of irradiated cells dropped from 100% to 2.5%, the p-NF-κB expression in the cell nuclei of irradiated NIH/3T3 cells increased significantly, while that in the cell nuclei of irradiated HeLa cells also increased although not significantly. Moreover, the p-NF-κB expression in the cell nuclei of irradiated HeLa cells and NIH/3T3 cells treated with CM also increased

  14. Differential responses to high- and low-dose ultraviolet-B stress in tobacco Bright Yellow-2 cells

    Science.gov (United States)

    Takahashi, Shinya; Kojo, Kei H.; Kutsuna, Natsumaro; Endo, Masaki; Toki, Seiichi; Isoda, Hiroko; Hasezawa, Seiichiro

    2015-01-01

    Ultraviolet (UV)-B irradiation leads to DNA damage, cell cycle arrest, growth inhibition, and cell death. To evaluate the UV-B stress–induced changes in plant cells, we developed a model system based on tobacco Bright Yellow-2 (BY-2) cells. Both low-dose UV-B (low UV-B: 740 J m−2) and high-dose UV-B (high UV-B: 2960 J m−2) inhibited cell proliferation and induced cell death; these effects were more pronounced at high UV-B. Flow cytometry showed cell cycle arrest within 1 day after UV-B irradiation; neither low- nor high-UV-B–irradiated cells entered mitosis within 12 h. Cell cycle progression was gradually restored in low-UV-B–irradiated cells but not in high-UV-B–irradiated cells. UV-A irradiation, which activates cyclobutane pyrimidine dimer (CPD) photolyase, reduced inhibition of cell proliferation by low but not high UV-B and suppressed high-UV-B–induced cell death. UV-B induced CPD formation in a dose-dependent manner. The amounts of CPDs decreased gradually within 3 days in low-UV-B–irradiated cells, but remained elevated after 3 days in high-UV-B–irradiated cells. Low UV-B slightly increased the number of DNA single-strand breaks detected by the comet assay at 1 day after irradiation, and then decreased at 2 and 3 days after irradiation. High UV-B increased DNA fragmentation detected by the terminal deoxynucleotidyl transferase dUTP nick end labeling assay 1 and 3 days after irradiation. Caffeine, an inhibitor of ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3-related (ATR) checkpoint kinases, reduced the rate of cell death in high-UV-B–irradiated cells. Our data suggest that low-UV-B–induced CPDs and/or DNA strand-breaks inhibit DNA replication and proliferation of BY-2 cells, whereas larger contents of high-UV-B–induced CPDs and/or DNA strand-breaks lead to cell death. PMID:25954287

  15. Pollen mitochondria in cytoplasmically male sterile tobacco zygotic and embryonic cells

    International Nuclear Information System (INIS)

    Symillides, Y.

    1985-09-01

    An attempt is being made to establish cytoplasmic organelles transmission during the process of fertilization, by using tobacco grain pollen labelled with leucine 14 C and tritiated thymidine. Through autoradiography the fate of pollen germination and its entry into the embryo sac has been studied. A few days after fertilization, labelled cytoplasmic organelles - mainly mitochondria - were detected in the embryo sac. However, labelling was not observed in cytoplasmic organelles by using tritiated thymidine. For more conclusive results labelled DNA incorporated in cytoplasmic organelles have to be traced during the embryo and endosperm development

  16. Cloning of transgenic tobacco BY-2 cells; an efficient method to analyse and reduce high natural heterogeneity of transgene expression.

    Science.gov (United States)

    Nocarova, Eva; Fischer, Lukas

    2009-04-22

    Phenotypic characterization of transgenic cell lines, frequently used in plant biology studies, is complicated because transgene expression in individual cells is often heterogeneous and unstable. To identify the sources and to reduce this heterogeneity, we transformed tobacco (Nicotiana tabacum L.) BY-2 cells with a gene encoding green fluorescent protein (GFP) using Agrobacterium tumefaciens, and then introduced a simple cloning procedure to generate cell lines derived from the individual transformed cells. Expression of the transgene was monitored by analysing GFP fluorescence in the cloned lines and also in lines obtained directly after transformation. The majority ( approximately 90%) of suspension culture lines derived from calli that were obtained directly from transformation consisted of cells with various levels of GFP fluorescence. In contrast, nearly 50% of lines generated by cloning cells from the primary heterogeneous suspensions consisted of cells with homogenous GFP fluorescence. The rest of the lines exhibited "permanent heterogeneity" that could not be resolved by cloning. The extent of fluorescence heterogeneity often varied, even among genetically identical clones derived from the primary transformed lines. In contrast, the offspring of subsequent cloning of the cloned lines was uniform, showing GFP fluorescence intensity and heterogeneity that corresponded to the original clone. The results demonstrate that, besides genetic heterogeneity detected in some lines, the primary lines often contained a mixture of epigenetically different cells that could be separated by cloning. This indicates that a single integration event frequently results in various heritable expression patterns, which are probably accidental and become stabilized in the offspring of the primary transformed cells early after the integration event. Because heterogeneity in transgene expression has proven to be a serious problem, it is highly advisable to use transgenes tagged with

  17. The NtAMI1 gene functions in cell division of tobacco BY-2 cells in the presence of indole-3-acetamide.

    Science.gov (United States)

    Nemoto, Keiichirou; Hara, Masamitsu; Suzuki, Masashi; Seki, Hikaru; Muranaka, Toshiya; Mano, Yoshihiro

    2009-01-22

    Tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) cells can be grown in medium containing indole-3-acetamide (IAM). Based on this finding, the NtAMI1 gene, whose product is functionally equivalent to the AtAMI1 gene of Arabidopsis thaliana and the aux2 gene of Agrobacterium rhizogenes, was isolated from BY-2 cells. Overexpression of the NtAMI1 gene allowed BY-2 cells to proliferate at lower concentrations of IAM, whereas suppression of the NtAMI1 gene by RNA interference (RNAi) caused severe growth inhibition in the medium containing IAM. These results suggest that IAM is incorporated into plant cells and converted to the auxin, indole-3-acetic acid, by NtAMI1.

  18. A Quantitative Assessment of the Morphofunctional Activity of the Population of Mast Cells Exposed to Biotechnological Strains of Microorganisms

    Directory of Open Access Journals (Sweden)

    Natalia Sheina

    2018-06-01

    Full Text Available In order to assess the sensitizing properties of bacteria, micromycetes and actinomycetes, the morphofunctional activity of the population of mast cells was tested in rats exposed to biotechnological microorganisms. The result showed the high informative value of the test of peritoneal must cell degranulation. Both the result and the intensity of the response of mast cells to the exposure to the tested strains depend on the taxonomy of microorganisms, their concentration and the mode of inoculation. The test of peritoneal must cell degranulation can be recommended for assessing the biological safety of industrial microorganisms.

  19. Analysis of Pseudomonas aeruginosa Cell Envelope Proteome by Capture of Surface-Exposed Proteins on Activated Magnetic Nanoparticles

    OpenAIRE

    Vecchietti, Davide; Di Silvestre, Dario; Miriani, Matteo; Bonomi, Francesco; Marengo, Mauro; Bragonzi, Alessandra; Cova, Lara; Franceschi, Eleonora; Mauri, Pierluigi; Bertoni, Giovanni

    2012-01-01

    We report on specific magneto-capturing followed by Multidimensional Protein Identification Technology (MudPIT) for the analysis of surface-exposed proteins of intact cells of the bacterial opportunistic pathogen Pseudomonas aeruginosa. The magneto-separation of cell envelope fragments from the soluble cytoplasmic fraction allowed the MudPIT identification of the captured and neighboring proteins. Remarkably, we identified 63 proteins captured directly by nanoparticles and 67 proteins embedde...

  20. Responses of well-differentiated nasal epithelial cells exposed to particles: Role of the epithelium in airway inflammation

    International Nuclear Information System (INIS)

    Auger, Floriane; Gendron, Marie-Claude; Chamot, Christophe; Marano, Francelyne; Dazy, Anne-Catherine

    2006-01-01

    Numerous epidemiological studies support the contention that ambient air pollution particles can adversely affect human health. To explain the acute inflammatory process in airways exposed to particles, a number of in vitro studies have been performed on cells grown submerged on plastic and poorly differentiated, and on cell lines, the physiology of which is somewhat different from that of well-differentiated cells. In order to obtain results using a model system in which epithelial cells are similar to those of the human airway in vivo, apical membranes of well-differentiated human nasal epithelial (HNE) cells cultured in an air-liquid interface (ALI) were exposed for 24 h to diesel exhaust particles (DEP) and Paris urban air particles (PM 2.5 ). DEP and PM 2.5 (10-80 μg/cm 2 ) stimulated both IL-8 and amphiregulin (ligand of EGFR) secretion exclusively towards the basal compartment. In contrast, there was no IL-1β secretion and only weak non-reproducible secretion of TNF-α. IL-6 and GM-CSF were consistently stimulated towards the apical compartment and only when cells were exposed to PM 2.5 . ICAM-1 protein expression on cell surfaces remained low after particle exposure, although it increased after TNF-α treatment. Internalization of particles, which is believed to initiate oxidative stress and proinflammatory cytokine expression, was restricted to small nanoparticles (≤ 40 nm). Production of reactive oxygen species (ROS) was detected, and DEP were more efficient than PM 2.5 . Collectively, our results suggest that airway epithelial cells exposed to particles augment the local inflammatory response in the lung but cannot alone initiate a systemic inflammatory response

  1. Apoptosis-inducing factor plays a critical role in caspase-independent, pyknotic cell death in hydrogen peroxide-exposed cells.

    Science.gov (United States)

    Son, Young-Ok; Jang, Yong-Suk; Heo, Jung-Sun; Chung, Wan-Tae; Choi, Ki-Choon; Lee, Jeong-Chae

    2009-06-01

    It has been proposed that continuously generated hydrogen peroxide (H(2)O(2)) inhibits typical apoptosis and instead initiates an alternate, apoptosis-inducing factor (AIF)-dependent process. Aside from the role of AIF, however, the detailed morphological characterization of H(2)O(2)-induced cell death is not complete. This study examined the cellular mechanism(s) by which the continuous presence of H(2)O(2) induces cell death. We also further analyzed the precise role of AIF by inhibiting its expression with siRNA. Exposure of cells to H(2)O(2) generated by glucose oxidase caused mitochondrion-mediated, caspase-independent cell death. In addition, H(2)O(2) exposure resulted in cell shrinkage and chromatin condensation without nuclear fragmentation, indicating that H(2)O(2) stimulates a pyknotic cell death. Further analysis of AIF-transfected cells clearly demonstrated that nuclear translocation of AIF is the most important event required for nuclear condensation, phosphatidyl serine translocation, and ultimately cell death in H(2)O(2)-exposed cells. Furthermore, ATP was rapidly and severely depleted in cells exposed to H(2)O(2) generated by glucose oxidase but not by H(2)O(2) added as a bolus. Suppression of the H(2)O(2)-mediated ATP depletion by 3-aminobenzamide led to a significant increase of nuclear fragmentation in glucose oxidase-exposed cells. Collectively, these findings suggest that an acute energy reduction by H(2)O(2) causes caspase-independent and AIF-dependent cell death.

  2. Potent antiproliferative cembrenoids accumulate in tobacco upon infection with Rhodococcus fascians and trigger unusual microtubule dynamics in human glioblastoma cells.

    Directory of Open Access Journals (Sweden)

    Aminata P Nacoulma

    Full Text Available AIMS: Though plant metabolic changes are known to occur during interactions with bacteria, these were rarely challenged for pharmacologically active compounds suitable for further drug development. Here, the occurrence of specific chemicals with antiproliferative activity against human cancer cell lines was evidenced in hyperplasia (leafy galls induced when plants interact with particular phytopathogens, such as the Actinomycete Rhodococcus fascians. METHODS: We examined leafy galls fraction F3.1.1 on cell proliferation, cell division and cytoskeletal disorganization of human cancer cell lines using time-lapse videomicroscopy imaging, combined with flow cytometry and immunofluorescence analysis. We determined the F3.1.1-fraction composition by gas chromatography coupled to mass spectrometry. RESULTS: The leafy galls induced on tobacco by R. fascians yielded fraction F3.1.1 which inhibited proliferation of glioblastoma U373 cells with an IC50 of 4.5 µg/mL, F.3.1.1 was shown to increase cell division duration, cause nuclear morphological deformations and cell enlargement, and, at higher concentrations, karyokinesis defects leading to polyploidization and apoptosis. F3.1.1 consisted of a mixture of isomers belonging to the cembrenoids. The cellular defects induced by F3.1.1 were caused by a peculiar cytoskeletal disorganization, with the occurrence of fragmented tubulin and strongly organized microtubule aggregates within the same cell. Colchicine, paclitaxel, and cembrene also affected U373 cell proliferation and karyokinesis, but the induced microtubule rearrangement was very different from that provoked by F3.1.1. Altogether our data indicate that the cembrenoid isomers in F3.1.1 have a unique mode of action and are able to simultaneously modulate microtubule polymerization and stability.

  3. Youth and Tobacco Use

    Science.gov (United States)

    ... past 30 days. † Any tobacco product includes cigarettes, cigars, smokeless tobacco (including chewing tobacco, snuff, dip, snus, and dissolvable tobacco), tobacco pipes, bidis, hookah, and electronic cigarettes. § Where percentages are missing, sample sizes were ...

  4. Transcriptome analysis of tobacco BY-2 cells elicited by cryptogein reveals new potential actors of calcium-dependent and calcium-independent plant defense pathways.

    Science.gov (United States)

    Amelot, Nicolas; Dorlhac de Borne, François; San Clemente, Hélène; Mazars, Christian; Grima-Pettenati, Jacqueline; Brière, Christian

    2012-02-01

    Cryptogein is a proteinaceous elicitor secreted by the oomycete Phytophthora cryptogea, which induces a hypersensitive response in tobacco plants. We have previously reported that in tobacco BY-2 cells treated with cryptogein, most of the genes of the phenylpropanoid pathway were upregulated and cell wall-bound phenolics accumulated. Both events were Ca(2+) dependent. In this study, we designed a microarray covering a large proportion of the tobacco genome and monitored gene expression in cryptogein-elicited BY-2 cells to get a more complete view of the transcriptome changes and to assess their Ca(2+) dependence. The predominant functional gene categories affected by cryptogein included stress- and disease-related proteins, phenylpropanoid pathway, signaling components, transcription factors and cell wall reinforcement. Among the 3819 unigenes whose expression changed more than fourfold, 90% were Ca(2+) dependent, as determined by their sensitivity to lanthanum chloride. The most Ca(2+)-dependent transcripts upregulated by cryptogein were involved in defense responses or the oxylipin pathway. This genome-wide study strongly supports the importance of Ca(2+)-dependent transcriptional regulation of regulatory and defense-related genes contributing to cryptogein responses in tobacco. Copyright © 2011 Elsevier Ltd. All rights reserved.

  5. Evaluation of Trace Elements in Augmentation of Statin-Induced Cytotoxicity in Uremic Serum-Exposed Human Rhabdomyosarcoma Cells

    Directory of Open Access Journals (Sweden)

    Hitoshi Uchiyama

    2018-01-01

    Full Text Available Patients with end-stage kidney disease (ESKD are at higher risk for rhabdomyolysis induced by statin than patients with normal kidney function. Previously, we showed that this increase in the severity of statin-induced rhabdomyolysis was partly due to uremic toxins. However, changes in the quantity of various trace elements in ESKD patients likely contribute as well. The purpose of this study is to determine the effect of trace elements on statin-induced toxicity in rhabdomyosarcoma cells exposed to uremic serum (US cells for a long time. Cell viability, apoptosis, mRNA expression, and intracellular trace elements were assessed by viability assays, flow cytometry, real-time RT-PCR, and ICP-MS, respectively. US cells exhibited greater simvastatin-induced cytotoxicity than cells long-time exposed with normal serum (NS cells (non-overlapping 95% confidence intervals. Intracellular levels of Mg, Mn, Cu, and Zn were significantly less in US cells compared to that in NS cells (p < 0.05 or 0.01. Pre-treatment with TPEN increased simvastatin-induced cytotoxicity and eliminated the distinction between both cells of simvastatin-induced cytotoxicity. These results suggest that Zn deficiencies may be involved in the increased risk for muscle complaints in ESKD patients. In conclusion, the increased severity of statin-induced rhabdomyolysis in ESKD patients may be partly due to trace elements deficiencies.

  6. Rifampicin sensitivity of residual RNA synthesis in Escherichia coli cells exposed to ultraviolet radiation and combined ultraviolet and γ radiations

    International Nuclear Information System (INIS)

    Prakash, R.K.; Netrawali, M.S.; Pradhan, D.S.

    1976-01-01

    UV-irradiation prevents rifampicin inhibition of the initiation of RNA synthesis by E.coli cells, but such rifampicin insensitivity is not exhibited by the residual RNA synthesis in γ-irradiated cells. Studies of the rate of [ 3 H]-uridine incorporation by E.coli cells at various times of incubation have been used to show that when γ-irradiation was given either before or after UV-irradiation of cells, the observed rifampicin insensitivity of residual RNA synthesis in the UV-irradiated cells was obliterated. RNA synthesis in cells subjected to combined exposures of UV- and γ-radiations was lowered to a lesser extent than that in the cells exposed to UV-irradiation alone. Possible mechanisms are discussed. (U.K.)

  7. Tetraploidy in monkey kidney epithelial cells exposed to various doses of radiation in vitro and in vivo. Comm.3

    International Nuclear Information System (INIS)

    Machavariani, M.G.

    1979-01-01

    The tetraploidy phenomenon in three and five day cultures of monkey kidney epithelial cells exposed to various doses of X-rays at Gsub(0) stage has been revealed. The data are presented on simple and complex tetraploidal enclo-reduplicated cells in monkey kidney epithelium after whole-body irradiaiton of animals by 60 Co γ-rays in dosage of 620-660 R. The frequency decrease of endoreduplicated cells at the second month coincides with the frequency increase of simple tetraploidal cells. In the investigated culture of monkey kidney epithelial cells, irradiated in vitro, a trend is observed towards the increase of the number of tetraploidal cells. An assumption is made on the possibility of using the frequency of tetraploidal cells ( including lymphocytes) for the purposes of biological dosimetry

  8. Rifampicin sensitivity of residual RNA synthesis in Escherichia coli cells exposed to ultraviolet radiation and combined ultraviolet and. gamma. radiations

    Energy Technology Data Exchange (ETDEWEB)

    Prakash, R K; Netrawali, M S; Pradhan, D S [Bhabha Atomic Research Centre, Bombay (India). Biochemistry and Food Technology Div.

    1976-09-01

    UV-irradiation prevents rifampicin inhibition of the initiation of RNA synthesis by E.coli cells, but such rifampicin insensitivity is not exhibited by the residual RNA synthesis in ..gamma..-irradiated cells. Studies of the rate of (/sup 3/H)-uridine incorporation by E.coli cells at various times of incubation have been used to show that when ..gamma.. irradiation was given either before or after uv-irradiation of cells, the observed rifampicin insensitivity of residual RNA synthesis in the uv-irradiated cells was obliterated. RNA synthesis in cells subjected to combined exposures of uv- and ..gamma..-radiations was lowered to a lesser extent than that in the cells exposed to uv-irradiation alone. Possible mechanisms are discussed.

  9. Enhancement of cell death by TNF α-related apoptosis-inducing ligand (TRAIL) in human lung carcinoma A549 cells exposed to X rays under hypoxia

    International Nuclear Information System (INIS)

    Takahashi, Momoko; Inanami, Osamu; Yasui, Hironobu; Ogura, Aki; Kuwabara, Mikinori; Kubota, Nobuo; Tsujitani, Michihiko

    2007-01-01

    Our previous study showed that ionizing radiation induced the expression of death receptor DR5 on the cell surface in tumor cell lines and that the death receptor of the TNF α-related apoptosis-inducing ligand TRAIL enhanced the apoptotic pathway (Hamasu et al., (2005) Journal of Radiation Research, 46:103-110). The present experiments were performed to examine whether treatment with TRAIL enhanced the cell killing in tumor cells exposed to ionizing radiation under hypoxia, since the presence of radioresistant cells in hypoxic regions of solid tumors is a serious problem in radiation therapy for tumors. When human lung carcinoma A549 cells were irradiated under normoxia and hypoxia, respectively, radiation-induced enhancement of expression of DR5 was observed under both conditions. Incubation in the presence of TRAIL enhanced the caspase-dependent and chymotrypsin-like-protease-dependent apoptotic cell death in A549 cells exposed to X rays. Furthermore, it was shown that treatment with TRAIL enhanced apoptotic cell death and loss of clonogenic ability in A549 cells exposed to X rays not only under normoxia but also under hypoxia, suggesting that combination treatment with TRAIL and X irradiation is effective for hypoxic tumor cells. (author)

  10. Relationship of CD86 surface marker expression and cytotoxicity on dendritic cells exposed to chemical allergen

    International Nuclear Information System (INIS)

    Hulette, Ben C.; Ryan, Cindy A.; Gildea, Lucy A.; Gerberick, G. Frank

    2005-01-01

    Human peripheral blood-derived dendritic cells (DC) respond to a variety of chemical allergens by up-regulating expression of the co-stimulatory molecule CD86. It has been postulated that this measure might provide the basis for an in vitro alternative approach for the identification of skin sensitizing chemicals. We recently reported that DC, exposed in culture to the highest non-cytotoxic concentrations of various chemical allergens, displayed marginal up-regulation of membrane CD86 expression; the interpretation being that such changes were insufficiently sensitive for the purposes of hazard identification. For the work presented here, immature DC were derived from human monocytes and treated with the chemical allergens 2,4-dinitrobenzenesulfonic acid (DNBS), nickel sulfate (NiSO 4 ), p-phenylenediamine (PPD), Bandrowski's base (BB), hydroquinone (HQ) and propyl gallate (PG) for 48 h at concentrations which induced both no to slight to moderate cytotoxicity. For comparison, DC were treated with the irritants sodium dodecyl sulfate (SDS), benzoic acid (BA), and benzalkonium chloride (BZC) at concentrations resulting in comparable levels of cytotoxicity. CD86 expression, as measured by flow cytometry, was consistently up-regulated (ranging from 162 to 386% control) on DC treated with concentrations of chemical allergens that induced approximately 10-15% cytotoxicity. The irritants BA and BZC did not induce up-regulation of CD86 expression when tested at concentrations that induced similar levels of cytotoxicity. SDS, however, up-regulated CD86 expression to 125-138% of control in 2/4 preparations when tested at concentrations which induced similar toxicity. Our results confirm that chemical allergens up-regulate CD86 expression on blood-derived DC and illustrate further that up-regulation of CD86 surface marker expression is more robust when DC are treated with concentrations of chemical allergen that induce slight to moderate cytotoxicity

  11. Low CD4+ T-cell levels and B-cell apoptosis in vertically HIV-exposed noninfected children and adolescents.

    Science.gov (United States)

    Miyamoto, Maristela; Pessoa, Silvana D; Ono, Erika; Machado, Daisy M; Salomão, Reinaldo; Succi, Regina C de M; Pahwa, Savita; de Moraes-Pinto, Maria Isabel

    2010-12-01

    Lymphocyte subsets, activation markers and apoptosis were assessed in 20 HIV-exposed noninfected (ENI) children born to HIV-infected women who were or not exposed to antiretroviral (ARV) drugs during pregnancy and early infancy. ENI children and adolescents were aged 6-18 years and they were compared to 25 age-matched healthy non-HIV-exposed children and adolescents (Control). ENI individuals presented lower CD4(+) T cells/mm(3) than Control group (control: 1120.3 vs. ENI: 876.3; t-test, p = 0.030). ENI individuals had higher B-cell apoptosis than Control group (Control: 36.6%, ARV exposed: 82.3%, ARV nonexposed: 68.5%; Kruskal-Wallis, p ENI and in Control children and adolescents. Subtle long-term immune alterations might persist among ENI individuals, but the clinical consequences if any are unknown, and these children require continued monitoring.

  12. The signal peptide-like segment of hpaXm is required for its association to the cell wall in transgenic tobacco plants.

    Science.gov (United States)

    Li, Le; Miao, Weiguo; Liu, Wenbo; Zhang, Shujian

    2017-01-01

    Harpins, encoded by hrp (hypersensitive response and pathogenicity) genes of Gram-negative plant pathogens, are elicitors of hypersensitive response (HR). HpaXm is a novel harpin-like protein described from cotton leaf blight bacteria, Xanthomonas citri subsp. malvacearum-a synonym of X. campestris pv. malvacearum (Smith 1901-1978). A putative signal peptide (1-MNSLNTQIGANSSFL-15) of hpaXm was predicted in the nitroxyl-terminal (N-terminal)by SignalP (SignalP 3.0 server). Here, we explored the function of the N-terminal leader peptide like segment of hpaXm using transgenic tobacco (Nicotiana tabacum cv. Xanthi nc.). Transgenic tobacco lines expressing the full-length hpaXm and the signal peptide-like segment-deleted mutant hpaXmΔLP were developed using transformation mediated by Agrobacterium tumefaciens. The target genes were confirmed integrated into the tobacco genomes and expressed normally. Using immune colloidal-gold detection technique, hpaXm protein was found to be transferred to the cytoplasm, the cell membrane, and organelles such as chloroplasts, mitochondria, and nucleus, as well as the cell wall. However, the deletion mutant hpaXmΔLP expressed in transgenic tobacco was found unable to cross the membrane to reach the cell wall. Additionally, soluble proteins extracted from plants transformed with hpaXm and hpaXmΔLP were bio-active. Defensive micro-HR induced by the transgene expression of hpaXm and hpaXmΔLP were observed on transgenic tobacco leaves. Disease resistance bioassays to tobacco mosaic virus (TMV) showed that tobacco plants transformed with hpaXm and with hpaXmΔLP exhibited enhanced resistance to TMV. In summary, the N-terminal signal peptide-like segment (1-45 bp) in hpaXm sequence is not necessary for transgene expression, bioactivity of hpaXm and resistance to TMV in transgenic tobacco, but is required for the protein to be translocated to the cell wall.

  13. Numerical study of induced current perturbations in the vicinity of excitable cells exposed to extremely low frequency magnetic fields

    International Nuclear Information System (INIS)

    Hassan, Noha; Chatterjee, Indira; Publicover, Nelson G; Craviso, Gale L

    2003-01-01

    Realistic three-dimensional cell morphologies were modelled to determine the current density induced in excitable cell culture preparations exposed to 60 Hz magnetic fields and to identify important factors that can influence the responses of cells to these fields. Cell morphologies representing single spherical adrenal chromaffin cells, single elongated smooth muscle cells and chromaffin cell aggregates in a Petri dish containing culture medium were modelled using the finite element method. The computations for a spherical cell revealed alterations in the magnitude and spatial distribution of the induced current density in the immediate vicinity of the cell. Maxima occurred at the equatorial sides and minima at the poles. Proximity of cells to each other as well as cell aggregate shape, size and orientation with respect to the induced current influenced the magnitude and spatial distribution of the induced current density. For an elongated cell, effects on the induced current density were highly dependent on cell orientation with respect to the direction of the induced current. These results provide novel insights into the perturbations in induced current that occur in excitable cell culture preparations and lay a foundation for understanding the mechanisms of interaction with extremely low frequency magnetic fields at the tissue level

  14. A multidirectional non-cell autonomous control and a genetic interaction restricting tobacco etch virus susceptibility in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Suresh Gopalan

    2007-10-01

    Full Text Available Viruses constitute a major class of pathogens that infect a variety of hosts. Understanding the intricacies of signaling during host-virus interactions should aid in designing disease prevention strategies and in understanding mechanistic aspects of host and pathogen signaling machinery.An Arabidopsis mutant, B149, impaired in susceptibility to Tobacco etch virus (TEV, a positive strand RNA virus of picoRNA family, was identified using a high-throughput genetic screen and a counterselection scheme. The defects include initiation of infection foci, rate of cell-to-cell movement and long distance movement.The defect in infectivity is conferred by a recessive locus. Molecular genetic analysis and complementation analysis with three alleles of a previously published mutant lsp1 (loss of susceptibility to potyviruses indicate a genetic interaction conferring haploinsufficiency between the B149 locus and certain alleles of lsp1 resulting in impaired host susceptibility. The pattern of restriction of TEV foci on leaves at or near the boundaries of certain cell types and leaf boundaries suggest dysregulation of a multidirectional non-cell autonomous regulatory mechanism. Understanding the nature of this multidirectional signal and the molecular genetic mechanism conferring it should potentially reveal a novel arsenal in the cellular machinery.

  15. USING PROTEOMICS TO MONITOR PROTEIN EXPRESSION IN HUMAN CELLS EXPOSED TO CARCINOGENS

    Science.gov (United States)

    People are continuously exposed exogenously to varying amounts of chemicals that have been shown to have carcinogenic properties in experimental systems. It has been estimated that exposure to environmental chemical carcinogens in the environment may contribute significantly to t...

  16. Environmental impact of heavy metals on the blood cells in professionally exposed workers

    OpenAIRE

    Velickova, Nevenka

    2017-01-01

    Aims of the study is to explain and research the effects of the heavy metals (lead, zinc and cadmium) on erythrocytes and leukocytes in miners with different work experience or exposure. The results and conclusions are made based on a three-year period of continuous testing on 120 miners, as professionally exposed workers. We confirmed that the miners long been professionally exposed to heavy metals, in the blood have an increased content of heavy metals (lead, zinc and cadmium) and they ha...

  17. The invasion of tobacco mosaic virus RNA induces endoplasmic reticulum stress-related autophagy in HeLa cells

    Science.gov (United States)

    Li, Li; Wang, Li; Xiao, Ruijing; Zhu, Guoguo; Li, Yan; Liu, Changxuan; Yang, Ru; Tang, Zhiqing; Li, Jie; Huang, Wei; Chen, Lang; Zheng, Xiaoling; He, Yuling; Tan, Jinquan

    2011-01-01

    The ability of human cells to defend against viruses originating from distant species has long been ignored. Owing to the pressure of natural evolution and human exploration, some of these viruses may be able to invade human beings. If their ‘fresh’ host had no defences, the viruses could cause a serious pandemic, as seen with HIV, SARS (severe acute respiratory syndrome) and avian influenza virus that originated from chimpanzees, the common palm civet and birds, respectively. It is unknown whether the human immune system could tolerate invasion with a plant virus. To model such an alien virus invasion, we chose TMV (tobacco mosaic virus) and used human epithelial carcinoma cells (HeLa cells) as its ‘fresh’ host. We established a reliable system for transfecting TMV-RNA into HeLa cells and found that TMV-RNA triggered autophagy in HeLa cells as shown by the appearance of autophagic vacuoles, the conversion of LC3-I (light chain protein 3-I) to LC3-II, the up-regulated expression of Beclin1 and the accumulation of TMV protein on autophagosomal membranes. We observed suspected TMV virions in HeLa cells by TEM (transmission electron microscopy). Furthermore, we found that TMV-RNA was translated into CP (coat protein) in the ER (endoplasmic reticulum) and that TMV-positive RNA translocated from the cytoplasm to the nucleolus. Finally, we detected greatly increased expression of GRP78 (78 kDa glucose-regulated protein), a typical marker of ERS (ER stress) and found that the formation of autophagosomes was closely related to the expanded ER membrane. Taken together, our data indicate that HeLa cells used ERS and ERS-related autophagy to defend against TMV-RNA. PMID:21729006

  18. Phosphatidylinositol (4,5)bisphosphate inhibits K+-efflux channel activity in NT1 tobacco cultured cells.

    Science.gov (United States)

    Ma, Xiaohong; Shor, Oded; Diminshtein, Sofia; Yu, Ling; Im, Yang Ju; Perera, Imara; Lomax, Aaron; Boss, Wendy F; Moran, Nava

    2009-02-01

    In the animal world, the regulation of ion channels by phosphoinositides (PIs) has been investigated extensively, demonstrating a wide range of channels controlled by phosphatidylinositol (4,5)bisphosphate (PtdInsP2). To understand PI regulation of plant ion channels, we examined the in planta effect of PtdInsP2 on the K+-efflux channel of tobacco (Nicotiana tabacum), NtORK (outward-rectifying K channel). We applied a patch clamp in the whole-cell configuration (with fixed "cytosolic" Ca2+ concentration and pH) to protoplasts isolated from cultured tobacco cells with genetically manipulated plasma membrane levels of PtdInsP2 and cellular inositol (1,4,5)trisphosphate: "Low PIs" had depressed levels of these PIs, and "High PIs" had elevated levels relative to controls. In all of these cells, K channel activity, reflected in the net, steady-state outward K+ currents (IK), was inversely related to the plasma membrane PtdInsP2 level. Consistent with this, short-term manipulations decreasing PtdInsP2 levels in the High PIs, such as pretreatment with the phytohormone abscisic acid (25 microM) or neutralizing the bath solution from pH 5.6 to pH 7, increased IK (i.e. NtORK activity). Moreover, increasing PtdInsP2 levels in controls or in abscisic acid-treated high-PI cells, using the specific PI-phospholipase C inhibitor U73122 (2.5-4 microM), decreased NtORK activity. In all cases, IK decreases stemmed largely from decreased maximum attainable NtORK channel conductance and partly from shifted voltage dependence of channel gating to more positive potentials, making it more difficult to activate the channels. These results are consistent with NtORK inhibition by the negatively charged PtdInsP2 in the internal plasma membrane leaflet. Such effects are likely to underlie PI signaling in intact plant cells.

  19. Over-expression of Trxo1 increases the viability of tobacco BY-2 cells under H2O2 treatment.

    Science.gov (United States)

    Ortiz-Espín, Ana; Locato, Vittoria; Camejo, Daymi; Schiermeyer, Andreas; De Gara, Laura; Sevilla, Francisca; Jiménez, Ana

    2015-09-01

    Reactive oxygen species (ROS), especially hydrogen peroxide, play a critical role in the regulation of plant development and in the induction of plant defence responses during stress adaptation, as well as in plant cell death. The antioxidant system is responsible for controlling ROS levels in these processes but redox homeostasis is also a key factor in plant cell metabolism under normal and stress situations. Thioredoxins (Trxs) are ubiquitous small proteins found in different cell compartments, including mitochondria and nuclei (Trxo1), and are involved in the regulation of target proteins through reduction of disulphide bonds, although their role under oxidative stress has been less well studied. This study describes over-expression of a Trxo1 for the first time, using a cell-culture model subjected to an oxidative treatment provoked by H2O2. Control and over-expressing PsTrxo1 tobacco (Nicotiana tabacum) BY-2 cells were treated with 35 mm H2O2 and the effects were analysed by studying the growth dynamics of the cultures together with oxidative stress parameters, as well as several components of the antioxidant systems involved in the metabolism of H2O2. Analysis of different hallmarks of programmed cell death was also carried out. Over-expression of PsTrxo1 caused significant differences in the response of TBY-2 cells to high concentrations of H2O2, namely higher and maintained viability in over-expressing cells, whilst the control line presented a severe decrease in viability and marked indications of oxidative stress, with generalized cell death after 3 d of treatment. In over-expressing cells, an increase in catalase activity, decreases in H2O2 and nitric oxide contents and maintenance of the glutathione redox state were observed. A decreased content of endogenous H2O2 may be responsible in part for the delayed cell death found in over-expressing cells, in which changes in oxidative parameters and antioxidants were less extended after the oxidative

  20. Gene expression profiling and pathway analysis of human bronchial epithelial cells exposed to airborne particulate matter collected from Saudi Arabia

    International Nuclear Information System (INIS)

    Sun, Hong; Shamy, Magdy; Kluz, Thomas; Muñoz, Alexandra B.; Zhong, Mianhua; Laulicht, Freda; Alghamdi, Mansour A.; Khoder, Mamdouh I.; Chen, Lung-Chi; Costa, Max

    2012-01-01

    Epidemiological studies have established a positive correlation between human mortality and increased concentration of airborne particulate matters (PM). However, the mechanisms underlying PM related human diseases, as well as the molecules and pathways mediating the cellular response to PM, are not fully understood. This study aims to investigate the global gene expression changes in human cells exposed to PM 10 and to identify genes and pathways that may contribute to PM related adverse health effects. Human bronchial epithelial cells were exposed to PM 10 collected from Saudi Arabia for 1 or 4 days, and whole transcript expression was profiled using the GeneChip human gene 1.0 ST array. A total of 140 and 230 genes were identified that significantly changed more than 1.5 fold after PM 10 exposure for 1 or 4 days, respectively. Ingenuity Pathway Analysis revealed that different exposure durations triggered distinct pathways. Genes involved in NRF2-mediated response to oxidative stress were up-regulated after 1 day exposure. In contrast, cells exposed for 4 days exhibited significant changes in genes related to cholesterol and lipid synthesis pathways. These observed changes in cellular oxidative stress and lipid synthesis might contribute to PM related respiratory and cardiovascular disease. -- Highlights: ► PM exposure modulated gene expression and associated pathways in BEAS-2B cells. ► One-day exposure to PM induced genes involved in responding to oxidative stress. ► 4-day exposure to PM changed genes associated to cholesterol and lipid synthesis.

  1. Phenotypic malignant changes and untargeted lipidomic analysis of long-term exposed prostate cancer cells to endocrine disruptors

    International Nuclear Information System (INIS)

    Bedia, Carmen; Dalmau, Núria; Jaumot, Joaquim; Tauler, Romà

    2015-01-01

    Endocrine disruptors (EDs) are a class of environmental toxic molecules able to interfere with the normal hormone metabolism. Numerous studies involve EDs exposure to initiation and development of cancers, including prostate cancer. In this work, three different EDs (aldrin, aroclor 1254 and chlorpyrifos (CPF)) were investigated as potential inducers of a malignant phenotype in DU145 prostate cancer cells after a chronic exposure. Epithelial to mesenchymal transition (EMT) induction, proliferation, migration, colony formation and release of metalloproteinase 2 (MMP-2) were analyzed in 50-day exposed cells to the selected EDs. As a result, aldrin and CPF exposure led to an EMT induction (loss of 16% and 14% of E-cadherin levels, respectively, compared to the unexposed cells). Aroclor and CPF presented an increased migration (134% and 126%, respectively), colony formation (204% and 144%, respectively) and MMP-2 release (137% in both cases) compared to the unexposed cells. An untargeted lipidomic analysis was performed to decipher the lipids involved in the observed transformations. As general results, aldrin exposure showed a global decrease in phospholipids and sphingolipids, and aroclor and CPF showed an increase of certain phospholipids, glycosphingolipids as well as a remarkable increase of some cardiolipin species. Furthermore, the three exposures resulted in an increase of some triglyceride species. In conclusion, some significant changes in lipids were identified and thus we postulate that some lipid compounds and lipid metabolic pathways could be involved in the acquisition of the malignant phenotype in exposed prostate cancer cells to the selected EDs. - Highlights: • Aldrin, aroclor and chlorpyrifos induced an aggressive phenotype in DU145 cells. • An untargeted lipidomic analysis has been performed on chronic exposed cells. • Lipidomic results showed changes in specific lipid species under chronic exposure. • These lipids may have a role in the

  2. Phenotypic malignant changes and untargeted lipidomic analysis of long-term exposed prostate cancer cells to endocrine disruptors

    Energy Technology Data Exchange (ETDEWEB)

    Bedia, Carmen, E-mail: carmen.bedia@idaea.csic.es; Dalmau, Núria, E-mail: nuria.dalmau@idaea.csic.es; Jaumot, Joaquim, E-mail: joaquim.jaumot@idaea.csic.es; Tauler, Romà, E-mail: roma.tauler@idaea.csic.es

    2015-07-15

    Endocrine disruptors (EDs) are a class of environmental toxic molecules able to interfere with the normal hormone metabolism. Numerous studies involve EDs exposure to initiation and development of cancers, including prostate cancer. In this work, three different EDs (aldrin, aroclor 1254 and chlorpyrifos (CPF)) were investigated as potential inducers of a malignant phenotype in DU145 prostate cancer cells after a chronic exposure. Epithelial to mesenchymal transition (EMT) induction, proliferation, migration, colony formation and release of metalloproteinase 2 (MMP-2) were analyzed in 50-day exposed cells to the selected EDs. As a result, aldrin and CPF exposure led to an EMT induction (loss of 16% and 14% of E-cadherin levels, respectively, compared to the unexposed cells). Aroclor and CPF presented an increased migration (134% and 126%, respectively), colony formation (204% and 144%, respectively) and MMP-2 release (137% in both cases) compared to the unexposed cells. An untargeted lipidomic analysis was performed to decipher the lipids involved in the observed transformations. As general results, aldrin exposure showed a global decrease in phospholipids and sphingolipids, and aroclor and CPF showed an increase of certain phospholipids, glycosphingolipids as well as a remarkable increase of some cardiolipin species. Furthermore, the three exposures resulted in an increase of some triglyceride species. In conclusion, some significant changes in lipids were identified and thus we postulate that some lipid compounds and lipid metabolic pathways could be involved in the acquisition of the malignant phenotype in exposed prostate cancer cells to the selected EDs. - Highlights: • Aldrin, aroclor and chlorpyrifos induced an aggressive phenotype in DU145 cells. • An untargeted lipidomic analysis has been performed on chronic exposed cells. • Lipidomic results showed changes in specific lipid species under chronic exposure. • These lipids may have a role in the

  3. Production and excision of thymine damage in the DNA of mammalian cells exposed to high-LET radiations

    International Nuclear Information System (INIS)

    Mattern, M.R.; Welch, G.P.

    1979-01-01

    HeLa S3 and Chinese hamster ovary cells were irradiated with high doses of carbon ions having linear energy transfers (LETs) of 170 and 780 keV/μm. The DNA was analyzed for 5,6-dihydroxydihydrothymine (t'-type) radiation products both before and after postirradiation incubation at 37 0 C. In HeLa cells, 2.1 x 10 -5 ring-damaged thymines were produced per kilorad per 10 6 daltons after irradiation with high-LET carbon ions - approximately one-fifth the efficiency of t' formation in HeLa cells exposed to low-LET x rays. t' products were also formed less efficiently in Chinese hamster ovary cells exposed to carbon ions than in those exposed to x rays. In both cell lines, up to 80% of the t' formed initially was excised selectively from the DNA during 60 min of postirradiation incubation at 37 0 C. Product excision was accompanied by small amounts of DNA degradation (less than 1%). Radiation with LET of 170 keV/μm - nearly the most effective LET for cell killing and the generation of unrejoined DNA strand breaks - produced ring-damaged thymines that were removed selectively from the DNA. This result is consistent with the conclusion that t'-type products do not contribute substantially to lethality after high-LET irradiation, although the alternative possibilities remain that t' is not excised as efficiently after biological doses, or that a particular subclass of t' or defective postexcision events contribute to cell killing

  4. Induction of chromosomal aberrations in human primary fibroblasts and immortalized cancer cells exposed to extremely-low-frequency electromagnetic fields

    International Nuclear Information System (INIS)

    Seyyedi, S. S.; Mozdarani, H.; Rezaei Tavirani, M.; Heydari, S.

    2010-01-01

    Rapidly increasing possibilities of exposure to environmental extremely low-frequency electromagnetic fields have become a topic of worldwide investigation. Epidemiological and laboratory studies suggest that exposure to extremely low-frequency electromagnetic fields may increase cancer risk therefore assessment of chromosomal damage in various cell lines might be of predictive value for future risk estimation. Materials and Methods: Primary cultures of fibroblasts from human skin biopsy were exposed to continuous extremely low-frequency electromagnetic fields (3, 50 and 60 Hz, sinusoidal, 3h, and 4 m T). Also immortalized cell lines, SW480, MCF-7 and 1321N1 were exposed to continuous extremely low-frequency electromagnetic fields (50 Hz, sinusoidal, 3 h, 4 m T). Metaphase plates Were prepared according to standard methods and stained in 5% Giemsa solution. Chromosomal aberrations of both chromosome and chromatid types were scored to evaluate the effects of extremely low-frequency electromagnetic fields on primary or established cell lines. Results: Results indicate that by increasing the frequency of extremely low-frequency electromagnetic fields, chromosomal aberrations were increased up to 7-fold above background levels in primary human fibroblast cells. In addition, continuous exposure to a 50 Hz electromagnetic field led to a significant increase in chromosomal aberrations in SW480, MCF-7 and 1321N1 cell lines compared to sham control. Conclusion: Results obtained indicate that extremely low-frequency electromagnetic fields has the potential for induction of chromosomal aberrations in all cell types.

  5. Mutagenic adaptive response to high-LET radiation in human lymphoblastoid cells exposed to X-rays.

    Science.gov (United States)

    Varès, Guillaume; Wang, Bing; Tanaka, Kaoru; Kakimoto, Ayana; Eguchi-Kasai, Kyomi; Nenoi, Mitsuru

    2011-01-10

    The ability of cells to adapt low-dose or low-dose rate radiation is well known. High-LET radiation has unique characteristics, and the data concerning low doses effects and high-LET radiation remain fragmented. In this study, we assessed in vitro the ability of low doses of X-rays to induce an adaptive response (AR) to a subsequent challenging dose of heavy-ion radiation. Lymphoblastoid cells (TK6, AHH-1, NH32) were exposed to priming 0.02-0.1Gy X-rays, followed 6h later by challenging 1Gy heavy-ion radiation (carbon-ion: 20 and 40keV/μm, neon-ion: 150keV/μm). Pre-exposure of p53-competent cells resulted in decreased mutation frequencies at hypoxanthine-guanine phosphoribosyl transferase locus and different H2AX phosphorylation kinetics, as compared to cells exposed to challenging radiation alone. This phenomenon did not seem to be linked with cell cycle effects or radiation-induced apoptosis. Taken together, our results suggested the existence of an AR to mutagenic effects of heavy-ion radiation in lymphoblastoid cells and the involvement of double-strand break repair mechanisms. Copyright © 2010 Elsevier B.V. All rights reserved.

  6. Cell viability, reactive oxygen species, apoptosis, and necrosis in myoblast cultures exposed to low-level infrared laser.

    Science.gov (United States)

    Alexsandra da Silva Neto Trajano, Larissa; da Silva, Camila Luna; de Carvalho, Simone Nunes; Cortez, Erika; Mencalha, André Luiz; de Souza da Fonseca, Adenilson; Stumbo, Ana Carolina

    2016-07-01

    Low-level infrared laser is considered safe and effective for treatment of muscle injuries. However, the mechanism involved on beneficial effects of laser therapy are not understood. The aim was to evaluate cell viability, reactive oxygen species, apoptosis, and necrosis in myoblast cultures exposed to low-level infrared laser at therapeutic fluences. C2C12 myoblast cultures at different (2 and 10 %) fetal bovine serum (FBS) concentrations were exposed to low-level infrared laser (808 nm, 100 mW) at different fluences (10, 35, and 70 J/cm(2)) and evaluated after 24, 48, and 72 h. Cell viability was evaluated by WST-1 assay; reactive oxygen species (ROS), apoptosis, and necrosis were evaluated by flow cytometry. Cell viability was decreased atthe lowest FBS concentration. Laser exposure increased the cell viability in myoblast cultures at 2 % FBS after 48 and 72 h, but no significant increase in ROS was observed. Apoptosis was decreased at the higher fluence and necrosis was increased at lower fluence in myoblast cultures after 24 h of laser exposure at 2 % FBS. No laser-induced alterations were obtained at 10 % FBS. Results show that level of reactive oxygen species is not altered, at least to those evaluated in this study, but low-level infrared laser exposure affects cell viability, apoptosis, and necrosis in myoblast cultures depending on laser fluence and physiologic conditions of cells.

  7. Sucrose Synthase Is Associated with the Cell Wall of Tobacco Pollen Tubes

    NARCIS (Netherlands)

    Persia, D.; Cai, G.; Casino, C.; Willemse, M.T.M.; Cresti, M.

    2008-01-01

    Sucrose synthase (Sus; EC 2.4.1.13) is a key enzyme of sucrose metabolism in plant cells, providing carbon for respiration and for the synthesis of cell wall polymers and starch. Since Sus is important for plant cell growth, insights into its structure, localization, and features are useful for

  8. Kinetics of hemopoietic stem cells and survival of mice treated with hydroxyurea and exposed to prolonged γ-radiation

    International Nuclear Information System (INIS)

    Chertkov, K.S.; Rogozkin, V.D.; Dikovenko, E.A.; Mosina, Z.M.

    1979-01-01

    A study was made of radioprotective efficiency of hydroxyurea in relation to mice exposed to prolonged 137 Cs-γ-radiation. It was found that a 30-day survival rate, under optimal conditions of treatment with hydroxyurea, was more than 40 per cent higher than that of the controls. The protective effect of hydroxyurea was manifested at the level of hemopoietic stem cells due to a quicker onset and accelerated rate of the repopulation process

  9. Tobacco BY-2 cell-free lysate: an alternative and highly-productive plant-based in vitro translation system.

    Science.gov (United States)

    Buntru, Matthias; Vogel, Simon; Spiegel, Holger; Schillberg, Stefan

    2014-05-03

    Cell-free protein synthesis is a rapid and efficient method for the production of recombinant proteins. Usage of prokaryotic cell-free extracts often leads to non-functional proteins. Eukaryotic counterparts such as wheat germ extract (WGE) and rabbit reticulocyte lysate (RLL) may improve solubility and promote the correct folding of eukaryotic multi-domain proteins that are difficult to express in bacteria. However, the preparation of WGEs is complex and time-consuming, whereas RLLs suffer from low yields. Here we report the development of a novel cell-free system based on tobacco Bright Yellow 2 (BY-2) cells harvested in the exponential growth phase. The highly-productive BY-2 lysate (BYL) can be prepared quickly within 4-5 h, compared to 4-5 d for WGE. The efficiency of the BYL was tested using three model proteins: enhanced yellow fluorescent protein (eYFP) and two versions of luciferase. The added mRNA was optimized by testing different 5' and 3' untranslated regions (UTRs). The protein yield in batch and dialysis reactions using BYL was much higher than that of a commercial Promega WGE preparation, achieving a maximum yield of 80 μg/mL of eYFP and 100 μg/mL of luciferase, compared to only 45 μg/mL of eYFP and 35 μg/mL of luciferase in WGEs. In dialysis reactions, the BYL yielded about 400 μg/mL eYFP, representing up to 50% more of the target protein than the Promega WGE, and equivalent to the amount using 5Prime WGE system. Due to the high yield and the short preparation time the BYL represents a remarkable improvement over current eukaryotic cell-free systems.

  10. An isoform of myosin XI is responsible for the translocation of endoplasmic reticulum in tobacco cultured BY-2 cells.

    Science.gov (United States)

    Yokota, Etsuo; Ueda, Shunpei; Tamura, Kentaro; Orii, Hidefumi; Uchi, Satoko; Sonobe, Seiji; Hara-Nishimura, Ikuko; Shimmen, Teruo

    2009-01-01

    The involvement of myosin XI in generating the motive force for cytoplasmic streaming in plant cells is becoming evident. For a comprehensive understanding of the physiological roles of myosin XI isoforms, it is necessary to elucidate the properties and functions of each isoform individually. In tobacco cultured BY-2 cells, two types of myosins, one composed of 175 kDa heavy chain (175 kDa myosin) and the other of 170 kDa heavy chain (170 kDa myosin), have been identified biochemically and immunocytochemically. From sequence analyses of cDNA clones encoding heavy chains of 175 kDa and 170 kDa myosin, both myosins have been classified as myosin XI. Immunocytochemical studies using a polyclonal antibody against purified 175 kDa myosin heavy chain showed that the 175 kDa myosin is distributed throughout the cytoplasm as fine dots in interphase BY-2 cells. During mitosis, some parts of 175 kDa myosin were found to accumulate in the pre-prophase band (PPB), spindle, the equatorial plane of a phragmoplast and on the circumference of daughter nuclei. In transgenic BY-2 cells, in which an endoplasmic reticulum (ER)-specific retention signal, HDEL, tagged with green fluorescent protein (GFP) was stably expressed, ER showed a similar behaviour to that of 175 kDa myosin. Furthermore, this myosin was co-fractionated with GFP-ER by sucrose density gradient centrifugation. From these findings, it was suggested that the 175 kDa myosin is a molecular motor responsible for translocating ER in BY-2 cells.

  11. DAF-fluorescence without NO: elicitor treated tobacco cells produce fluorescing DAF-derivatives not related to DAF-2 triazol.

    Science.gov (United States)

    Rümer, Stefan; Krischke, Markus; Fekete, Agnes; Mueller, Martin J; Kaiser, Werner M

    2012-08-15

    Diaminofluorescein-dyes (DAFs) are widely used for visualizing NO· production in biological systems. Here it was examined whether DAF-fluorescence could be evoked by other means than nitrosation. Tobacco (Nicotiana tabacum) suspension cells treated with the fungal elicitor cryptogein released compound(s) which gave a fluorescence increase in the cell-free filtrate after addition of DAF-2 or DAF-FM or DAR-4M. DAF-reactive compounds were relatively stable and identified as reaction products of H(2)O(2) plus apoplastic peroxidase (PO). CPTIO prevented formation of these products. Horseradish-peroxidase (HR-PO) plus H(2)O(2) also generated DAF-fluorescence in vitro. Using RP-HPLC with fluorescence detection, DAF derivatives were further analyzed. In filtrates from cryptogein-treated cells, fluorescence originated from two novel DAF-derivatives also obtained in vitro with DAF-2+HR-PO+H(2)O(2). DAF-2T was only detected when an NO donor (DEA-NO) was present. Using high resolution mass spectrometry, the two above-described novel DAF-reaction products were tentatively identified as dimers. In cells preloaded with DAF-2 DA and incubated with or without cryptogein, DAF-fluorescence originated from a complex pattern of multiple products different from those obtained in vitro. One specific peak was responsive to exogenous H(2)O(2), and another, minor peak eluted at or close to DAF-2T. Thus, in contrast to the prevailing opinion, DAF-2 can be enzymatically converted into a variety of highly fluorescing derivatives, both inside and outside cells, of which none (outside) or only a minor part (inside) appeared NO· dependent. Accordingly, DAF-fluorescence and its prevention by cPTIO do not necessarily indicate NO· production. Copyright © 2012 Elsevier Inc. All rights reserved.

  12. Recovery of tobacco BY-2 cells after high hydrostatic pressure treatment.

    Science.gov (United States)

    Kusube, Masataka; Nishino, Takumi; Nishikawa, Yuki; Goto, Masaki; Matsuki, Hitoshi; Iwahashi, Hitoshi

    2010-02-01

    The recovery of Nicotiana tabacum L. cv. Bright Yellow 2 (BY-2) cells in Linsmaire and Skoog medium after treatment at high hydrostatic pressure was investigated using an Evans Blue staining method to discriminate live from dead cells. The survival of BY-2 cells just after the high-pressure treatment at 5 degrees C and 25 degrees C decreased abruptly at pressures higher than 50 MPa and 100 MPa, respectively. Furthermore, almost all of the BY-2 cells treated at 5 degrees C and 25 degrees C recovered pressures below 25 MPa and 75 MPa, respectively. However, no BY-2 cells recovered at pressures above 100 MPa at either temperature.

  13. Antioxidant Activity of Ixora parviflora in a Cell/Cell-Free System and in UV-Exposed Human Fibroblasts

    Directory of Open Access Journals (Sweden)

    Hsiu-Mei Chiang

    2011-07-01

    Full Text Available Polyphenols and flavonoids possess a variety of biological activities including antioxidant and anti-tumor activities. Ixora parviflora is a member of the flavonoid-rich Rubiaceae family of flowering plants and used as folk medicine in India. The aim of this study was to investigate the antioxidant activity of Ixora parviflora extract (IPE in a cell-free system and erythrocytes, and the ability of IPE to inhibit reactive oxygen species (ROS generation in human fibroblasts (Hs68 after ultraviolet (UV exposure. Various in vitro antioxidant assays were employed in this study. The extraction yield of IPE was 17.4 ± 3.9%, the total phenolic content of IPE was 26.2 μg gallic acid equivalent (GAE/mg leaves dry weight and the total flavonoids content was 54.2 ± 4.4 μg quercetin equvalent (QE/mg extract. The content of chlorogenic acid was 9.7 ± 1.2 mg/g extract. IPE at 1000 μg/mL exhibited a reducing capacity of 90.5 ± 0.6%, a 1,1-diphenyl-2-picrylhydrazy (DPPH radical scavenging activity of 96.0 ± 0.4%, a ferrous chelating activity of 72.2 ± 3.5%, a hydroxyl radical scavenging activity of 96.8 ± 1.4%, and a hydrogen peroxide scavenging activity of 99.5 ± 3.3%. IPE at 500 μg/mL also possessed inhibitory activity against 2,2'-azobis (2-methylpropionamidine dihydrochloride (AAPH-induced hemolysis of erythrocytes (89.4 ± 1.8% and resulted in a 52.9% reduction in ROS generation in UV-exposed fibroblasts. According to our findings, IPE is a potent antioxidant and a potential anti-photoaging agent.

  14. Transcriptome-wide analysis of jasmonate-treated BY-2 cells reveals new transcriptional regulators associated with alkaloid formation in tobacco.

    Science.gov (United States)

    Yang, Yuping; Yan, Pengcheng; Yi, Che; Li, Wenzheng; Chai, Yuhui; Fei, Lingling; Gao, Ping; Zhao, Heping; Wang, Yingdian; Timko, Michael P; Wang, Bingwu; Han, Shengcheng

    2017-08-01

    Jasmonates (JAs) are well-known regulators of stress, defence, and secondary metabolism in plants, with JA perception triggering extensive transcriptional reprogramming, including both activation and/or repression of entire metabolic pathways. We performed RNA sequencing based transcriptomic profiling of tobacco BY-2 cells before and after treatment with methyl jasmonate (MeJA) to identify novel transcriptional regulators associated with alkaloid formation. A total of 107,140 unigenes were obtained through de novo assembly, and at least 33,213 transcripts (31%) encode proteins, in which 3419 transcription factors (TFs) were identified, representing 72 gene families, as well as 840 transcriptional regulators (TRs) distributed among 19 gene families. After MeJA treatment BY-2 cells, 7260 differentially expressed transcripts were characterised, which include 4443 MeJA-upregulated and 2817 MeJA-downregulated genes. Of these, 227 TFs/TRs in 36 families were specifically upregulated, and 102 TFs/TRs in 38 families were downregulated in MeJA-treated BY-2 cells. We further showed that the expression of 12 ethylene response factors and four basic helix-loop-helix factors increased at the transcriptional level after MeJA treatment in BY-2 cells and displayed specific expression patterns in nic mutants with or without MeJA treatments. Our data provide a catalogue of transcripts of tobacco BY-2 cells and benefit future study of JA-modulated regulation of secondary metabolism in tobacco. Copyright © 2017 Elsevier GmbH. All rights reserved.

  15. LASER CAPTURE MICRODISSECTION AND GENE ARRAY ANALYSIS OF PALATAL EPITHELIAL AND MESENCHYMAL CELLS EXPOSED TO TCDD

    Science.gov (United States)

    Palatal shelves from embryos exposed on gestation day (GD) 12 to either retinoic acid (RA) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) contact but fail to fuse. It is of interest to know if diverse agents that induce clefting via the same etiology also activate the same biochem...

  16. Oxygen enhancement ratios in synchronous HeLa cells exposed to low-LET radiation

    International Nuclear Information System (INIS)

    Sapozink, M.D.

    1977-01-01

    HeLa cells were synchronized by the mitotic selection method and rendered hypoxic by coincubation with an excess of heavily irradiated, but metabolically active, feeder cells. An oxygen enhancement ratio (OER) of about 3 was obtained in interphase HeLa cells irradiated with x or gamma rays. A significantly lower OER was obtained with cells in, or close to, mitosis. The significance of this decrease in the oxygen effect in mitotic cells is discussed

  17. Smokeless tobacco use among adolescents in Ilala Municipality ...

    African Journals Online (AJOL)

    Smokeless tobacco use is a significant part of the overall world tobacco problem. When the habit is introduced early in life, it increases the chance for permanent addiction and primes adolescents for use of harder drugs, exposing them to higher risk of oral cancer and other adverse effects of tobacco. This baseline study ...

  18. Polonium-210 in tobacco

    International Nuclear Information System (INIS)

    Harley, N.H.; Cohen, B.S.; Tso, T.C.

    1978-01-01

    The purpose of this paper was to present the measurements that have been made on tobacco and tobacco products and to indicate the studies that show the amount of 210 Po transferred to mainstream smoke and inhaled. The amounts reported to be in the lung are summarized. The authors have shown what average values might be expected in the lung due to normal deposition and clearance of the smoke aerosol and to compare these values with the measurements. The average dose to cells in the bronchial epithelium was estimated for the activities reported to be on the bronchial surface, and a comparison of this dose with a known tumorigenic alpha dose was made

  19. Oxidative stress and apoptosis induction in human thyroid carcinoma cells exposed to the essential oil from Pistacia lentiscus aerial parts.

    Directory of Open Access Journals (Sweden)

    Simona Catalani

    Full Text Available Essential oils from the aerial parts (leaves, twigs and berries of Pistacia lentiscus (PLEO have been well characterized for their antibacterial and anti-inflammatory properties; however, poor information exists on their potential anticancer activity.Increasing concentrations of PLEO (0.01-0.1% v/v, 80-800 μg/ml were administered to a wide variety of cultured cancer cells from breast, cervix, colon, liver, lung, prostate, and thyroid carcinomas. Fibroblasts were also included as healthy control cells. Cell viability was monitored by WST-8 assay up to 72 hours after PLEO administration. The intracellular formation of reactive oxygen species (ROS, the induction of apoptosis, and the enhancement of chemotherapeutic drug cytotoxicity by PLEO were further investigated in the most responsive cancer cell line.A dose-dependent reduction of tumor cell viability was observed upon PLEO exposure; while no cytotoxic effect was revealed in healthy fibroblasts. FTC-133 thyroid cancer cells were found to be the most sensitive cells to PLEO treatment; accordingly, an intracellular accumulation of ROS and an activation of both the extrinsic and intrinsic apoptotic pathways were evidenced in FTC-133 cells after PLEO administration. Furthermore, the cytotoxic effect of the antineoplastic drugs cisplatin, 5-fluorouracil and etoposide was enhanced in PLEO-exposed FTC-133 cells.Taking into account its mode of action, PLEO might be considered as a promising source of natural antitumor agents which might have therapeutic potential in integrated oncology.

  20. Oxidative stress and apoptosis induction in human thyroid carcinoma cells exposed to the essential oil from Pistacia lentiscus aerial parts.

    Science.gov (United States)

    Catalani, Simona; Palma, Francesco; Battistelli, Serafina; Benedetti, Serena

    2017-01-01

    Essential oils from the aerial parts (leaves, twigs and berries) of Pistacia lentiscus (PLEO) have been well characterized for their antibacterial and anti-inflammatory properties; however, poor information exists on their potential anticancer activity. Increasing concentrations of PLEO (0.01-0.1% v/v, 80-800 μg/ml) were administered to a wide variety of cultured cancer cells from breast, cervix, colon, liver, lung, prostate, and thyroid carcinomas. Fibroblasts were also included as healthy control cells. Cell viability was monitored by WST-8 assay up to 72 hours after PLEO administration. The intracellular formation of reactive oxygen species (ROS), the induction of apoptosis, and the enhancement of chemotherapeutic drug cytotoxicity by PLEO were further investigated in the most responsive cancer cell line. A dose-dependent reduction of tumor cell viability was observed upon PLEO exposure; while no cytotoxic effect was revealed in healthy fibroblasts. FTC-133 thyroid cancer cells were found to be the most sensitive cells to PLEO treatment; accordingly, an intracellular accumulation of ROS and an activation of both the extrinsic and intrinsic apoptotic pathways were evidenced in FTC-133 cells after PLEO administration. Furthermore, the cytotoxic effect of the antineoplastic drugs cisplatin, 5-fluorouracil and etoposide was enhanced in PLEO-exposed FTC-133 cells. Taking into account its mode of action, PLEO might be considered as a promising source of natural antitumor agents which might have therapeutic potential in integrated oncology.

  1. Immunostimulatory Activity of the Cytokine-Based Biologic, IRX-2, on Human Papillomavirus-Exposed Langerhans Cells.

    Science.gov (United States)

    Da Silva, Diane M; Woodham, Andrew W; Naylor, Paul H; Egan, James E; Berinstein, Neil L; Kast, W Martin

    2016-05-01

    Langerhans cells (LCs) are the antigen-presenting cells of the epithelial layer and are responsible for initiating immune responses against skin and mucosa-invading viruses. Human papillomavirus (HPV)-mediated suppression of LC function is a crucial mechanism of HPV immune evasion, which can lead to persistent infection and development of several human cancers, including cervical, anal, and head and neck cancers. The cell-derived cytokine-based biologic, IRX-2, consists of multiple well-defined cytokines and is broadly active on various immune cell subsets. In this study, we investigated primary human LC activation after exposure to HPV16, followed by treatment with IRX-2 in vitro, and evaluated their subsequent ability to induce HPV16-specific T cells. In contrast to its activity on dendritic cells, HPV16 alone is not sufficient to induce phenotypic and functional activation of LCs. However, IRX-2 induces a significant upregulation of antigen presentation and costimulatory molecules, T helper 1 (Th1)-associated cytokine release, and chemokine-directed migration of LCs pre-exposed to HPV16. Furthermore, LCs treated with IRX-2 after HPV16 exposure induced CD8(+) T-cell responses against specific HLA-A*0201-binding HPV16 T-cell epitopes. The present study suggests that IRX-2 is an attractive immunomodulator for assisting the immune response in eradication of HPV-infected cells, thereby potentially preventing HPV-induced cancers.

  2. Abnormal regulation of DNA replication and increased lethality in ataxia telangiectasia cells exposed to carcinogenic agents

    International Nuclear Information System (INIS)

    Jaspers, N.G.; de Wit, J.; Regulski, M.R.; Bootsma, D.

    1982-01-01

    The effect of different carcinogenic agents on the rate of semiconservative DNA replication in normal and ataxia telangiectasis (AT) cells was investigated. The rate of DNA synthesis in all AT cell strains tested was depressed to a significantly lesser extent than in normal cells after exposure to X-rays under oxia or hypoxia or to bleomycin, agents to which AT cells are hypersensitive. In contrast, inhibition of DNA replication in normal human and AT cells was similar after treatment with some DNA-methylating agents or mitomycin C. Colony-forming ability of AT cells treated with these agents was not different from normal cells. Treatment with 4-nitroquinoline 1-oxide elicited a variable response in both AT and normal cell strains. In some strains, including those shown to be hypersensitive to the drug by other workers, the inhibition of DNA synthesis was more pronounced than in other cell strains, but no significant difference between AT and normal cells could be detected. The rejoining of DNA strand breaks induced by X-rays, measured by DNA elution techniques, occurred within l2 hr after treatment and could not be correlated with the difference in DNA synthesis inhibition in AT and normal cells. After low doses of X-rays, AT cells rejoined single-strand breaks slightly more slowly than did normal cells. The rate of DNA replication in X-irradiation AT and normal cells was not affected by nicotinamide, an inhibitor of poly(adenosine diphosphate ribose) synthesis. These data indicate that the diminished inhibition of DNA replication in carcinogen-treated AT cells (a) is a general characteristic of all AT cell strains, (b) correlates with AT cellular hypersensitivity, (c) is not directly caused by the bulk of the DNA strand breaks produced by carcinogenic agents, and (d) is not based on differences in the induction of poly(adenosine diphosphate ribose) synthesis between X-irradiated AT and normal cells

  3. Dose-rate effects on mammalian cells exposed to ionizing radiation

    International Nuclear Information System (INIS)

    Mitchell, J.B.

    1978-01-01

    The effect of irradiation on the life cycle and on cell survival was studied for a range of different dose rates. Log phase, plateau phase and synchronized cultures of different mammalian cells were used. Cell cycle redistribution during the radiation exposure was found to be a very important factor in determining the overall dose-rate effect for log phase and synchronized cells. In fact, cell cycle redistribution during the exposure, in some instances, resulted in a lower dose rate being more effective in cell killing per unit dose than a higher dose rate. For plateau phase cultures, where cell cycle times are greatly lengthened, the effects of redistribution in regard to cell killing was virtually eliminated. Both fed and unfed plateau phase cultures exhibited a dose-rate effect, but it was found that below dose rates of 154 rad/h there is no further loss in effectiveness

  4. Lucifer Yellow uptake by CHO cells exposed to magnetic and electric pulses

    OpenAIRE

    Miklavčič, Damijan; Towhidi, Leila; Firoozabadi, S. M. P.; Mozdarani, Hossein

    2015-01-01

    Background The cell membrane acts as a barrier that hinders free entrance of most hydrophilic molecules into the cell. Due to numerous applications in medicine, biology and biotechnology, the introduction of impermeant molecules into biological cells has drawn considerable attention in the past years. One of the most famous methods in this field is electroporation, in which electric pulses with high intensity and short duration are applied to the cells. The aim of our study was to investigate...

  5. Microtubule reorganization in tobacco BY-2 cells stably expressing GFP-MBD

    Science.gov (United States)

    Granger, C. L.; Cyr, R. J.

    2000-01-01

    Microtubule organization plays an important role in plant morphogenesis; however, little is known about how microtubule arrays transit from one organized state to another. The use of a genetically incorporated fluorescent marker would allow long-term observation of microtubule behavior in living cells. Here, we have characterized a Nicotiana tabacum L. cv. Bright Yellow 2 (BY-2) cell line that had been stably transformed with a gfp-mbd construct previously demonstrated to label microtubules (J. Marc et al., 1998, Plant Cell 10: 1927-1939). Fluorescence levels were low, but interphase and mitotic microtubule arrays, as well as the transitions between these arrays, could be observed in individual gfp-mbd-transformed cells. By comparing several attributes of transformed and untransformed cells it was concluded that the transgenic cells are not adversely affected by low-level expression of the transgene and that these cells will serve as a useful and accurate model system for observing microtubule reorganization in vivo. Indeed, some initial observations were made that are consistent with the involvement of motor proteins in the transition between the spindle and phragmoplast arrays. Our observations also support the role of the perinuclear region in nucleating microtubules at the end of cell division with a progressive shift of these microtubules and/or nucleating activity to the cortex to form the interphase cortical array.

  6. You(th) & Tobacco

    Science.gov (United States)

    ... Exposure is High in Multiunit Housing Smokeless Products Electronic Cigarettes Youth Tobacco Prevention Tobacco Products Tobacco Ingredient ... Performance Don’t get trapped. Nicotine in cigarettes, cigars, and spit tobacco is addictive. Nicotine narrows your ...

  7. Construction and identification of subtracted cDNA library in bone marrow cells of radon-exposed mice

    International Nuclear Information System (INIS)

    Li Jianxiang; Nie Jihua; Tong Jian; Fu Chunling; Zhou Jianwei

    2008-01-01

    Objective: To construct and identify subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation. Methods: Adult male BALB/c mice, weighing 18-22 g, were placed in a multi- functional radon chamber. One group of mice was exposed to radon up to the accumulative dose of 105 work level month (WLM). The control group of mice was housed in a room with an accumulative dose of 1 WLM. To construct a subtracted cDNA library enriched with differentially expressed genes, the SMART technique and the suppression subtractive hybridization were performed. The obtained forward and reverse cDNA fragments were directly inserted into pMD18-T vector and transformed into E. coli JM109. The inserting cDNA fragments were screened by the blue-and-white blot screening and nested PCR of bacterium liquid. Results: The 244 of 285 white bacteria clones obtained randomly were positive clones contained 100-1100 bp inserted cDNA fragments. Conclusions: The forward and reverse subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation is successfully constructed. (authors)

  8. Comparison of photovoltaic cell temperatures in modules operating with exposed and enclosed back surfaces

    Science.gov (United States)

    Namkoong, D.; Simon, F. F.

    1981-01-01

    Four different photovoltaic module designs were tested to determine the cell temperature of each design. The cell temperatures were compared to those obtained on identical design, using the same nominal operating cell temperature (NOCT) concept. The results showed that the NOCT procedure does not apply to the enclosed configurations due to continuous transient conditions. The enclosed modules had higher cell temperatures than the open modules, and insulated modules higher than the uninsulated. The severest performance loss - when translated from cell temperatures - 17.5 % for one enclosed, insulated module as a compared to that module mounted openly.

  9. Heat shock protein 70 modulates neural progenitor cells dynamics in human neuroblastoma SH-SY5Y cells exposed to high glucose content.

    Science.gov (United States)

    Salimi, Leila; Rahbarghazi, Reza; Jafarian, Vahab; Biray Avci, Çıgır; Goker Bagca, Bakiye; Pinar Ozates, Neslihan; Khaksar, Majid; Nourazarian, Alireza

    2018-01-18

    In the current experiment, detrimental effects of high glucose condition were investigated on human neuroblastoma cells. Human neuroblastoma cell line SH-SY5Y were exposed to 5, 40, and 70 mM glucose over a period of 72 h. Survival rate and the proliferation of cells were analyzed by MTT and BrdU incorporation assays. Apoptosis was studied by the assays of flow cytometry and PCR array. In order to investigate the trans-differentiation capacity of the cell into mature neurons, we used immunofluorescence imaging to follow NeuN protein level. The transcription level of HSP70 was shown by real-time PCR analysis. MMP-2 and -9 activities were shown by gelatin Zymography. According to data from MTT and BrdU incorporation assay, 70 mM glucose reduced cell viability and proliferation rate as compared to control (5 mM glucose) and cells treated with 40 mM glucose (P Cell exposure to 70 mM glucose had potential to induced apoptosis after 72 h (P SH-SY5Y cells to detrimental effects of high glucose condition during trans-differentiation into mature neuron-like cells. Real-time PCR analysis confirmed the expression of HSP70 in cells under high content glucose levels, demonstrating the possible cell compensatory response to an insulting condition (p control vs 70 mM group  cells being exposed to 70 mM glucose. High glucose condition could abrogate the dynamics of neural progenitor cells. The intracellular level of HSP70 was proportional to cell damage in high glucose condition. © 2018 Wiley Periodicals, Inc.

  10. Telomere elongation protects heart and lung tissue cells from fatal damage in rats exposed to severe hypoxia.

    Science.gov (United States)

    Wang, Yaping; Zhao, Zhen; Zhu, Zhiyong; Li, Pingying; Li, Xiaolin; Xue, Xiaohong; Duo, Jie; Ma, Yingcai

    2018-02-17

    The effects of acute hypoxia at high altitude on the telomere length of the cells in the heart and lung tissues remain unclear. This study aimed to investigate the change in telomere length of rat heart and lung tissue cells in response to acute exposure to severe hypoxia and its role in hypoxia-induced damage to heart and lung tissues. Forty male Wistar rats (6-week old) were randomized into control group (n = 10) and hypoxia group (n = 30). Rats in control group were kept at an altitude of 1500 m, while rats in hypoxia group were exposed to simulated hypoxia with an altitude of 5000 m in a low-pressure oxygen chamber for 1, 3, and 7 days (n = 10). The left ventricular and right middle lobe tissues of each rat were collected for measurement of telomere length and reactive oxygen species (ROS) content, and the mRNA and protein levels of telomerase reverse transcriptase (TERT), hypoxia-inducible factor1α (HIF-1α), and hypoxia-inducible factor1α (HIF-2α). Increased exposure to hypoxia damaged rat heart and lung tissue cells and increased ROS production and telomere length. The mRNA and protein levels of TERT and HIF-1α were significantly higher in rats exposed to hypoxia and increased with prolonged exposure; mRNA and protein levels of HIF-2α increased only in rats exposed to hypoxia for 7 days. TERT was positively correlated with telomere length and the levels of HIF-1α but not HIF-2α. Acute exposure to severe hypoxia causes damage to heart and lung tissues due to the production of ROS but promotes telomere length and adaptive response by upregulating TERT and HIF-1α, which protect heart and lung tissue cells from fatal damage.

  11. Blot hybridization analysis of TCR genes of T cells for five people exposed in a radiation accident

    International Nuclear Information System (INIS)

    Min Rui; Liu Benti; Cheng Tianmin; Yang Rujun; Meng Xiangshun; Xiao Jinsong

    1996-01-01

    Human lymphocyte total DNA was prepared in agarose plug by mixing cells with low melting agarose, and two restriction endonucleases were used for digestion of the total DNA with human α and β TCR cDNA probes. The total digested DNA from five people who were whole body exposed to 2.0-2.5 Gy ionizing radiation in an accident 4.5 years ago was hybridized by Southern blot method. The results showed that no obvious difference in hybridization bands was found between controls and the five victims when hybridizations were fulfilled in the total DNA which was digested by Hind III restriction endonuclease with both α and β probes. However, when the total DNA was digested with restriction endonuclease EcoR I and was hybridized with TCR α probe, four of the five exposed people showed a different hybridizing band pattern compared with the controls. The results are also discussed

  12. A DP based scheme for real-time reconfiguration of solar cell arrays exposed to dynamic changing inhomogeneous illuminations

    DEFF Research Database (Denmark)

    Shi, Liping; Brehm, Robert

    2016-01-01

    The overall energy conversion efficiency of solar cell arrays is highly effected by partial shading effects. Especially for solar panel arrays installed in environments which are exposed to inhomogeneous dynamic changing illuminations such as on roof tops of electrical vehicles the overall system...... efficiency is drastically reduced. Dynamic real-time reconfiguration of the solar panel array can reduce effects on the output efficiency due to partial shading. This results in a maximized power output of the panel array when exposed to dynamic changing illuminations. The optimal array configuration...... with respect to shading patterns can be stated as a combinatorial optimization problem and this paper proposes a dynamic programming (DP) based algorithm which finds the optimal feasible solution to reconfigure the solar panel array for maximum efficiency in real-time with linear time complexity. It is shown...

  13. DNA damage in male gonad cells of Green mussel (Perna viridis) upon exposure to tobacco products

    Digital Repository Service at National Institute of Oceanography (India)

    Nagarajappa; Ganguly, A.; Goswami, U.

    DNA damage (determined by the Comet Assay) and the occurrence of deformed nuclei were measured as endpoints of genotoxicity in male gonad cells of the marine mussel (Perna viridis). Upon exposure of the organism to varying concentrations...

  14. FGF2 mediates DNA repair in epidermoid carcinoma cells exposed to ionizing radiation

    International Nuclear Information System (INIS)

    Marie, Melanie; Hafner, Sophie; Moratille, Sandra; Vaigot, Pierre; Rigaud, Odile; Martin, Michele T.; Mine, Solene

    2012-01-01

    Fibroblast growth factor 2 (FGF2) is a well-known survival factor. However, its role in DNA repair is poorly documented. The present study was designed to investigate in epidermoid carcinoma cells the potential role of FGF2 in DNA repair. The side population (SP) with cancer stem cell-like properties and the main population (MP) were isolated from human A431 squamous carcinoma cells. Radiation-induced DNA damage and repair were assessed using the alkaline comet assay. FGF2 expression was quantified by enzyme linked immunosorbent assay (ELISA). SP cells exhibited rapid repair of radiation induced DNA damage and a high constitutive level of nuclear FGF2. Blocking FGF2 signaling abrogated the rapid DNA repair. In contrast, in MP cells, a slower repair of damage was associated with low basal expression of FGF2. Moreover, the addition of exogenous FGF2 accelerated DNA repair in MP cells. When irradiated, SP cells secreted FGF2, whereas MP cells did not. FGF2 was found to mediate DNA repair in epidermoid carcinoma cells. We postulate that carcinoma stem cells would be intrinsically primed to rapidly repair DNA damage by a high constitutive level of nuclear FGF2. In contrast, the main population with a low FGF2 content exhibits a lower repair rate which can be increased by exogenous FGF2. (authors)

  15. Metabolomics reveals metabolic changes in male reproductive cells exposed to thirdhand smoke

    Science.gov (United States)

    Xu, Bo; Chen, Minjian; Yao, Mengmeng; Ji, Xiaoli; Mao, Zhilei; Tang, Wei; Qiao, Shanlei; Schick, Suzaynn F.; Mao, Jian-Hua; Hang, Bo; Xia, Yankai

    2015-10-01

    Thirdhand smoke (THS) is a new term for the toxins in cigarette smoke that linger in the environment long after the cigarettes are extinguished. The effects of THS exposure on male reproduction have not yet been studied. In this study, metabolic changes in male germ cell lines (GC-2 and TM-4) were analyzed after THS treatment for 24 h. THS-loaded chromatography paper samples were generated in a laboratory chamber system and extracted in DMEM. At a paper: DMEM ratio of 50 μg/ml, cell viability in both cell lines was normal, as measured by the MTT assay and markers of cytotoxicity, cell cycle, apoptosis and ROS production were normal as measured by quantitative immunofluorescence. Metabolomic analysis was performed on methanol extracts of GC-2 and TM-4 cells. Glutathione metabolism in GC-2 cells, and nucleic acid and ammonia metabolism in TM-4 cells, was changed significantly by THS treatment. RT-PCR analyses of mRNA for enzyme genes Gss and Ggt in GC-2 cells, and TK, SMS and Glna in TM-4 cells reinforced these findings, showing changes in the levels of enzymes involved in the relevant pathways. In conclusion, exposure to THS at very low concentrations caused distinct metabolic changes in two different types of male reproductive cell lines.

  16. Enhanced heterologous expression of biologically active human granulocyte colony stimulating factor in transgenic tobacco BY-2 cells by localization to endoplasmic reticulum.

    Science.gov (United States)

    Nair, Nisha R; Chidambareswaren, M; Manjula, S

    2014-09-01

    Tobacco Bright Yellow-2 (BY-2) cells, one of the best characterized cell lines is an attractive expression system for heterologous protein expression. However, the expression of foreign proteins is currently hampered by their low yield, which is partially the result of proteolytic degradation. Human granulocyte colony stimulating factor (hG-CSF) is a hematopoietic cytokine. Recombinant hG-CSF is successfully being used for the treatment of chemotherapy-induced neutropenia in cancer patients. Here, we describe a simple strategy for producing biologically active hG-CSF in tobacco BY-2 cells, localized in the apoplast of BY-2 cells, as well as targeted to the endoplasmic reticulum (ER). ER targeting significantly enhanced recombinant production which scaled to 17.89 mg/l from 4.19 mg/l when expressed in the apoplasts. Southern blotting confirmed the stable integration of hG-CSF in the BY-2 nuclear genome, and the expression of hG-CSF was analysed by Western blotting. Total soluble protein containing hG-CSF isolated from positive calli showed proliferative potential when tested on HL-60 cell lines by MTT assay. We also report the potential of a Fluorescence-activated cell sorting approach for an efficient sorting of the hG-CSF-expressing cell lines, which will enable the generation of homogenous high-producing cell lines.

  17. Effect of ATP sulfurylase overexpression in bright yellow 2 tobacco cells: regulation of ATP sulfurylase and SO4(-2) transport activities

    International Nuclear Information System (INIS)

    Hatzfeld, Y.; Cathala, N.; Grignon, C.; Davidian, J.C.

    1998-01-01

    To determine if the ATP sulfurylase reaction is a regulatory step for the SO4(2-)-assimilation pathway in plants, an Arabidopsis thaliana ATP sulfurylase cDNA, APS2, was fused to the 355 promoter of the cauliflower mosaic virus and introduced by Agrobacterium tumefaciens-mediated transformation into isolated Bright Yellow 2 tobacco (Nicotiana tabacum) cells. The ATP sulfurylase activity in transgenic cells was 8-fold that in control cells, and was correlated with the expression of a specific polypeptide revealed by western analysis using an anti-ATP sulfurylase antibody. The molecular mass of this polypeptide agreed with that for the overexpressed mature protein. ATP sulfurylase overexpression had no effect on [35S]SO4(2-) influx or ATP sulfurylase activity regulation by S availability, except that ATP sulfurylase activity variations in response to S starvation in transgenic cells were 8 times higher than in the wild type. There were also no differences in cell growth or sensitivity to SeO4(2-) (a toxic SO4(2-) analog) between transgenic and wild-type cells. We propose that in Bright Yellow 2 tobacco cells, the ATP sulfurylase derepression by S deficiency may involve a posttranscriptional mechanism, and that the ATP sulfurylase abundance is not limiting for cell metabolism

  18. Genotoxicity assessment data for exfoliated buccal cells exposed to mobile phone radiation

    Directory of Open Access Journals (Sweden)

    F.M. de Oliveira

    2017-12-01

    Full Text Available Healthy mobile phone users aged 18–30 y.o. provided exfoliated buccal cells samples from the right and left inner cheeks. A total of 2000 cells per subject were screened for the presence of micronuclei as a sign of genotoxic damage, according to the mobile phone use profile of each user. Keywords: Electromagnetic fields, Mobile phones, Genotoxicity, Micronuclei, Exfoliated buccal cells, Feulgen stain

  19. Diminished excretion of polyamines from BHK-21/C13 cells exposed to methylglyoxal bis(guanylhydrazone).

    Science.gov (United States)

    Melvin, M A; Keir, H M

    1978-01-01

    Methylglyoxal bis(guanylhydrazone) (1,1'-[methylethanediylidine)dinitrilo]diguanidine) inhibited the growth of BHK-21/C13 cells in monolayer cultures. Accumulation of spermidine and spermine was inhibited, whereas the accumulation of putrescine was increased. The intracellular spermidine/spermine molar ratio decreased conly slightly after exposure of the cells to 20 micrometer-methylglyoxal bis(guanylhydrazone) for 1 day. Cells incubated in the presence of the drug released less polyamine into the culture medium that did control cells, the polyamine released consisting almost exclusively of spermidine, both free and as a conjugated form. PMID:697761

  20. Structural damage of chicken red blood cells exposed to platinum nanoparticles and cisplatin

    DEFF Research Database (Denmark)

    Kutwin, Marta; Sawosz, Ewa; Jaworski, Sławomir

    2014-01-01

    of platinum nanoparticles (NP-Pt) and cisplatin with blood compartments are important for future applications. This study investigated structural damage, cell membrane deformation and haemolysis of chicken embryo red blood cells (RBC) after treatment with cisplatin and NP-Pt. Cisplatin (4 μg/ml) and NP-Pt (2......,6 μg/ml), when incubated with chicken embryo RBC, were detrimental to cell structure and induced haemolysis. The level of haemolytic injury was increased after cisplatin and NP-Pt treatments compared to the control group. Treatment with cisplatin caused structural damage to cell membranes...

  1. ATP Release from Human Airway Epithelial Cells Exposed to Staphylococcus aureus Alpha-Toxin

    Directory of Open Access Journals (Sweden)

    Romina Baaske

    2016-12-01

    Full Text Available Airway epithelial cells reduce cytosolic ATP content in response to treatment with S. aureus alpha-toxin (hemolysin A, Hla. This study was undertaken to investigate whether this is due to attenuated ATP generation or to release of ATP from the cytosol and extracellular ATP degradation by ecto-enzymes. Exposure of cells to rHla did result in mitochondrial calcium uptake and a moderate decline in mitochondrial membrane potential, indicating that ATP regeneration may have been attenuated. In addition, ATP may have left the cells through transmembrane pores formed by the toxin or through endogenous release channels (e.g., pannexins activated by cellular stress imposed on the cells by toxin exposure. Exposure of cells to an alpha-toxin mutant (H35L, which attaches to the host cell membrane but does not form transmembrane pores, did not induce ATP release from the cells. The Hla-mediated ATP-release was completely blocked by IB201, a cyclodextrin-inhibitor of the alpha-toxin pore, but was not at all affected by inhibitors of pannexin channels. These results indicate that, while exposure of cells to rHla may somewhat reduce ATP production and cellular ATP content, a portion of the remaining ATP is released to the extracellular space and degraded by ecto-enzymes. The release of ATP from the cells may occur directly through the transmembrane pores formed by alpha-toxin.

  2. Unscheduled DNA synthesis in spleen cells of mice exposed to low doses of total body irradiation

    International Nuclear Information System (INIS)

    Tuschl, H.; Kovac, R.; Hruby, E.

    1983-07-01

    Unscheduled DNA synthesis was induced by UV irradiation of spleen cells obtained from C 57 Bl mice after repeated total body irradiation of 0.05 Gy 60 Co (0.00125 Gy/mice) and determined autoradiographically. An enhancement in the ability for repair of UV induced DNA lesions was observed in cells of gamma irradiated animals. While the amount of 3 H-thymidine incorporated per cell was increased, the percentage of labeled cells remained unchanged. The present results are compared with previous data on low dose radiation exposure in men. (Author) [de

  3. Dehydration-induced WRKY genes from tobacco and soybean respond to jasmonic acid treatments in BY-2 cell culture.

    Science.gov (United States)

    Rabara, Roel C; Tripathi, Prateek; Lin, Jun; Rushton, Paul J

    2013-02-15

    Drought is one of the important environmental factors affecting crop production worldwide and therefore understanding the molecular response of plant to stress is an important step in crop improvement. WRKY transcription factors are one of the 10 largest transcription factor families across the green lineage. In this study, highly upregulated dehydration-induced WRKY and enzyme-coding genes from tobacco and soybean were selected from microarray data for promoter analyses. Putative stress-related cis-regulatory elements such as TGACG motif, ABRE-like elements; W and G-like sequences were identified by an in silico analyses of promoter region of the selected genes. GFP quantification of transgenic BY-2 cell culture showed these promoters direct higher expression in-response to 100 μM JA treatment compared to 100 μM ABA, 10% PEG and 85 mM NaCl treatments. Thus promoter activity upon JA treatment and enrichment of MeJA-responsive elements in the promoter of the selected genes provides insights for these genes to be jasmonic acid responsive with potential of mediating cross-talk during dehydration responses. Copyright © 2013 Elsevier Inc. All rights reserved.

  4. Analysis of Pseudomonas aeruginosa cell envelope proteome by capture of surface-exposed proteins on activated magnetic nanoparticles.

    Directory of Open Access Journals (Sweden)

    Davide Vecchietti

    Full Text Available We report on specific magneto-capturing followed by Multidimensional Protein Identification Technology (MudPIT for the analysis of surface-exposed proteins of intact cells of the bacterial opportunistic pathogen Pseudomonas aeruginosa. The magneto-separation of cell envelope fragments from the soluble cytoplasmic fraction allowed the MudPIT identification of the captured and neighboring proteins. Remarkably, we identified 63 proteins captured directly by nanoparticles and 67 proteins embedded in the cell envelope fragments. For a high number of proteins, our analysis strongly indicates either surface exposure or localization in an envelope district. The localization of most identified proteins was only predicted or totally unknown. This novel approach greatly improves the sensitivity and specificity of the previous methods, such as surface shaving with proteases that was also tested on P. aeruginosa. The magneto-capture procedure is simple, safe, and rapid, and appears to be well-suited for envelope studies in highly pathogenic bacteria.

  5. Analysis of Pseudomonas aeruginosa Cell Envelope Proteome by Capture of Surface-Exposed Proteins on Activated Magnetic Nanoparticles

    Science.gov (United States)

    Vecchietti, Davide; Di Silvestre, Dario; Miriani, Matteo; Bonomi, Francesco; Marengo, Mauro; Bragonzi, Alessandra; Cova, Lara; Franceschi, Eleonora; Mauri, Pierluigi; Bertoni, Giovanni

    2012-01-01

    We report on specific magneto-capturing followed by Multidimensional Protein Identification Technology (MudPIT) for the analysis of surface-exposed proteins of intact cells of the bacterial opportunistic pathogen Pseudomonas aeruginosa. The magneto-separation of cell envelope fragments from the soluble cytoplasmic fraction allowed the MudPIT identification of the captured and neighboring proteins. Remarkably, we identified 63 proteins captured directly by nanoparticles and 67 proteins embedded in the cell envelope fragments. For a high number of proteins, our analysis strongly indicates either surface exposure or localization in an envelope district. The localization of most identified proteins was only predicted or totally unknown. This novel approach greatly improves the sensitivity and specificity of the previous methods, such as surface shaving with proteases that was also tested on P. aeruginosa. The magneto-capture procedure is simple, safe, and rapid, and appears to be well-suited for envelope studies in highly pathogenic bacteria. PMID:23226459

  6. Anti-Inflammatory Effects of Pomegranate Peel Extract in THP-1 Cells Exposed to Particulate Matter PM10

    Directory of Open Access Journals (Sweden)

    Soojin Park

    2016-01-01

    Full Text Available Epidemiological and experimental evidence support health risks associated with the exposure to airborne particulate matter with a diameter of <10 μM (PM10. PM10 stimulates the production of reactive oxygen species (ROS and inflammatory mediators. Thus, we assumed that natural antioxidants might provide health benefits attenuating hazardous effects of PM10. In the present study, we examined the effects of pomegranate peel extract (PPE on THP-1 monocytic cells exposed to PM10. PM10 induced cytotoxicity and the production of ROS. It also increased the expression and secretion of inflammatory cytokines, such as tumor necrosis factor-α (TNF-α, interleukin-1β (IL-1β, and monocyte chemoattractant protein-1 (MCP-1, and cell adhesion molecules, such as intercellular adhesion molecule-1 (ICAM-1 and vascular cell adhesion molecule-1 (VCAM-1. PPE at 10–100 μg mL−1 attenuated the production of ROS and the expression of TNF-α, IL-1β, MCP-1, and ICAM-1, but not VCAM-1, in THP-1 cells stimulated by PM10 (100 μg mL−1. PPE also attenuated the adhesion of PM10-stimulated THP-1 cells to EA.hy926 endothelial cells. PPE constituents, punicalagin and ellagic acid, attenuated PM10-induced monocyte adhesion to endothelial cells, and punicalagin was less cytotoxic compared to ellagic acid. The present study suggests that PPE and punicalagin may be useful in alleviating inflammatory reactions due to particulate matter.

  7. Transcriptome analysis of the human T lymphocyte cell line Jurkat and human peripheral blood mononuclear cells exposed to deoxynivalenol (DON): New mechanistic insights

    Energy Technology Data Exchange (ETDEWEB)

    Katika, Madhumohan R. [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Department of Health Risk Analysis and Toxicology, Maastricht University (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Hendriksen, Peter J.M. [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Shao, Jia [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Department of Health Risk Analysis and Toxicology, Maastricht University (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Loveren, Henk van [Department of Health Risk Analysis and Toxicology, Maastricht University (Netherlands); National Institute for Public Health and the Environment (RIVM), Bilthoven (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Peijnenburg, Ad, E-mail: ad.peijnenburg@wur.nl [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Netherlands Toxicogenomics Centre (Netherlands)

    2012-10-01

    Deoxynivalenol (DON) or vomitoxin is a commonly encountered type-B trichothecene mycotoxin, produced by Fusarium species predominantly found in cereals and grains. DON is known to exert toxic effects on the gastrointestinal, reproductive and neuroendocrine systems, and particularly on the immune system. Depending on dose and exposure time, it can either stimulate or suppress immune function. The main objective of this study was to obtain a deeper insight into DON-induced effects on lymphoid cells. For this, we exposed the human T-lymphocyte cell line Jurkat and human peripheral blood mononuclear cells (PBMCs) to various concentrations of DON for various times and examined gene expression changes by DNA microarray analysis. Jurkat cells were exposed to 0.25 and 0.5 μM DON for 3, 6 and 24 h. Biological interpretation of the microarray data indicated that DON affects various processes in these cells: It upregulates genes involved in ribosome structure and function, RNA/protein synthesis and processing, endoplasmic reticulum (ER) stress, calcium-mediated signaling, mitochondrial function, oxidative stress, the NFAT and NF-κB/TNF-α pathways, T cell activation and apoptosis. The effects of DON on the expression of genes involved in ER stress, NFAT activation and apoptosis were confirmed by qRT-PCR. Other biochemical experiments confirmed that DON activates calcium-dependent proteins such as calcineurin and M-calpain that are known to be involved in T cell activation and apoptosis. Induction of T cell activation was also confirmed by demonstrating that DON activates NFATC1 and induces its translocation from the cytoplasm to the nucleus. For the gene expression profiling of PBMCs, cells were exposed to 2 and 4 μM DON for 6 and 24 h. Comparison of the Jurkat microarray data with those obtained with PBMCs showed that most of the processes affected by DON in the Jurkat cell line were also affected in the PBMCs. -- Highlights: ► The human T cell line Jurkat and human

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

    International Nuclear Information System (INIS)

    Suzuki, Katsura; Inageda, Kiyoshi; Nishitai, Gen; Matsuoka, Masato

    2007-01-01

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

  9. Low-level laser therapy: Effects on human face aged skin and cell viability of HeLa cells exposed to UV radiation

    Directory of Open Access Journals (Sweden)

    Mezghani Sana

    2015-01-01

    Full Text Available Chronic and excessive exposure to UV radiation leads to photoaging and photocarcinogenesis. Adequate protection of the skin against the deleterious effects of UV irradiation is essential. Low-level laser therapy (LLLT is a light source in the red to near-infrared range that has been accepted in a variety of medical applications. In this study, we explored the effect of LLLT in human face aged skin and the cell viability of HeLa cells exposed to UV radiation. We found that LLLT significantly reduced visible wrinkles and the loss of firmness of facial skin in aging subjects. Additionally, treatment of cultured HeLa cells with LLLT prior to or post UVA or UVB exposure significantly protected cells from UV-mediated cell death. All results showed the beneficial effects of LLLT on relieving signs of skin aging and its prevention and protection of the cell viability against UV-induced damage.

  10. Oxidative stress, mitochondrial permeability transition, and cell death in Cu-exposed trout hepatocytes

    International Nuclear Information System (INIS)

    Krumschnabel, Gerhard; Manzl, Claudia; Berger, Christian; Hofer, Bettina

    2005-01-01

    We have previously shown that, in trout hepatocytes, exposure to a high dose of copper (Cu) leads to disruption of Ca 2+ homeostasis and elevated formation of reactive oxygen species (ROS), with the latter ultimately causing cell death. In the present study, we aimed at identifying, using a lower Cu concentration, the role of mitochondria in this scenario, the potential involvement of the mitochondrial permeability transition (MPT), and the mode of cell death induced by the metal. Incubation with 10 μM Cu resulted in a strong stimulation of ROS formation, and after 2 h of exposure a significant increase of both apoptotic and necrotic cells was seen. Co-incubation of Cu-treated hepatocytes with the iron-chelator deferoxamine significantly inhibited ROS production and completely prevented cell death. The origin of the radicals generated was at least partly mitochondrial, as visualized by confocal laser scanning microscopy. Furthermore, ROS production was diminished by inhibition of mitochondrial respiration, but since this also aggravated the elevation of intracellular Ca 2+ induced by Cu, it did not preserve cell viability. In a sub-population of cells, Cu induced a decrease of mitochondrial membrane potential and occurrence of the MPT. Cyclosporin A, which did not inhibit ROS formation, prevented the onset of the MPT and inhibited apoptotic, but not necrotic, cell death. Cu-induced apoptosis therefore appears to be dependent on induction of the MPT, but the prominent contribution of mitochondria to ROS generation also suggests an important role of mitochondria in necrotic cell death

  11. Sensitivity of HTB140 cell exposed to protons and alkylating agents

    International Nuclear Information System (INIS)

    Koricanac, L.; Petrovic, I.; Privitera, G.; Cuttone, G.; Ristic-Fira, A.

    2006-01-01

    Malignant melanoma is a highly aggressive cancer with a poor prognosis due to resistance to radiotherapy and chemotherapy regimens. The mainstay of treatment remains DNA-alkylating agent dacarbazine (DTIC). Fotemustine (FM), chloroethylnitrosourea agent, also has demonstrated significant antitumoral effects in malignant melanoma. However, the resistance of melanoma cells limits their clinical application. In order to enhance the inhibition of melanoma cell growth, in this study, combined treatment of FM and DTIC with proton irradiation, was investigated. We analyzed the effects of combined treatment on HTB140 melanoma cell viability and proliferation. Significant inhibition of cell growth, especially cell proliferation, was obtained after treatment with protons and FM compare to single irradiation or drug treatment. Treatment with protons and DTIC has shown improved growth inhibition compare to appropriate single drug treatment, but not compare to irradiation as a single treatment. (author)

  12. Gene expression profiles of glucose toxicity-exposed islet microvascular endothelial cells.

    Science.gov (United States)

    Liu, Mingming; Lu, Wenbao; Hou, Qunxing; Wang, Bing; Sheng, Youming; Wu, Qingbin; Li, Bingwei; Liu, Xueting; Zhang, Xiaoyan; Li, Ailing; Zhang, Honggang; Xiu, Ruijuan

    2018-03-25

    Islet microcirculation is mainly composed by IMECs. The aim of the study was to investigate the differences in gene expression profiles of IMECs upon glucose toxicity exposure and insulin treatment. IMECs were treated with 5.6 mmol L -1 glucose, 35 mmol L -1 glucose, and 35 mmol L -1 glucose plus 10 -8  mol L -1 insulin, respectively. Gene expression profiles were determined by microarray and verified by qPCR. GO terms and KEGG analysis were performed to assess the potential roles of differentially expressed genes. The interaction and expression tendency of differentially expressed genes were analyzed by Path-Net algorithm. Compared with glucose toxicity-exposed IMECs, 1574 mRNAs in control group and 2870 mRNAs in insulin-treated IMECs were identified with differential expression, respectively. GO and KEGG pathway analysis revealed that these genes conferred roles in regulation of apoptosis, proliferation, migration, adhesion, and metabolic process etc. Additionally, MAPK signaling pathway and apoptosis were the dominant nodes in Path-Net. IMECs survival and function pathways were significantly changed, and the expression tendency of genes from euglycemia and glucose toxicity exposure to insulin treatment was revealed and enriched in 7 patterns. Our study provides a microcirculatory framework for gene expression profiles of glucose toxicity-exposed IMECs. © 2018 John Wiley & Sons Ltd.

  13. Conveying misinformation: Top-ranked Japanese books on tobacco.

    Science.gov (United States)

    Kanamori, Yuko; Malone, Ruth E

    2011-01-24

    Tobacco control efforts in Japan have lagged other high income countries, possibly because the Japanese government partially owns Japan Tobacco, Inc. In Japan, tobacco use is still often regarded as an issue of manners rather than an issue of health. Information about tobacco is available, but may not always be accurate. We explored what information Japanese consumers might access by reading popular Japanese books about tobacco. We searched Amazon.com Japan using the term "Tobacco", identifying the top 12 books by "relevance" and "bestselling." We eliminated duplicates and books not concerned with tobacco use and classified the remaining books as pro-smoking, anti-smoking, or neutral. We reviewed the pro-smoking books, published 2004-2009, and analyzed examples of misinformation by theme. Pro-smoking popular books conveyed five types of misinformation: doubt about science; suggestions that smoking increased health, longevity, virility, etc.; trivializing tobacco's effects; attacking public health advocates/authorities; and linking tobacco use with authenticity, history, or civil rights. At least one book was authored by a former Japan Tobacco employee; another used a popular Japan Tobacco advertising phrase. Creating doubt and confusion about tobacco serves tobacco industry interests and re-creates a strategy developed by US tobacco interests more than 40 years ago. Japanese readers may be misled by texts such as those reviewed. Tobacco control and public health advocates in Japan and globally should expose and counter such misinformation. "Naming and shaming" may be effective.

  14. Expression of Epstein-Barr virus among oral potentially malignant disorders and oral squamous cell carcinomas in the South Indian tobacco-chewing population.

    Science.gov (United States)

    Reddy, Sujatha S; Sharma, Shivani; Mysorekar, Vijaya

    2017-07-01

    Oral cancer is the sixth most common malignancy in the world. Viruses are the causative agents of approximately 10-15% of all cancers worldwide (Cancers, 6, 2014 and 2155). The tumorigenic roles of Epstein-Barr virus in oral cancer are unclear. Literature search results are conflicting and dependent on various factors such as geographical/regional variations, sociocultural lifestyles, dietary habits, chewing/smoking tobacco habit. This study is the first original observation about frequency of Epstein-Barr virus among South Indian tobacco-chewing patients to elucidate its involvement in oral carcinogenesis and to know whether this can be a valuable diagnostic and prognostic indicator. A total number of 75 tobacco chewer subjects aged between 23 and 76 years with histopathologically confirmed oral potentially malignant disorders (25), oral squamous cell carcinoma (25), and age-matched healthy controls (25) formed the study group. Immunohistochemical expression of Epstein-Barr virus latent membrane protein 1 was assessed among cases and healthy controls. Out of the total 75 subjects, six subjects (8%) were positive for Epstein-Barr virus antigen and 69 subjects (92%) negative. The antigen positivity was observed among two cases of moderately differentiated oral squamous cell carcinoma, two cases of leukoplakia, and two healthy controls. No significant association between Epstein-Barr virus positivity was observed among oral potentially malignant disorders and oral squamous cell carcinoma among South Indian tobacco-chewing patients. This can be partially explained by the methodology employed, by the patient population analyzed and different habits in various geographical regions. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  15. High Level of Tobacco Carcinogen-Derived DNA Damage in Oral Cells Is an Independent Predictor of Oral/Head and Neck Cancer Risk in Smokers.

    Science.gov (United States)

    Khariwala, Samir S; Ma, Bin; Ruszczak, Chris; Carmella, Steven G; Lindgren, Bruce; Hatsukami, Dorothy K; Hecht, Stephen S; Stepanov, Irina

    2017-09-01

    Exposure to tobacco-specific nitrosamines (TSNA) and polycyclic aromatic hydrocarbons (PAH) is recognized to play an important role in the development of oral/head and neck squamous cell cancer (HNSCC). We recently reported higher levels of TSNA-associated DNA adducts in the oral cells of smokers with HNSCC as compared with cancer-free smokers. In this study, we further investigated the tobacco constituent exposures in the same smokers to better understand the potential causes for the elevated oral DNA damage in smokers with HNSCC. Subjects included cigarette smokers with HNSCC (cases, n = 30) and cancer-free smokers (controls, n = 35). At recruitment, tobacco/alcohol use questionnaires were completed, and urine and oral cell samples were obtained. Analysis of urinary 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and N '-Nitrosonornicotine (NNN; TSNA biomarkers), 1-hydroxypyrene (1-HOP, a PAH), cotinine, 3'-hydroxycotinine, and the nicotine metabolite ratio (NMR) were performed. Cases and controls differed in mean age, male preponderance, and frequency of alcohol consumption (but not total alcoholic drinks). Univariate analysis revealed similar levels of NNN, 1-HOP, and cotinine between groups but, as reported previously, significantly higher DNA adduct formation in the cases. Multiple regression adjusting for potential confounders showed persistent significant difference in DNA adduct levels between cases and controls [ratio of geometric means, 20.0; 95% CI, 2.7-148.6). Our cohort of smokers with HNSCC demonstrates higher levels of TSNA-derived oral DNA damage in the setting of similar exposure to nicotine and tobacco carcinogens. Among smokers, DNA adduct formation may act as a predictor of eventual development of HNSCC that is independent of carcinogen exposure indicators. Cancer Prev Res; 10(9); 507-13. ©2017 AACR See related editorial by Johnson and Bauman, p. 489 . ©2017 American Association for Cancer Research.

  16. Biotransformation of isonitrosoacetophenone (2-keto-2-phenyl-acetaldoxime) in tobacco cell suspensions

    CSIR Research Space (South Africa)

    Madala, NE

    2012-07-01

    Full Text Available Nicotiana tabacum cell suspensions, 2g wet wt/ml, rapidly took up 1 mM isonitrosoacetophenone (INAP), a plant-derived stress metabolite with anti-oxidative and anti-fungal properties, producing 40-hexopyranosyloxy-30-methoxyisonitrosoacetophenone...

  17. Cytological changes and alterations in polyamine contents induced by cadmium in tobacco BY-2 cells

    Czech Academy of Sciences Publication Activity Database

    Kuthanová, A.; Gemperlová, Lenka; Zelenková, S.; Eder, Josef; Macháčková, Ivana; Opatrný, Z.; Cvikrová, Milena

    2004-01-01

    Roč. 42, č. 2 (2004), s. 149-156 ISSN 0981-9428 R&D Projects: GA MŠk LN00A081 Institutional research plan: CEZ:AV0Z5038910 Keywords : BY-2 cells * cadmium * DAO Subject RIV: EF - Botanics Impact factor: 1.414, year: 2004

  18. NtGNL1a ARF-GEF acts in endocytosis in tobacco cells

    Czech Academy of Sciences Publication Activity Database

    Jelínková, Adriana; Müller, Karel; Pařezová, Markéta; Petrášek, Jan

    2015-01-01

    Roč. 15, NOV 5 (2015), s. 272 ISSN 1471-2229 R&D Projects: GA ČR GPP305/11/P797 Institutional support: RVO:61389030 Keywords : Endocytosis * PIN1 protein trafficking * Inhibitors of endomembrane trafficking Subject RIV: EA - Cell Biology Impact factor: 3.631, year: 2015

  19. Comparative analysis of the surface exposed proteome of two canine osteosarcoma cell lines and normal canine osteoblasts.

    Science.gov (United States)

    Milovancev, Milan; Hilgart-Martiszus, Ian; McNamara, Michael J; Goodall, Cheri P; Seguin, Bernard; Bracha, Shay; Wickramasekara, Samanthi I

    2013-06-13

    Osteosarcoma (OSA) is the most common primary bone tumor of dogs and carries a poor prognosis despite aggressive treatment. An improved understanding of the biology of OSA is critically needed to allow for development of novel diagnostic, prognostic, and therapeutic tools. The surface-exposed proteome (SEP) of a cancerous cell includes a multifarious array of proteins critical to cellular processes such as proliferation, migration, adhesion, and inter-cellular communication. The specific aim of this study was to define a SEP profile of two validated canine OSA cell lines and a normal canine osteoblast cell line utilizing a biotinylation/streptavidin system to selectively label, purify, and identify surface-exposed proteins by mass spectrometry (MS) analysis. Additionally, we sought to validate a subset of our MS-based observations via quantitative real-time PCR, Western blot and semi-quantitative immunocytochemistry. Our hypothesis was that MS would detect differences in the SEP composition between the OSA and the normal osteoblast cells. Shotgun MS identified 133 putative surface proteins when output from all samples were combined, with good consistency between biological replicates. Eleven of the MS-detected proteins underwent analysis of gene expression by PCR, all of which were actively transcribed, but varied in expression level. Western blot of whole cell lysates from all three cell lines was effective for Thrombospondin-1, CYR61 and CD44, and indicated that all three proteins were present in each cell line. Semi-quantitative immunofluorescence indicated that CD44 was expressed at much higher levels on the surface of the OSA than the normal osteoblast cell lines. The results of the present study identified numerous differences, and similarities, in the SEP of canine OSA cell lines and normal canine osteoblasts. The PCR, Western blot, and immunocytochemistry results, for the subset of proteins evaluated, were generally supportive of the mass spectrometry data

  20. Silver ions increase plasma membrane permeability through modulation of intracellular calcium levels in tobacco BY-2 cells.

    Science.gov (United States)

    Klíma, Petr; Laňková, Martina; Vandenbussche, Filip; Van Der Straeten, Dominique; Petrášek, Jan

    2018-05-01

    Silver ions increase plasma membrane permeability for water and small organic compounds through their stimulatory effect on plasma membrane calcium channels, with subsequent modulation of intracellular calcium levels and ion homeostasis. The action of silver ions at the plant plasma membrane is largely connected with the inhibition of ethylene signalling thanks to the ability of silver ion to replace the copper cofactor in the ethylene receptor. A link coupling the action of silver ions and cellular auxin efflux has been suggested earlier by their possible direct interaction with auxin efflux carriers or by influencing plasma membrane permeability. Using tobacco BY-2 cells, we demonstrate here that besides a dramatic increase of efflux of synthetic auxins 2,4-dichlorophenoxyacetic acid (2,4-D) and 1-naphthalene acetic acid (NAA), treatment with AgNO 3 resulted in enhanced efflux of the cytokinin trans-zeatin (tZ) as well as the auxin structural analogues tryptophan (Trp) and benzoic acid (BA). The application of AgNO 3 was accompanied by gradual water loss and plasmolysis. The observed effects were dependent on the availability of extracellular calcium ions (Ca 2+ ) as shown by comparison of transport assays in Ca 2+ -rich and Ca 2+ -free buffers and upon treatment with inhibitors of plasma membrane Ca 2+ -permeable channels Al 3+ and ruthenium red, both abolishing the effect of AgNO 3 . Confocal microscopy of Ca 2+ -sensitive fluorescence indicator Fluo-4FF, acetoxymethyl (AM) ester suggested that the extracellular Ca 2+ availability is necessary to trigger the response to silver ions and that the intracellular Ca 2+ pool alone is not sufficient for this effect. Altogether, our data suggest that in plant cells the effects of silver ions originate from the primal modification of the internal calcium levels, possibly by their interaction with Ca 2+ -permeable channels at the plasma membrane.

  1. Myosin XI-dependent formation of tubular structures from endoplasmic reticulum isolated from tobacco cultured BY-2 cells.

    Science.gov (United States)

    Yokota, Etsuo; Ueda, Haruko; Hashimoto, Kohsuke; Orii, Hidefumi; Shimada, Tomoo; Hara-Nishimura, Ikuko; Shimmen, Teruo

    2011-05-01

    The reticular network of the endoplasmic reticulum (ER) consists of tubular and lamellar elements and is arranged in the cortical region of plant cells. This network constantly shows shape change and remodeling motion. Tubular ER structures were formed when GTP was added to the ER vesicles isolated from tobacco (Nicotiana tabacum) cultured BY-2 cells expressing ER-localized green fluorescent protein. The hydrolysis of GTP during ER tubule formation was higher than that under conditions in which ER tubule formation was not induced. Furthermore, a shearing force, such as the flow of liquid, was needed for the elongation/extension of the ER tubule. The shearing force was assumed to correspond to the force generated by the actomyosin system in vivo. To confirm this hypothesis, the S12 fraction was prepared, which contained both cytosol and microsome fractions, including two classes of myosins, XI (175-kD myosin) and VIII (BY-2 myosin VIII-1), and ER-localized green fluorescent protein vesicles. The ER tubules and their mesh-like structures were arranged in the S12 fraction efficiently by the addition of ATP, GTP, and exogenous filamentous actin. The tubule formation was significantly inhibited by the depletion of 175-kD myosin from the S12 fraction but not BY-2 myosin VIII-1. Furthermore, a recombinant carboxyl-terminal tail region of 175-kD myosin also suppressed ER tubule formation. The tips of tubules moved along filamentous actin during tubule elongation. These results indicated that the motive force generated by the actomyosin system contributes to the formation of ER tubules, suggesting that myosin XI is responsible not only for the transport of ER in cytoplasm but also for the reticular organization of cortical ER.

  2. Polyethylene glycol and polyvinylpirrolidone effect on bacterial rRNA extraction and hybridization from cells exposed to tannins.

    OpenAIRE

    ARCURI, P.B.; THONNEY, M.L.; SCHOFIELD, P.; PELL, A.N.

    2003-01-01

    In order to detect fluctuations in ruminal microbial populations due to forage tannins using 16S ribosomal RNA (rRNA) probes, recovery of intact rRNA is required. The objective of this work was to evaluate the effect of polyethylene glycol (PEG) and polyvinylpirrolidone (PVP) on extraction of bacterial rRNA, in the presence of tannins from tropical legume forages and other sources, that hybridize with oligonucleotide probes. Ruminococcus albus 8 cells were exposed to 8 g/L tannic acid or 1 g/...

  3. Transformation of Balb 3T3 cells exposed to a germicidal UV lamp and a sunlamp

    International Nuclear Information System (INIS)

    Withrow, T.J.; Lugo, M.H.; Dempsey, M.J.

    1980-01-01

    The effect of germicidal UV and sunlamp exposure on direct and simian virus-40 (SV-40) transformatioon of Balb 3T3 cells was studied. Transformation was determined by the ability of transformed cells to grow as clones in agar. Radiation from these lamps enhanced direct transformation, and enhanced viral transformation to approximately the same degree. Enhanced transformation was seen with exposures of light that caused no measurable cell killing, which suggests that the induction of new transformants is involved rather than the selection of pre-existing transformants. Induction is also suggested by post-irradiation growth kinetics experiments. (author)

  4. TOBACCO CONTROL

    International Development Research Centre (IDRC) Digital Library (Canada)

    Tobacco is farmed in more than 125 countries and the problems associated with this ... Canada's International Development Research Centre (IDRC) is one of the world's leading institutions in the generation and application of new ... assumptions about the relative safety ... In Kenya, researchers at Maseno University work.

  5. Rcan1-1L overexpression induces mitochondrial autophagy and improves cell survival in angiotensin II-exposed cardiomyocytes

    International Nuclear Information System (INIS)

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling

    2015-01-01

    Mitochondrial autophagy is an important adaptive stress response and can be modulated by various key molecules. A previous study found that the regulator of calcineurin 1-1L (Rcan1-1L) may regulate mitochondrial autophagy and cause mitochondria degradation in neurocytes. However, the effect of Rcan1-1L on cardiomyocytes has not been determined. In the present study, we aimed to investigate the role of Rcan1-1L in angiotensin II (Ang II)-exposed human cardiomyocytes. Above all, Human adult cardiac myocytes (HACMs) were exposed to 200 nmol/L Ang II for 4 days. Enhanced H 2 O 2 production, cytochrome C release and mitochondrial permeability were observed in these cells, which were blocked by valsartan. Consistently, Ang II exposure significantly reduced cardiomyocyte viability. However, transfection of Rcan1-1L vector promoted cell viability and ameliorated the apoptosis caused by Ang II. Rcan1-1L clearly promoted mitochondrial autophagy in HACMs, with elevated autophagy protein (ATG) 5 and light chain 3 (LC3) expression. Transient mitochondrial biogenesis and reduced cytochrome C release was also induced by Rcan1-1L. Additionally, Rcan1-1L significantly inhibited calcineurin/nuclear factor of activated T cells (NFAT) signaling. We thus conclude that Rcan1-1L may play a protective role in Ang II-treated cardiomyocytes through the induction of mitochondrial autophagy, and may be an alternative method of cardiac protection. - Highlights: • Transfection of Rcan1-1L into HACMs promoted cell viability and reduced apoptosis. • Transfection of Rcan1-1L promoted mitochondrial autophagy in HACMs. • Rcan1-1L inhibited the calcineurin/nuclear factor of activated T cells signaling

  6. Rcan1-1L overexpression induces mitochondrial autophagy and improves cell survival in angiotensin II-exposed cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling, E-mail: shanglingwang@126.com

    2015-07-01

    Mitochondrial autophagy is an important adaptive stress response and can be modulated by various key molecules. A previous study found that the regulator of calcineurin 1-1L (Rcan1-1L) may regulate mitochondrial autophagy and cause mitochondria degradation in neurocytes. However, the effect of Rcan1-1L on cardiomyocytes has not been determined. In the present study, we aimed to investigate the role of Rcan1-1L in angiotensin II (Ang II)-exposed human cardiomyocytes. Above all, Human adult cardiac myocytes (HACMs) were exposed to 200 nmol/L Ang II for 4 days. Enhanced H{sub 2}O{sub 2} production, cytochrome C release and mitochondrial permeability were observed in these cells, which were blocked by valsartan. Consistently, Ang II exposure significantly reduced cardiomyocyte viability. However, transfection of Rcan1-1L vector promoted cell viability and ameliorated the apoptosis caused by Ang II. Rcan1-1L clearly promoted mitochondrial autophagy in HACMs, with elevated autophagy protein (ATG) 5 and light chain 3 (LC3) expression. Transient mitochondrial biogenesis and reduced cytochrome C release was also induced by Rcan1-1L. Additionally, Rcan1-1L significantly inhibited calcineurin/nuclear factor of activated T cells (NFAT) signaling. We thus conclude that Rcan1-1L may play a protective role in Ang II-treated cardiomyocytes through the induction of mitochondrial autophagy, and may be an alternative method of cardiac protection. - Highlights: • Transfection of Rcan1-1L into HACMs promoted cell viability and reduced apoptosis. • Transfection of Rcan1-1L promoted mitochondrial autophagy in HACMs. • Rcan1-1L inhibited the calcineurin/nuclear factor of activated T cells signaling.

  7. DNA damage and apoptosis of endometrial cells cause loss of the early embryo in mice exposed to carbon disulfide

    International Nuclear Information System (INIS)

    Zhang, Bingzhen; Shen, Chunzi; Yang, Liu; Li, Chunhui; Yi, Anji; Wang, Zhiping

    2013-01-01

    Carbon disulfide (CS 2 ) may lead to spontaneous abortion and very early pregnancy loss in women exposed in the workplace, but the mechanism remains unclear. We designed an animal model in which gestating Kunming strain mice were exposed to CS 2 via i.p. on gestational day 4 (GD4). We found that the number of implanted blastocysts on GD8 was significantly reduced by each dose of 0.1 LD 50 (157.85 mg/kg), 0.2 LD 50 (315.7 mg/kg) and 0.4 LD 50 (631.4 mg/kg). In addition, both the level of DNA damage and apoptosis rates of endometrial cells on GD4.5 were increased, showed definite dose–response relationships, and inversely related to the number of implanted blastocysts. The expressions of mRNA and protein for the Bax and caspase-3 genes in the uterine tissues on GD4.5 were up-regulated, while the expressions of mRNA and protein for the Bcl-2 gene were dose-dependently down-regulated. Our results indicated that DNA damage and apoptosis of endometrial cells were important reasons for the loss of implanted blastocysts induced by CS 2 . - Highlights: • We built an animal model of CS2 exposure during blastocyst implantation. • Endometrial cells were used in the comet assay to detect DNA damage. • CS2 exposure caused DNA damage and endometrial cell apoptosis. • DNA damage and endometrial cell apoptosis were responsible for embryo loss

  8. DNA damage and apoptosis of endometrial cells cause loss of the early embryo in mice exposed to carbon disulfide

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Bingzhen [Department of Epidemiology and Health Statistics, School of Public Health, Shandong University, Jinan (China); Shen, Chunzi [Centers for Disease Control and Prevention, Zibo (China); Yang, Liu; Li, Chunhui; Yi, Anji [Department of Epidemiology and Health Statistics, School of Public Health, Shandong University, Jinan (China); Wang, Zhiping, E-mail: zhipingw@sdu.edu.cn [Department of Epidemiology and Health Statistics, School of Public Health, Shandong University, Jinan (China)

    2013-12-01

    Carbon disulfide (CS{sub 2}) may lead to spontaneous abortion and very early pregnancy loss in women exposed in the workplace, but the mechanism remains unclear. We designed an animal model in which gestating Kunming strain mice were exposed to CS{sub 2} via i.p. on gestational day 4 (GD4). We found that the number of implanted blastocysts on GD8 was significantly reduced by each dose of 0.1 LD{sub 50} (157.85 mg/kg), 0.2 LD{sub 50} (315.7 mg/kg) and 0.4 LD{sub 50} (631.4 mg/kg). In addition, both the level of DNA damage and apoptosis rates of endometrial cells on GD4.5 were increased, showed definite dose–response relationships, and inversely related to the number of implanted blastocysts. The expressions of mRNA and protein for the Bax and caspase-3 genes in the uterine tissues on GD4.5 were up-regulated, while the expressions of mRNA and protein for the Bcl-2 gene were dose-dependently down-regulated. Our results indicated that DNA damage and apoptosis of endometrial cells were important reasons for the loss of implanted blastocysts induced by CS{sub 2}. - Highlights: • We built an animal model of CS2 exposure during blastocyst implantation. • Endometrial cells were used in the comet assay to detect DNA damage. • CS2 exposure caused DNA damage and endometrial cell apoptosis. • DNA damage and endometrial cell apoptosis were responsible for embryo loss.

  9. Monitoring Intracellular Redox Changes in Ozone-exposed airway epithelial cells

    Science.gov (United States)

    Background: The toxicity of many compounds involves oxidative injury to cells. Direct assessment of mechanistic events involved in xenobiotic-induced oxidative stress is not easily achievable. Development of genetically-encoded probes designed for monitoring intracellular redox s...

  10. DNA repair in mammalian cells exposed to combinations of carcinogenic agents

    International Nuclear Information System (INIS)

    Setlow, R.B.; Ahmed, F.E.

    1979-01-01

    Cells defective in one or more aspects of repair are killed and often mutagenized more readily than normal cells by DNA damaging agents, and humans whose cells are deficient in repair are at an increased carcinogenic risk compared to normal individuals. The excision repair of uv induced pyrimidine dimers is a well studied system, but the details of the steps in this repair system are far from being understood in human cells. We know that there are a number of chemicals that mimic uv in that normal human cells repair DNA damage from both these agents and from uv by a long patch excision repair system, and that xeroderma pigmentosum cells defective in repair of uv are also defective in the repair of damage from these chemicals. The chemicals we have investigated are AAAF, 4-NQO, DMBA-epoxide, and ICR-170. We describe experiments, using several techniques, in which DNA excision repair is measured after treatment of various human cell strains with combinations of uv and these agents. If two agents have a common rate limiting step then, at doses high enough to saturate the repair system, one would expect the observed repair after a treatment with a combination of agents to be equal to that from one agent alone. Such is not the case for normal human or excision-deficient XP cells. In the former repair is additive and in the latter repair is usually appreciably less than that observed with either agent alone. Models that attempt to explain these surprising results involve complexes of enzymes and cofactors

  11. Ion transport in a human lens epithelial cell line exposed to hyposmotic and apoptotic stress.

    Science.gov (United States)

    Chimote, Ameet A; Adragna, Norma C; Lauf, Peter K

    2010-04-01

    Membrane transport changes in human lens epithelial (HLE-B3) cells under hyposmotic and apoptotic stress were compared. Cell potassium content, K(i), uptake of the K congener rubidium, Rb(i), and water content were measured after hyposmotic stress induced by hypotonicity, and apoptotic stress by the protein-kinase inhibitor staurosporine (STP). Cell water increased in hyposmotic (150 mOsm) as compared to isosmotic (300 mOsm) balanced salt solution (BSS) by >2-fold at 5 min and decreased within 15 min to baseline values accompanied by a 40% K(i) loss commensurate with cell swelling and subsequent cell shrinkage likely due to regulatory volume decrease (RVD). Loss of K(i), and accompanying water, and Rb(i) uptake in hyposmotic BSS were prevented by clotrimazole (CTZ) suggesting water shifts associated with K and Rb flux via intermediate conductance K (IK) channels, also detected at the mRNA and protein level. In contrast, 2 h after 2 microM STP exposure, the cells lost approximately 40% water and approximately 60% K(i), respectively, consistent with apoptotic volume decrease (AVD). Indeed, water and K(i) loss was at least fivefold greater after hyposmotic than after apoptotic stress. High extracellular K and 2 mM 4-aminopyridine (4-AP) but not CTZ significantly reduced apoptosis. Annexin labeling phosphatidylserine (PS) at 15 min suggested loss of lipid asymmetry. Quantitative PCR revealed significant IK channel expression during prolonged hyposmotic stress. Results suggest in HLE-B3 cells, IK channels likely partook in and were down regulated after RVD, whereas pro-apoptotic STP-activation of 4-AP-sensitive voltage-gated K channels preceded or accompanied PS externalization before subsequent apoptosis. J. Cell. Physiol. 223: 110-122, 2010. (c) 2009 Wiley-Liss, Inc.

  12. Influence of shear stress and size on viability of endothelial cells exposed to gold nanoparticles

    Science.gov (United States)

    Fede, C.; Albertin, Giovanna; Petrelli, L.; De Caro, R.; Fortunati, I.; Weber, V.; Ferrante, Camilla

    2017-09-01

    Screening nanoparticle toxicity directly on cell culture can be a fast and cheap technique. Nevertheless, to obtain results in accordance with those observed in live animals, the conditions in which cells are cultivated should resemble the one encountered in live systems. Microfluidic devices offer the possibility to satisfy this requirement, in particular with endothelial cell lines, because they are capable to reproduce the flowing media and shear stress experienced by these cell lines in vivo. In this work, we exploit a microfluidic device to observe how human umbilical vein endothelial cells (HUVEC) viability changes when subject to a continuous flow of culture medium, in which spherical citrate-stabilized gold nanoparticles of different sizes and at varying doses are investigated. For comparison, the same experiments are also run in multiwells where the cells do not experience the shear stress induced by the flowing medium. We discuss the results considering the influence of mode of exposure and nanoparticle size (24 and 13 nm). We observed that gold nanoparticles show a lower toxicity under flow conditions with respect to static and the HUVEC viability decreases as the nanoparticle surface area per unit volume increases, regardless of size.

  13. Effects of tyrosine kinase and phosphatase inhibitors on mitosis progression in synchronized tobacco BY-2 cells.

    Science.gov (United States)

    Sheremet, Ya A; Yemets, A I; Azmi, A; Vissenberg, K; Verbelen, J P; Blume, Ya B

    2012-01-01

    To test whether reversible tubulin phosphorylation plays any role in the process of plant mitosis the effects of inhibitors of tyrosine kinases, herbimycin A, genistein and tyrphostin AG 18, and of an inhibitor of tyrosine phosphatases, sodium orthovanadate, on microtubule organization and mitosis progression in a synchronized BY-2 culture has been investigated. It was found that treatment with inhibitors of tyrosine kinases of BY-2 cells at the G2/M transition did not lead to visible disturbances of mitotic microtubule structures, while it did reduce the frequency of their appearance. We assume that a decreased tyrosine phosphorylation level could alter the microtubule dynamic instability parameters during interphase/prophase transition. All types of tyrosine kinase inhibitors used caused a prophase delay: herbimycin A and genistein for 2 h, and tyrphostin AG18 for 1 h. Thereafter the peak of mitosis was displaced for 1 h by herbimycin A or genistein exposure, but after tyrphostin AG18 treatment the timing of the mitosis-peak was comparable to that in control cells. Enhancement of tyrosine phosphorylation induced by the tyrosine phosphatase inhibitor resulted in the opposite effect on BY-2 mitosis transition. Culture treatment with sodium orthovanadate during 1 h resulted in an accelerated start of the prophase and did not lead to the alteration in time of the mitotic index peak formation, as compared to control cells. We suppose that the reversible tyrosine phosphorylation can be involved in the regulation of interphase to M phase transition possibly through regulation of microtubule dynamics in plant cells.

  14. An Exocyst Complex Functions in Plant Cell Growth in Arabidopsis and Tobacco

    Czech Academy of Sciences Publication Activity Database

    Hála, Michal; Cole, R.A.; Synek, Lukáš; Drdová, Edita; Pečenková, Tamara; Nordheim, A.; Lamkemeyer, T.; Madlung, J.; Hochholdinger, F.; Fowler, J.E.; Žárský, Viktor

    2008-01-01

    Roč. 20, č. 5 (2008), s. 1330-1345 ISSN 1040-4651 R&D Projects: GA MŠk ME 841; GA MŠk(CZ) LC06034; GA AV ČR IAA6038410 Institutional research plan: CEZ:AV0Z50380511 Keywords : EXOCYST * PROTEIN COMPLEX * CELL POLARITY * MORPHOGENESIS * GTPASES Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.296, year: 2008

  15. Alamethicin permeabilizes the plasma membrane and mitochondria but not the tonoplast in tobacco (Nicotiana tabacum L. cv Bright Yellow) suspension cells

    DEFF Research Database (Denmark)

    Matic, S.; Geisler, D.A.; Møller, I.M.

    2005-01-01

    remained intact, as indicated by an unaffected tonoplast proton gradient. Low-flux permeabilization of plasma membranes and mitochondria at moderate AlaM concentrations was reversible and did not affect cell vigour. Higher AlaM concentrations induced cell death. After the addition of catalase that removes...... concentrations. Possible uses and limitations of this method for plant cell research are discussed.......The ion channel-forming peptide AlaM (alamethicin) is known to permeabilize isolated mitochondria as well as animal cells. When intact tobacco (Nicotiana tabacum L.) Bright Yellow-2 cells were treated with AlaM, the cells became permeable for low-molecular-mass molecules as shown by induced leakage...

  16. Secondhand Tobacco Smoke (Environmental Tobacco Smoke)

    Science.gov (United States)

    Learn about secondhand tobacco smoke, which can raise your risk of lung cancer. Secondhand tobacco smoke is the combination of the smoke given off by a burning tobacco product and the smoke exhaled by a smoker. Also called environmental tobacco smoke, involuntary smoke, and passive smoke.

  17. Induction of anchorage-independent growth in primary human cells exposed to protons or HZE ions separately or in dual exposures.

    Science.gov (United States)

    Sutherland, B M; Cuomo, N C; Bennett, P V

    2005-10-01

    Travelers on space missions will be exposed to a complex radiation environment that includes protons and heavy charged particles. Since protons are present at much higher levels than are heavy ions, the most likely scenario for cellular radiation exposure will be proton exposure followed by a hit by a heavy ion. Although the effects of individual ion species on human cells are being investigated extensively, little is known about the effects of exposure to both radiation types. One useful measure of mammalian cell damage is induction of the ability to grow in a semi-solid agar medium highly inhibitory to the growth of normal human cells, termed neoplastic transformation. Using primary human cells, we evaluated induction of soft-agar growth and survival of cells exposed to protons only or to heavy charged particles (600 MeV/nucleon silicon) only as well as of cells exposed to protons followed after a 4-day interval by silicon ions. Both ions alone efficiently transformed the human cells to anchorage-independent growth. Initial experiments indicate that the dose responses for neoplastic transformation of cells exposed to protons and then after 4 days to silicon ions appear similar to that of cells exposed to silicon ions alone.

  18. Sulindac enhances the killing of cancer cells exposed to oxidative stress.

    Directory of Open Access Journals (Sweden)

    Maria Marchetti

    2009-06-01

    Full Text Available Sulindac is an FDA-approved non-steroidal anti-inflammatory drug (NSAID that affects prostaglandin production by inhibiting cyclooxygenases (COX 1 and 2. Sulindac has also been of interest for more than decade as a chemopreventive for adenomatous colorectal polyps and colon cancer.Pretreatment of human colon and lung cancer cells with sulindac enhances killing by an oxidizing agent such as tert-butyl hydroperoxide (TBHP or hydrogen peroxide. This effect does not involve cyclooxygenase (COX inhibition. However, under the conditions used, there is a significant increase in reactive oxygen species (ROS within the cancer cells and a loss of mitochondrial membrane potential, suggesting that cell death is due to apoptosis, which was confirmed by Tunel assay. In contrast, this enhanced killing was not observed with normal lung or colon cells.These results indicate that normal and cancer cells handle oxidative stress in different ways and sulindac can enhance this difference. The combination of sulindac and an oxidizing agent could have therapeutic value.

  19. Cell proliferation and 3H-proline incorporation in periodontal ligament exposed to mechanical stress

    International Nuclear Information System (INIS)

    Kunz, J.; Plascke, C.; Duncker, M.

    1988-01-01

    In order to study the metabolic processes induced in the periodontal ligament by mechanical influences, a tension spring was implanted in rats between the incisor and the first maxillary molar on the right-hand side, while the left maxilla of these animals as well as non-operated rats served as controls. Under such mechanical stress, there occurred at 3, 10 and 21 days after implantation a significant increase in the 3 H-thymidine labelling index, which was demonstrate histoautoradiographically. A change in cell density was not discovered. Therefore, the increase in S-phase fraction as equally recorded in both pressure and tension zones is regarded as an expression of an enhanced cell turnover. Cell renewal in the periodontal ligament can be modified by inflammatory processes within the gingival region. There is a slight enlargement of the periodontal space in the tension zone. Under experimental conditions, no change occurs in the silver grain number per cell after 3 H-proline administration. The results indicate that, following the impact of orthodontic forces, the reactivity of periodontal cell proliferation as compared to collagen synthesis is enhanced. (author)

  20. Regulation of SUMO2 Target Proteins by the Proteasome in Human Cells Exposed to Replication Stress

    DEFF Research Database (Denmark)

    Bursomanno, Sara; McGouran, Joanna F; Kessler, Benedikt M

    2015-01-01

    In human cells, SUMO2 is predominantly conjugated to target proteins in response to cellular stress. Previous studies suggested that proteins conjugated to SUMO2, but not to SUMO1, could be regulated by the ubiquitin-mediated proteasome system. Hence, we set out to understand the role...... of the proteasome in determining the fate of proteins conjugated to SUMO2 when cells are treated with DNA replication stress conditions. We conducted a quantitative proteomic analysis in a U2OS cell line stably expressing SUMO2(Q87R) tagged with StrepHA in the presence or absence of epoxomicin (EPOX), a proteasome...... inhibitor. We identified subgroups of putative SUMO2 targets that were either degraded or stabilized by EPOX upon SUMO2 conjugation in response to replication stress. Interestingly, the subgroup of proteins degraded upon SUMO2 conjugation was enriched in proteins playing roles in DNA damage repair...

  1. Ultrastructural alterations in ciliary cells exposed to ionizing radiation. A scanning and transmission electron microscopic study

    Energy Technology Data Exchange (ETDEWEB)

    Baldetorp, L; Mecklenburg, C v; Haakansson, C H [Lund Univ. (Sweden). Hospital; Lund Univ. (Sweden). Dept. of Zoology)

    1977-01-01

    Early effects of ionizing radiation were investigated in an experimental in vitro system using the ciliary cells of the tracheal mucous membrane of the rabbit, irradiated at 30/sup 0/C and at more than 90% humidity. The changes in physiological activities of the ciliary cells caused by irradation were continously registered during the irradation. The specimens were examined immediately after irradiation electron microscopically. The morphological changes in irradiated material after 10-70 Gy are compared with normal material. After 40-70 Gy, scanning electron microscopy revealed the formation of vesicles on cilia, and club-like protrusions and adhesion of their tips. After 30-70 Gy, a swelling of mitochondrial membranes and cristae was apparent transmission electron microscopically. The membrane alterations caused by irradiation are assumed to disturb the permeability and flow of ATP from the mitochondria, which in turn leads to the recorded changes in the activity of the ciliated cells.

  2. Measurement of oxidative damage to DNA in nanomaterial exposed cells and animals

    DEFF Research Database (Denmark)

    Møller, Peter; Jensen, Ditte Marie; Christophersen, Daniel Vest

    2015-01-01

    -reactivity with other molecules in cells. This review provides an overview of efforts to reliably detect oxidatively damaged DNA and a critical assessment of the published studies on DNA damage levels. Animal studies with high baseline levels of oxidatively damaged DNA are more likely to show positive associations...... of oxidatively damaged DNA in lung tissue. Oral exposure to nanosized carbon black, TiO2 , carbon nanotubes and ZnO is associated with elevated levels of oxidatively damaged DNA in tissues. These observations are supported by cell culture studies showing concentration-dependent associations between ENM exposure...... and oxidatively damaged DNA measured by the comet assay. Cell culture studies show relatively high variation in the ability of ENMs to oxidatively damage DNA; hence, it is currently impossible to group ENMs according to their DNA damaging potential. Environ. Mol. Mutagen., 2014. © 2014 Wiley Periodicals, Inc....

  3. Inactivation of Cronobacter malonaticus cells and inhibition of its biofilm formation exposed to hydrogen peroxide stress.

    Science.gov (United States)

    Ye, Yingwang; Zhang, Maofeng; Jiao, Rui; Ling, Na; Zhang, Xiyan; Tong, Liaowang; Zeng, Haiyang; Zhang, Jumei; Wu, Qingping

    2018-01-01

    Presence of Cronobacter malonaticus in powdered infant formula (PIF) poses a high risk to infant and public health. Cronobacter malonaticus has been widely distributed in food and food processing environments, and the true origin of C. malonaticus in PIF is poorly understood. Control and prevention of C. malonaticus is necessary for achieving microbial safety of PIF. However, little information about decontamination of C. malonaticus is available. In this study, effects of hydrogen peroxide on inactivation and morphological changes of C. malonaticus cells were determined. Furthermore, inhibitory effects of H 2 O 2 on biofilm formation in C. malonaticus were also performed. Results indicated that H 2 O 2 could completely inactivate C. malonaticus in sterile water with 0.06% H 2 O 2 for 25 min, 0.08% H 2 O 2 for 15 min, and 0.10% for 10 min, respectively, whereas the survival rates of C. malonaticus in tryptic soy broth medium significantly increased with the same treatment time and concentration of H 2 O 2 . In addition, morphological changes of C. malonaticus cells, including cell shrinkage, disruption of cells, cell intercession, and leakage of intercellular material in sterile water after H 2 O 2 treatment, were more predominant than those in tryptic soy broth. Finally, significant reduction in biofilm formation by H 2 O 2 was found using crystal violet staining, scanning electron microscopy, and confocal laser scanning microscopy detection compared with control samples. This is the first report to determine the effects of H 2 O 2 on C. malonaticus cells and biofilm formation. The findings provided valuable information for practical application of H 2 O 2 for decontamination of C. malonaticus in dairy processing. Copyright © 2018 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  4. Transient permeabilization of cell membranes by ultrasound-exposed microbubbles is related to formation of hydrogen peroxide.

    Science.gov (United States)

    Juffermans, L J M; Dijkmans, P A; Musters, R J P; Visser, C A; Kamp, O

    2006-10-01

    In the present study, we addressed the interactions among ultrasound, microbubbles, and living cells as well as consequent arising bioeffects. We specifically investigated whether hydrogen peroxide (H(2)O(2)) is involved in transient permeabilization of cell membranes in vitro after ultrasound exposure at low diagnostic power, in the presence of stable oscillating microbubbles, by measuring the generation of H(2)O(2) and Ca(2+) influx. Ultrasound, in the absence or presence of SonoVue microbubbles, was applied to H9c2 cells at 1.8 MHz with a mechanical index (MI) of 0.1 or 0.5 during 10 s. This was repeated every minute, for a total of five times. The production of H(2)O(2) was measured intracellularly with CM-H(2)DCFDA. Cell membrane permeability was assessed by measuring real-time changes in intracellular Ca(2+) concentration with fluo-4 using live-cell fluorescence microscopy. Ultrasound, in the presence of microbubbles, caused a significant increase in intracellular H(2)O(2) at MI 0.1 of 50% and MI 0.5 of 110% compared with control (P ultrasound exposure was completely blocked at MI 0.1 (P ultrasound-exposed microbubbles.

  5. Patient with pure red cell aplasia who was exposed to the atomic bomb near the site of blast

    Energy Technology Data Exchange (ETDEWEB)

    Toyoda, N; Matsukuma, G; Uchiyama, E; Tsunoo, S; Anpi, S [Nagasaki Atomic Bomb Hospital (Japan)

    1976-03-01

    This paper describes a female patient who had been exposed at the age of 26 to the atomic bomb at a distance of 1.3 km from the bombed area. She was pointed out to have anemia and the disturbance of the liver in 1961. In 1963, she had iron deficiency, hypoleucocytosis, and hyper-..gamma.. globulinemia, and showed a high value of serum colloids. In 1969, she had more frequently anemia together with hyper ..gamma..-globulinemia. The erythrocyte sedimentation rate and serum colloids were still high. The positive reaction to the LE cells began to appear. The liver biopsy findings showed proliferation of Kupper cells in the lobule, decidual degeneration of some liver cells, infiltration of many round cells in the Glisson's sheath. Therefore, she was diagnosed as having pure red cell aplasia and lupoid hepatitis. However, in 1970 she was suspected of having pulmonary valvular insufficiency caused by pulmonary hypertension in spite of the continuous steroid therapy. In 1974, she died of jundice, hyper-..gamma..-globulinemia which accompanied urea in the blood and the increased values of nitrogen and NH/sub 3/. This patient was considered to be interested on the assumption of the relationships between exposure to the atomic bomb and disturbance of immunity.

  6. Bone-Marrow Stem-Cell Survival in the Non-Uniformly Exposed Mammal

    Energy Technology Data Exchange (ETDEWEB)

    Bond, V. P.; Robinson, C. V. [Brookhaven National Laboratory, Medical Research Center, Upton, Long Island, NY (United States)

    1967-07-15

    For comparison of the effectiveness of non-uniform versus uniform irradiations in causing haematological death in mammals, a model of the irradiated haemopoietic system has been proposed. The essential features of this model are: (1) that different parts of the haemopoietic system have numbers of stem cells which are proportioned to the amounts of active marrow in those parts as measured by {sup 59}Fe uptake, (2) that stem cells in the different parts are subject to the, same dose-survival relationship, and (3) that survival of the animal depends on survival of a critical fraction of the total number of stem cells independent of their distribution among the parts of the total marrow mass. To apply this model one needs to know: (a) the relative {sup 59}Fe uptakes of the different parts of the haemopoietic system, (b) the doses delivered to those parts by each of the exposures to be compared, and (c) the dose-survival curve applicable to the stem cells. From these one can calculate the fraction of stem cells surviving each exposure. In a preliminary communication the applicability of the model was investigated using data obtained entirely from the literature. Additional data, particularly on bone-marrow distribution, have since been obtained and are included here. The primary object of the present paper is to test further the validity of the above 'stem-cell survival model'. Data on bilateral (essentially uniform) versus unilateral and non-uniform rotational exposures in mammals are examined with respect to the surviving fraction of stem cells at the LD{sub 50/30} day dose level. Although an adequate test is not possible at present for lack of a full set of data in any one species, a partial test indicates compatibility with data for dogs and rats. Other possible mortality determinants such as doses or exposures at entrance, midline or exit, or the gram-rads or average dose to the marrow, appear to be less useful than the critical stem-cell survival fraction.

  7. Smad2/3 Upregulates the Expression of Vimentin and Affects Its Distribution in DBP-Exposed Sertoli Cells

    Directory of Open Access Journals (Sweden)

    Xi Zhang

    2015-01-01

    Full Text Available Sertoli cells (SCs in the testes provide physical and nutritional support to germ cells. The vimentin cytoskeleton in SCs is disrupted by dibutyl phthalate (DBP, which leads to SCs dysfunction. In a previous study, we found that peroxisome proliferator-activated receptor alpha (PPARα influenced the distribution of vimentin by affecting its phosphorylation in DBP-exposed SCs. In the present study, we investigated the role of Smad2/3 in regulating the expression of vimentin in DBP-exposed SCs. We hypothesized that Smad2/3 affects the distribution of vimentin by regulating its expression and that there is cross talk between Smad2/3 and PPARα. The real-time PCR and ChIP-qPCR results showed that SB431542 (an inhibitor of Smad2/3 could significantly attenuate the expression of vimentin induced by DBP in SCs. Phosphorylated and soluble vimentin were both downregulated by SB431542 pretreatment. WY14643 (an agonist of PPARα pretreatment stimulated, while GW6471 (an antagonist of PPARα inhibited, the activity of Smad2/3; SB431542 pretreatment also inhibited the activity of PPARα, but it did not rescue the DBP-induced collapse in vimentin. Our results suggest that, in addition to promoting the phosphorylation of vimentin, DBP also stimulates the expression of vimentin by activating Smad2/3 in SCs and thereby induces irregular vimentin distribution.

  8. Heterogeneity in c-jun gene expression in normal and malignant cells exposed to either ionizing radiation or hydrogen peroxide

    International Nuclear Information System (INIS)

    Horio, M.; Collart, F.R.; Huberman, E.

    1993-01-01

    We investigated the role of reactive oxygen intermediates and protein kinase C (PKC) in induction of c-jun gene expression in human ML-2 leukemic cells and normal DET-551 fibroblasts by comparing the effects of either ionizing radiation or H 2 O 2 exposure in the presence or absence of appropriate inhibitors. In these cell types, the radiation and H 2 O 2 -mediated increase in c-jun mRNA levels could be prevented by pretreatment of the cells with N-acetylcysteine, an antioxidant, or H7, an inhibitor of PKC and cAMP-dependent protein kinase (PKA), but not by HA1004, an inhibitor of PKA. These results suggest a role for PKC and reactive oxygen intermediates in the induction of c-jun gene expression in both normal and tumor cells. We also investigated potential differences in radiation- or H 2 O 2 -induced c-jun gene expression in normal and tumor cells by examining steady-state c-jun mRNA levels in a number of human fibroblast, leukemia, melanoma, sarcoma, and carcinoma cell types. We observed heterogeneity in the steady-state level of c-jun mRNA in both the untreated normal and tumor cells and in such cells exposed to ionizing radiation or to H 2 O 2 . Exposure to radiation or to hydrogen peroxide produced a varied response which ranged from little or no induction to a more than two orders of magnitude increase in the steady-state level of the c-jun mRNA

  9. DJ1 Expression Downregulates in Neuroblastoma Cells (SK-N-MC Chronically Exposed to HIV-1 and Cocaine.

    Directory of Open Access Journals (Sweden)

    Upal eRoy

    2015-07-01

    Full Text Available Background: HIV-associated neurological disorder (HAND has long been recognized as a consequence of Human Immunodeficiency Virus (HIV infection in the brain. The pathology of HAND gets more complicated with the recreational drug use such as cocaine. Recent studies have suggested multiple genetic influences involved in the pathology of addiction and HAND but only a fraction of the entire genetic risk has been investigated so far. In this regard, role of DJ1 protein (a gene linked to autosomal recessive early-onset Parkinson’s disease in regulating dopamine transmission and reactive oxygen species (ROS production in neuronal cells will be worth investigating in HIV-1 and cocaine exposed microenvironment. Being a very abundant protein in the brain, DJ1 could serve as a potential marker for early detection of HIV-1 and/or cocaine related neurological disorder.Methods: In vitro analysis was done to observe the effect of HIV-1 and/or cocaine on DJ1 protein expression in neuroblastoma cells (SK-N-MC. Gene expression and protein analysis of DJ1 was done on the HIV infected and/or cocaine treated SK-N-MC and compared to untreated cells using real time PCR, Western Blot and flow cytometry.Results: Gene expression and protein analysis indicated that there was a significant decrease in DJ1 expression in SK-N-MC chronically exposed to HIV-1 and/or cocaine.Conclusion: This is the first study to establish that DJ1 expression level in the neuronal cells significantly decreased in presence of HIV-1and/or cocaine indicating oxidative stress level of dopamine neurons.

  10. Global gene expression changes in human urothelial cells exposed to low-level monomethylarsonous acid

    Czech Academy of Sciences Publication Activity Database

    Medeiros, M.; Zheng, X.; Novák, Petr; Wnek, S.M.; Chyan, V.; Escudero-Lourdes, C.; Gandolfi, A.J.

    2012-01-01

    Roč. 291, 1-3 (2012), s. 102-112 ISSN 0300-483X Institutional research plan: CEZ:AV0Z50510513 Institutional support: RVO:60077344 Keywords : HUMAN BLADDER CELLS * METHYLATED TRIVALENT ARSENICALS * MALIGNANT-TRANSFORMATION0300 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.017, year: 2012

  11. Nanomimics of host cell membranes block invasion and expose invasive malaria parasites.

    Science.gov (United States)

    Najer, Adrian; Wu, Dalin; Bieri, Andrej; Brand, Françoise; Palivan, Cornelia G; Beck, Hans-Peter; Meier, Wolfgang

    2014-12-23

    The fight against most infectious diseases, including malaria, is often hampered by the emergence of drug resistance and lack or limited efficacies of vaccines. Therefore, new drugs, vaccines, or other strategies to control these diseases are needed. Here, we present an innovative nanotechnological strategy in which the nanostructure itself represents the active substance with no necessity to release compounds to attain therapeutic effect and which might act in a drug- and vaccine-like dual function. Invasion of Plasmodium falciparum parasites into red blood cells was selected as a biological model for the initial validation of this approach. Stable nanomimics-polymersomes presenting receptors required for parasite attachment to host cells-were designed to efficiently interrupt the life cycle of the parasite by inhibiting invasion. A simple way to build nanomimics without postformation modifications was established. First, a block copolymer of the receptor with a hydrophobic polymer was synthesized and then mixed with a polymersome-forming block copolymer. The resulting nanomimics bound parasite-derived ligands involved in the initial attachment to host cells and they efficiently blocked reinvasion of malaria parasites after their egress from host cells in vitro. They exhibited efficacies of more than 2 orders of magnitude higher than the soluble form of the receptor, which can be explained by multivalent interactions of several receptors on one nanomimic with multiple ligands on the infective parasite. In the future, our strategy might offer interesting treatment options for severe malaria or a way to modulate the immune response.

  12. Analysis of miRNA expression profiles in melatonin-exposed GC-1 spg cell line.

    Science.gov (United States)

    Zhu, Xiaoling; Chen, Shuxiong; Jiang, Yanwen; Xu, Ying; Zhao, Yun; Chen, Lu; Li, Chunjin; Zhou, Xu

    2018-02-05

    Melatonin is an endocrine neurohormone secreted by pinealocytes in the pineal gland. It exerts diverse physiological effects, such as circadian rhythm regulator and antioxidant. However, the functional importance of melatonin in spermatogenesis regulation remains unclear. The objectives of this study are to: (1) detect melatonin affection on miRNA expression profiles in GC-1 spg cells by miRNA deep sequencing (DeepSeq) and (2) define melatonin affected miRNA-mRNA interactions and associated biological processes using bioinformatics analysis. GC-1 spg cells were cultured with melatonin (10 -7 M) for 24h. DeepSeq data were validated using quantitative real-time reverse transcription polymerase chain reaction analysis (qRT-PCR). A total of 176 miRNA expressions were found to be significantly different between two groups (fold change of >2 or melatonin could regulate the expression of miRNA to perform its physiological effects in GC-1 spg cells. These results should be useful to investigate the biological function of miRNAs regulated by melatonin in spermatogenesis and testicular germ cell tumor. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Disruption of glucagon receptor signaling causes hyperaminoacidemia exposing a possible liver - alpha-cell axis

    DEFF Research Database (Denmark)

    Galsgaard, Katrine D; Winther-Sørensen, Marie; Ørskov, Cathrine

    2018-01-01

    Glucagon secreted from the pancreatic alpha-cells is essential for regulation of blood glucose levels. However, glucagon may play an equally important role in the regulation of amino acid metabolism by promoting ureagenesis. We hypothesized that disruption of glucagon receptor signaling would lead...

  14. Photobiomodulation Protects and Promotes Differentiation of C2C12 Myoblast Cells Exposed to Snake Venom.

    Directory of Open Access Journals (Sweden)

    Luciana Miato Gonçalves Silva

    Full Text Available Snakebites is a neglected disease and in Brazil is considered a serious health problem, with the majority of the snakebites caused by the genus Bothrops. Antivenom therapy and other first-aid treatments do not reverse local myonecrose which is the main sequel caused by the envenomation. Several studies have shown the effectiveness of low level laser (LLL therapy in reducing local myonecrosis induced by Bothropic venoms, however the mechanism involved in this effect is unknown. In this in vitro study, we aimed to analyze the effect of LLL irradiation against cytotoxicity induced by Bothrops jararacussu venom on myoblast C2C12 cells.C2C12 were utilized as a model target and were incubated with B. jararacussu venom (12.5 μg/mL and immediately irradiated with LLL at wavelength of red 685 nm or infrared 830 nm with energy density of 2.0, 4.6 and 7.0 J/cm2. Effects of LLL on cellular responses of venom-induced cytotoxicity were examined, including cell viability, measurement of cell damage and intra and extracellular ATP levels, expression of myogenic regulatory factors, as well as cellular differentiation.In non-irradiated cells, the venom caused a decrease in cell viability and a massive release of LDH and CK levels indicating myonecrosis. Infrared and red laser at all energy densities were able to considerably decrease venom-induced cytotoxicity. Laser irradiation induced myoblasts to differentiate into myotubes and this effect was accompanied by up regulation of MyoD and specially myogenin. Moreover, LLL was able to reduce the extracellular while increased the intracellular ATP content after venom exposure. In addition, no difference in the intensity of cytotoxicity was shown by non-irradiated and irradiated venom.LLL irradiation caused a protective effect on C2C12 cells against the cytotoxicity caused by B. jararacussu venom and promotes differentiation of these cells by up regulation of myogenic factors. A modulatory effect of ATP synthesis may

  15. Comparative Analysis of WUSCHEL-Related Homeobox Genes Revealed Their Parent-of-Origin and Cell Type-Specific Expression Pattern During Early Embryogenesis in Tobacco

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    Xuemei Zhou

    2018-03-01

    Full Text Available WUSCHEL-related homeobox (WOX gene is a plant-specific clade of homeobox transcription factors. Increasing evidences reveal that WOXs play critical roles in early embryogenesis, which involves zygote development, initiation of zygote division, and apical or basal cell lineage establishment. However, how WOXs regulate these developmental events remains largely unknown, and even detailed expression pattern in gametes and early proembryos is not yet available. Here, 13 WOX family genes were identified in Nicotiana tabacum genome. Comparative analysis of 13 WOX family genes with their homologs in Arabidopsis thaliana reveals relatively conserved expression pattern of WUS and WOX5 in shoot/root apical meristem. Whereas variations were also found, e.g., lacking homolog of WOX8 (a marker for suspensor cell in tobacco genome and the expression of WOX2/WOX9 in both apical cell and basal cell. Transient transcriptional activity analysis revealed that WOXs in WUS clade have repressive activities for their target's transcription, whereas WOXs in ancient and intermediate clade have activation activities, giving a molecular basis for the phylogenetic classification of tobacco WOXs into three major clades. Expression pattern analysis revealed that some WOXs (e.g., WOX 13a expressed in both male and female gametes and some WOXs (e.g., WOX 11 and WOX 13b displayed the characteristics of parent-of-origin genes. Interestingly, some WOXs (e.g., WOX2 and WOX9, which are essential for early embryo patterning, were de novo transcribed in zygote, indicating relevant mechanism for embryo pattern formation is only established in zygote right after fertilization and not carried in by gametes. We also found that most WOXs displayed a stage-specific and cell type-specific expression pattern. Taken together, this work provides a detailed landscape of WOXs in tobacco during fertilization and early embryogenesis, which will facilitate the understanding of their specific roles

  16. The morphological and molecular changes of brain cells exposed to direct current electric field stimulation.

    Science.gov (United States)

    Pelletier, Simon J; Lagacé, Marie; St-Amour, Isabelle; Arsenault, Dany; Cisbani, Giulia; Chabrat, Audrey; Fecteau, Shirley; Lévesque, Martin; Cicchetti, Francesca

    2014-12-07

    The application of low-intensity direct current electric fields has been experimentally used in the clinic to treat a number of brain disorders, predominantly using transcranial direct current stimulation approaches. However, the cellular and molecular changes induced by such treatment remain largely unknown. Here, we tested various intensities of direct current electric fields (0, 25, 50, and 100V/m) in a well-controlled in vitro environment in order to investigate the responses of neurons, microglia, and astrocytes to this type of stimulation. This included morphological assessments of the cells, viability, as well as shape and fiber outgrowth relative to the orientation of the direct current electric field. We also undertook enzyme-linked immunosorbent assays and western immunoblotting to identify which molecular pathways were affected by direct current electric fields. In response to direct current electric field, neurons developed an elongated cell body shape with neurite outgrowth that was associated with a significant increase in growth associated protein-43. Fetal midbrain dopaminergic explants grown in a collagen gel matrix also showed a reorientation of their neurites towards the cathode. BV2 microglial cells adopted distinct morphological changes with an increase in cyclooxygenase-2 expression, but these were dependent on whether they had already been activated with lipopolysaccharide. Finally, astrocytes displayed elongated cell bodies with cellular filopodia that were oriented perpendicularly to the direct current electric field. We show that cells of the central nervous system can respond to direct current electric fields both in terms of their morphological shape and molecular expression of certain proteins, and this in turn can help us to begin understand the mechanisms underlying the clinical benefits of direct current electric field. © The Author 2015. Published by Oxford University Press on behalf of CINP.

  17. Induction of Cell Death through Alteration of Oxidants and Antioxidants in Epithelial Cells Exposed to High Energy Protons

    Science.gov (United States)

    Ramesh, Govindarajan; Wu, Honglu

    2012-01-01

    Radiation affects several cellular and molecular processes including double strand breakage, modifications of sugar moieties and bases. In outer space, protons are the primary radiation source which poses a range of potential health risks to astronauts. On the other hand, the use of proton radiation for tumor radiation therapy is increasing as it largely spares healthy tissues while killing tumor tissues. Although radiation related research has been conducted extensively, the molecular toxicology and cellular mechanisms affected by proton radiation remain poorly understood. Therefore, in the present study, we irradiated rat epithelial cells (LE) with different doses of protons and investigated their effects on cell proliferation and cell death. Our data showed an inhibition of cell proliferation in proton irradiated cells with a significant dose dependent activation and repression of reactive oxygen species (ROS) and antioxidants, glutathione and superoxide dismutase respectively as compared to control cells. In addition, apoptotic related genes such as caspase-3 and -8 activities were induced in a dose dependent manner with corresponding increased levels of DNA fragmentation in proton irradiated cells than control cells. Together, our results show that proton radiation alters oxidant and antioxidant levels in the cells to activate apoptotic pathway for cell death.

  18. Radiosensitizing effect of Chitosan on HeLa and LN 18 brain tumor cells exposed to electron beam radiation

    International Nuclear Information System (INIS)

    Rao, Shama; Shetty, Sukanya; Suchetha Kumari, N.; Madhu, L.N.

    2014-01-01

    Chitosan has been widely used for multiple applications because it is a non-toxic biocompatible, biodegradable, and adsorptive material. A previous study has shown that low-molecular-weight chitosan (LMWC) exerts a cytotoxic effect on oral cancer cells. Although a higher concentration of LMWC in comparison to cisplatin was needed in order to kill cancer cells, it was relatively less cytotoxic to non-cancer cells. Some of the well known anticancer drugs have the property of sensitizing the cell to radiation, which will be more applicable during combination therapy of cancer. The present study was undertaken to find the radiosensitizing effect of chitosan on Hela and Brain tumor (LN18) cells against electron beam radiation (EBR). Both the cancer cell lines, Hela and LN 18 were treated with different concentration of chitosan (50 and 100 μg/ml) pre and post exposure to 4 Gy EBR. The percentage of cell viability, percentage of apoptosis and ssDNA damage in the treated cells were assessed by MTT assay, DNA diffusion assay and comet assay respectively. The obtained results showed 62.13 1 5.08 and 65.24 1 2.45 percent Hela and LN 18 viable cells at 24 hour after the exposure to 4 Gy EBR. The percentage of viability was found to be decreased in cells exposed to EBR in the presence of chitosan. Supporting to this, percentage of apoptotic cells was found to be more in treated groups (28.13 1 4.34 and 25.13 1 3.76) when compared with control (23.19 1 1.07 and 20.79 1 4.86). Treatment of HeLa and LN18 before and after the exposure of EBR showed significantly (P<0.05) more frequency of micronucleus and % of DNA damage than the 4 Gy EBR control group. These results conclude the sensitizing effect of chitosan on cancer cell line against EBR exposure. (author)

  19. Somatic cell chromosome changes in a population exposed to low levels of ionizing radiation

    International Nuclear Information System (INIS)

    Brandom, W.F.

    1982-01-01

    The analysis of chromosomes from the cells of 897 plutonium workers is reported. Within three years, the number of controls alone analyzed for this study approximated the largest plutonium cytogenetic studies today including workers plus controls (81 compared to 84 in a 1979 French study and 94 in a 1982 British report). The number of subjects analyzed in the first three years were: new employees - 245; new employees assigned to plutonium work areas - 7; workers with less than 3% of maximum permissible systemic burden (MPSB) - 35; workers with less than 50% MPSB - 274; workers with greater than 50% of MPSB - 65; follow-up familial congenital cytogenetics at worker request (through Medical) - 6; polymorphic/variant chromosome constitutions - 242; re-sampling of workers with elevated aberration yields - 26; cell sample study - 28; sister-chromatid-exchange (SCE) study - 23; beryllium workers at Rocky Flats - 10; Hanford worker analyses - 5). 20 refs., 3 figs., 5 tabs

  20. DNA repair in human cells exposed to combinations of carcinogenic agents

    International Nuclear Information System (INIS)

    Setlow, R.B.; Ahmed, F.E.

    1980-01-01

    Normal human and XP 2 fibroblasts were treated with uv plus uv-mimetic chemicals. The uv dose used was sufficient to saturate the uv excision repair system. Excision repair after combined treatments was estimated by unscheduled DNA synthesis, BrdUrd photolysis, and the loss of sites sensitive to a uv specific endonuclease. Since the repair of damage from uv and its mimetics is coordinately controlled we expected that there would be similar rate-limiting steps in the repair of uv and chemical damage and that after a combined treatment the total amount of repair would be the same as from uv or the chemicals separately. The expectation was not fulfilled. In normal cells repair after a combined treatment was additive whereas in XP cells repair after a combined treatment was usually less than after either agent separately. The chemicals tested were AAAF, DMBA-epoxide, 4NQO, and ICR-170

  1. Disruption of cardiogenesis in human embryonic stem cells exposed to trichloroethylene.

    Science.gov (United States)

    Jiang, Yan; Wang, Dan; Zhang, Guoxing; Wang, Guoqing; Tong, Jian; Chen, Tao

    2016-11-01

    Trichloroethylene (TCE) is ubiquitous in our living environment, and prenatal exposure to TCE is reported to cause congenital heart disease in humans. Although multiple studies have been performed using animal models, they have limited value in predicting effects on humans due to the unknown species-specific toxicological effects. To test whether exposure to low doses of TCE induces developmental toxicity in humans, we investigated the effect of TCE on human embryonic stem cells (hESCs) and cardiomyocytes (derived from the hESCs). In the current study, hESCs cardiac differentiation was achieved by using differentiation medium consisting of StemPro-34. We examined the effects of TCE on cell viability by cell growth assay and cardiac inhibition by analysis of spontaneously beating cluster. The expression levels of genes associated with cardiac differentiation and Ca 2+ channel pathways were measured by immunofluorescence and qPCR. The overall data indicated the following: (1) significant cardiac inhibition, which was characterized by decreased beating clusters and beating rates, following treatment with low doses of TCE; (2) significant up-regulation of the Nkx2.5/Hand1 gene in cardiac progenitors and down regulation of the Mhc-7/cTnT gene in cardiac cells; and (3) significant interference with Ca 2+ channel pathways in cardiomyocytes, which contributes to the adverse effect of TCE on cardiac differentiation during early embryo development. Our results confirmed the involvement of Ca 2+ turnover network in TCE cardiotoxicity as reported in animal models, while the inhibition effect of TCE on the transition of cardiac progenitors to cardiomyocytes is unique to hESCs, indicating a species-specific effect of TCE on heart development. This study provides new insight into TCE biology in humans, which may help explain the development of congenital heart defects after TCE exposure. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1372-1380, 2016. © 2015 Wiley

  2. CYTOKINESIS-BLOCK MICRONUCLEUS ASSAY IN HUMAN GLIOMA CELLS EXPOSED TO RADIATION

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    Jerzy Slowinski

    2011-05-01

    Full Text Available Biological tests are efficient in reflecting the biological influences of several types of generally harmful exposures. The micronucleus assay is widely used in genotoxicity studies or studies on genomic damage in general. We present methodological aspects of cytokinesis-block micronucleus assay performed in human gliomas irradiated in vitro. Eight human glioblastoma cell lines obtained from DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Germany were gamma-irradiated (60Co over a dose range of 0-10 Gy. Cytokinesis-block micronucleus assay was performed to quantitate cytogenetic damage. The cells were fixed directly on dishes, stained with fluorochrome DAPI and evaluated under fluorescent and phase contrast microscope. The micronucleus frequency was expressed as a micronuclei (MN per binucleated cell (BNC ratio, calculated after scoring at least 100 BNC per dish. The frequency of spontaneous MN ranged from 0.17 to 0.613 (mean: 0.29 ± 0.14. After irradiation increase of MN frequency in the range of 0.312 - 2.241 (mean: 0.98 ± 0.68 was found at 10 Gy. Gliomas are extremely heterogenous in regard to cytogenetic effects of irradiation, as shown in this study by cytokinesis-block micronucleus assay. This test is easily performed on irradiated glioma cell lines and can assist in determining their radiosensitivity. However, in order to obtain reliable and reproducible results, precise criteria for MN scoring must be strictly followed. Simultaneous use of fluorescent and phase contrast equipment improves imaging of morphological details and can further optimize MN scoring.

  3. Ultrastructural study of mitochondrial damage in CHO cells exposed to hyperthermia.

    Science.gov (United States)

    Cole, A; Armour, E P

    1988-09-01

    A unique direct-view stereo electron microscope technique was used to visualize the structure and three-dimensional distributions of mitochondria in CHO cells in situ following hyperthermic treatments. Aberrations induced by various heating regimens were recorded. The protocol included a trypsin digestion that may have enhanced the expression of the initial heat damage. The developed damage was observed as increasing levels of mitochondrial distortion, swelling, and dissociation. Minimal damage was induced at 42 degrees C for exposures of up to 4 h, while significant damage was induced at 43 degrees C for exposures of more than 30 min and at 45 degrees C for exposures of more than 10 min. For moderate exposures, a partial recovery of mitochondrial integrity was observed when the heat treatment was followed by incubation at 37 degrees C for 24 h. Mitochondrial damage was related to the heat dose in that increasing treatment temperature resulted in greater damage, but when compared to cell survival the damage did not parallel cell killing under all time-temperature conditions.

  4. Alpha-fetoprotein is a biomarker of unfolded protein response and altered proteostasis in hepatocellular carcinoma cells exposed to sorafenib.

    Science.gov (United States)

    Houessinon, Aline; Gicquel, Albane; Bochereau, Flora; Louandre, Christophe; Nyga, Rémy; Godin, Corinne; Degonville, James; Fournier, Emma; Saidak, Zuzana; Drullion, Claire; Barbare, Jean-Claude; Chauffert, Bruno; François, Catherine; Pluquet, Olivier; Galmiche, Antoine

    2016-01-28

    Sorafenib is the treatment of reference for advanced hepatocellular carcinoma (HCC). A decrease in the serum levels of Alpha-fetoprotein (AFP) is reported to be the biological parameter that is best associated with disease control by sorafenib. In order to provide a biological rationale for the variations of AFP, we analyzed the various steps of AFP production in human HCC cell lines exposed to sorafenib. Sorafenib dramatically reduced the levels of AFP produced by HCC cells independently of its effect on cell viability. The mRNA levels of AFP decreased upon sorafenib treatment, while the AFP protein remained localized in the Golgi apparatus. Sorafenib activated the Regulated Inositol-Requiring Enzyme-1α (IRE-1α) and the PKR-like ER Kinase (PERK)-dependent arms of the Unfolded Protein Response (UPR). The inhibition of IRE-1α partially restored the mRNA levels of AFP upon treatment with sorafenib. The inhibition of both pathways partially prevented the drop in the production of AFP induced by sorafenib. The findings provide new insights on the regulation of AFP, and identify it as a biomarker suitable for the exploration of HCC cell proteostasis in the context of therapeutic targeting. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  5. Raman micro-spectroscopy analysis of human lens epithelial cells exposed to a low-dose-range of ionizing radiation

    Science.gov (United States)

    Allen, Christian Harry; Kumar, Achint; Qutob, Sami; Nyiri, Balazs; Chauhan, Vinita; Murugkar, Sangeeta

    2018-01-01

    Recent findings in populations exposed to ionizing radiation (IR) indicate dose-related lens opacification occurs at much lower doses (micro-spectroscopy was used to investigate the effects of varying doses of radiation, ranging from 0.01 Gy to 5 Gy, on human lens epithelial (HLE) cells which were chemically fixed 24 h post-irradiation. Raman spectra were acquired from the nucleus and cytoplasm of the HLE cells. Spectra were collected from points in a 3  ×  3 grid pattern and then averaged. The raw spectra were preprocessed and principal component analysis followed by linear discriminant analysis was used to discriminate between dose and control for 0.25, 0.5, 2, and 5 Gy. Using leave-one-out cross-validation accuracies of greater than 74% were attained for each dose/control combination. The ultra-low doses 0.01 and 0.05 Gy were included in an analysis of band intensities for Raman bands found to be significant in the linear discrimination, and an induced repair model survival curve was fit to a band-difference-ratio plot of this data, suggesting HLE cells undergo a nonlinear response to low-doses of IR. A survival curve was also fit to clonogenic assay data done on the irradiated HLE cells, showing a similar nonlinear response.

  6. PACAP and VIP inhibit the invasiveness of glioblastoma cells exposed to hypoxia through the regulation of HIFs and EGFR expression

    Directory of Open Access Journals (Sweden)

    Grazia eMaugeri

    2016-05-01

    Full Text Available Pituitary adenylate cyclase-activating polypeptide (PACAP and vasoactive intestinal peptide (VIP through the binding of vasoactive intestinal peptide receptors (VIPRs, perform a wide variety of effects in human cancers, including glioblastoma multiforme (GBM. This tumor is characterized by extensive areas of hypoxia, which triggers the expression of hypoxia-inducible factors (HIFs. HIFs not only mediate angiogenesis but also tumor cell migration and invasion. Furthermore, HIFs activation is linked to epidermal growth factor receptor (EGFR overexpression. Previous studies have shown that VIP interferes with the invasive nature of gliomas by regulating cell migration. However, the role of VIP family members in GBM infiltration under low oxygen tension has not been clarified yet. Therefore, in the present study we have investigated, for the first time, the molecular mechanisms involved in the anti-invasive effect of PACAP or VIP in U87MG glioblastoma cells exposed to hypoxia induced by treatment with desferrioxamine (DFX. The results suggest that either PACAP or VIP exert an anti-infiltrative effect under low oxygen tension by modulating HIFs and EGFR expression, key elements involved in cell migration and angiogenesis. These peptides act through the inhibition of PI3K/Akt and MAPK/ERK signaling pathways, which are known to have a crucial role in HIFs regulation. In conclusion, the modulation of hypoxic event and the anti-invasive effect exerted by some VIP family members might open new insights in the therapeutic approach to GBM.

  7. Lineage-related cytotoxicity and clonogenic profile of 1,4-benzoquinone-exposed hematopoietic stem and progenitor cells

    Energy Technology Data Exchange (ETDEWEB)

    Chow, Paik Wah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Abdul Hamid, Zariyantey, E-mail: zyantey@ukm.edu.my [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Chan, Kok Meng [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Inayat-Hussain, Salmaan Hussain [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Rajab, Nor Fadilah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia)

    2015-04-01

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) are sensitive targets for benzene-induced hematotoxicity and leukemogenesis. The impact of benzene exposure on the complex microenvironment of HSCs and HPCs remains elusive. This study aims to investigate the mechanism linking benzene exposure to targeting HSCs and HPCs using phenotypic and clonogenic analyses. Mouse bone marrow (BM) cells were exposed ex vivo to the benzene metabolite, 1,4-benzoquinone (1,4-BQ), for 24 h. Expression of cellular surface antigens for HSC (Sca-1), myeloid (Gr-1, CD11b), and lymphoid (CD45, CD3e) populations were confirmed by flow cytometry. The clonogenicity of cells was studied using the colony-forming unit (CFU) assay for multilineage (CFU-GM and CFU-GEMM) and single-lineage (CFU-E, BFU-E, CFU-G, and CFU-M) progenitors. 1,4-BQ demonstrated concentration-dependent cytotoxicity in mouse BM cells. The percentage of apoptotic cells increased (p < 0.05) following 1,4-BQ exposure. Exposure to 1,4-BQ showed no significant effect on CD3e{sup +} cells but reduced the total counts of Sca-1{sup +}, CD11b{sup +}, Gr-1{sup +}, and CD45{sup +} cells at 7 and 12 μM (p < 0.05). Furthermore, the CFU assay showed reduced (p < 0.05) clonogenicity in 1,4-BQ-treated cells. 1,4-BQ induced CFU-dependent cytotoxicity by significantly inhibiting colony growth for CFU-E, BFU-E, CFU-G, and CFU-M starting at a low concentration of exposure (5 μM); whereas for the CFU-GM and CFU-GEMM, the inhibition of colony growth was remarkable only at 7 and 12 μM of 1,4-BQ, respectively. Taken together, 1,4-BQ caused lineage-related cytotoxicity in mouse HPCs, demonstrating greater toxicity in single-lineage progenitors than in those of multi-lineage. - Highlights: • We examine 1,4-BQ toxicity targeting mouse hematopoietic cell lineages. • 1,4-BQ induces concentration-dependent cytotoxicity in bone marrow (BM) cells. • 1,4-BQ shows lineage-related toxicity on hematopoietic stem and

  8. Lineage-related cytotoxicity and clonogenic profile of 1,4-benzoquinone-exposed hematopoietic stem and progenitor cells

    International Nuclear Information System (INIS)

    Chow, Paik Wah; Abdul Hamid, Zariyantey; Chan, Kok Meng; Inayat-Hussain, Salmaan Hussain; Rajab, Nor Fadilah

    2015-01-01

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) are sensitive targets for benzene-induced hematotoxicity and leukemogenesis. The impact of benzene exposure on the complex microenvironment of HSCs and HPCs remains elusive. This study aims to investigate the mechanism linking benzene exposure to targeting HSCs and HPCs using phenotypic and clonogenic analyses. Mouse bone marrow (BM) cells were exposed ex vivo to the benzene metabolite, 1,4-benzoquinone (1,4-BQ), for 24 h. Expression of cellular surface antigens for HSC (Sca-1), myeloid (Gr-1, CD11b), and lymphoid (CD45, CD3e) populations were confirmed by flow cytometry. The clonogenicity of cells was studied using the colony-forming unit (CFU) assay for multilineage (CFU-GM and CFU-GEMM) and single-lineage (CFU-E, BFU-E, CFU-G, and CFU-M) progenitors. 1,4-BQ demonstrated concentration-dependent cytotoxicity in mouse BM cells. The percentage of apoptotic cells increased (p < 0.05) following 1,4-BQ exposure. Exposure to 1,4-BQ showed no significant effect on CD3e + cells but reduced the total counts of Sca-1 + , CD11b + , Gr-1 + , and CD45 + cells at 7 and 12 μM (p < 0.05). Furthermore, the CFU assay showed reduced (p < 0.05) clonogenicity in 1,4-BQ-treated cells. 1,4-BQ induced CFU-dependent cytotoxicity by significantly inhibiting colony growth for CFU-E, BFU-E, CFU-G, and CFU-M starting at a low concentration of exposure (5 μM); whereas for the CFU-GM and CFU-GEMM, the inhibition of colony growth was remarkable only at 7 and 12 μM of 1,4-BQ, respectively. Taken together, 1,4-BQ caused lineage-related cytotoxicity in mouse HPCs, demonstrating greater toxicity in single-lineage progenitors than in those of multi-lineage. - Highlights: • We examine 1,4-BQ toxicity targeting mouse hematopoietic cell lineages. • 1,4-BQ induces concentration-dependent cytotoxicity in bone marrow (BM) cells. • 1,4-BQ shows lineage-related toxicity on hematopoietic stem and progenitors. • 1,4-BQ

  9. Repair and cell cycle response in cells exposed to environmental biohazards. Progress report, February 1, 1976--May 31, 1977

    International Nuclear Information System (INIS)

    Billen, D.

    1977-01-01

    Progress is reported on the following research projects: DNA polymerase III dependent repair of x-ray damage in Escherichia coli; regulation of reinsertation of nucleotides by DNA ligase; DNA synthesis in permeabilized CHO cells; measurement of damage to DNA in Bacillus subtilis; repair defect in rec A cells; inactivation of transforming DNA; and mutagenesis of transforming DNA

  10. Reactive oxygen species regulated mitochondria-mediated apoptosis in PC12 cells exposed to chlorpyrifos

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jeong Eun [Department of Pharmacology, College of Medicine, Hanyang University, Seoul (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Park, Jae Hyeon [Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of); Shin, In Chul [Department of Pharmacology, College of Medicine, Hanyang University, Seoul (Korea, Republic of); Koh, Hyun Chul, E-mail: hckoh@hanyang.ac.kr [Department of Pharmacology, College of Medicine, Hanyang University, Seoul (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of)

    2012-09-01

    Reactive oxidative species (ROS) generated by environmental toxicants including pesticides could be one of the factors underlying the neuronal cell damage in neurodegenerative diseases. In this study we found that chlorpyrifos (CPF) induced apoptosis in dopaminergic neuronal components of PC12 cells as demonstrated by the activation of caspases and nuclear condensation. Furthermore, CPF also reduced the tyrosine hydroxylase-positive immunoreactivity in substantia nigra of the rat. In addition, CPF induced inhibition of mitochondrial complex I activity. Importantly, N-acetyl cysteine (NAC) treatment effectively blocked apoptosis via the caspase-9 and caspase-3 pathways while NAC attenuated the inhibition of mitochondrial complex I activity as well as the oxidative metabolism of dopamine (DA). These results demonstrated that CPF-induced apoptosis was involved in mitochondrial dysfunction through the production of ROS. In the response of cellular antioxidant systems to CPF, we found that CPF treatment increased HO-1 expression while the expression of CuZnSOD and MnSOD was reduced. In addition, we found that CPF treatment activated MAPK pathways, including ERK 1/2, the JNK, and the p38 MAP kinase in a time-dependent manner. NAC treatment abolished MAPK phosphorylation caused by CPF, indicating that ROS are upstream signals of MAPK. Interestingly, MAPK inhibitors abolished cytotoxicity and reduced ROS generation by CPF treatment. Our results demonstrate that CPF induced neuronal cell death in part through MAPK activation via ROS generation, suggesting its potential to generate oxidative stress via mitochondrial damage and its involvement in oxidative stress-related neurodegenerative disease. -- Highlights: ► Chlorpyrifos induces apoptosis. ► Chlorpyrifos inhibits mitochondrial complex I activity. ► ROS is involved in chlorpyrifos-induced apoptosis. ► Chlorpyrifos affects cellular antioxidant systems. ► Chlorpyrifos-induced apoptosis mediates activation of MAPK.

  11. Reactive oxygen species regulated mitochondria-mediated apoptosis in PC12 cells exposed to chlorpyrifos

    International Nuclear Information System (INIS)

    Lee, Jeong Eun; Park, Jae Hyeon; Shin, In Chul; Koh, Hyun Chul

    2012-01-01

    Reactive oxidative species (ROS) generated by environmental toxicants including pesticides could be one of the factors underlying the neuronal cell damage in neurodegenerative diseases. In this study we found that chlorpyrifos (CPF) induced apoptosis in dopaminergic neuronal components of PC12 cells as demonstrated by the activation of caspases and nuclear condensation. Furthermore, CPF also reduced the tyrosine hydroxylase-positive immunoreactivity in substantia nigra of the rat. In addition, CPF induced inhibition of mitochondrial complex I activity. Importantly, N-acetyl cysteine (NAC) treatment effectively blocked apoptosis via the caspase-9 and caspase-3 pathways while NAC attenuated the inhibition of mitochondrial complex I activity as well as the oxidative metabolism of dopamine (DA). These results demonstrated that CPF-induced apoptosis was involved in mitochondrial dysfunction through the production of ROS. In the response of cellular antioxidant systems to CPF, we found that CPF treatment increased HO-1 expression while the expression of CuZnSOD and MnSOD was reduced. In addition, we found that CPF treatment activated MAPK pathways, including ERK 1/2, the JNK, and the p38 MAP kinase in a time-dependent manner. NAC treatment abolished MAPK phosphorylation caused by CPF, indicating that ROS are upstream signals of MAPK. Interestingly, MAPK inhibitors abolished cytotoxicity and reduced ROS generation by CPF treatment. Our results demonstrate that CPF induced neuronal cell death in part through MAPK activation via ROS generation, suggesting its potential to generate oxidative stress via mitochondrial damage and its involvement in oxidative stress-related neurodegenerative disease. -- Highlights: ► Chlorpyrifos induces apoptosis. ► Chlorpyrifos inhibits mitochondrial complex I activity. ► ROS is involved in chlorpyrifos-induced apoptosis. ► Chlorpyrifos affects cellular antioxidant systems. ► Chlorpyrifos-induced apoptosis mediates activation of MAPK.

  12. DNA methylation analysis in rat kidney epithelial cells exposed to 3-MCPD and glycidol.

    Science.gov (United States)

    Senyildiz, Mine; Alpertunga, Buket; Ozden, Sibel

    2017-10-01

    3-Monochloropropane-1,2-diol (3-MCPD) is a well-known food processing contaminant that has been regarded as a rat carcinogen, which is known to induce Leydig-cell and mammary gland tumors in males, as well as kidney tumors in both genders. 3-MCPD is highly suspected to be a non-genotoxic carcinogen. 2,3-Epoxy-1-propanol (glycidol) can be formed via dehalogenation from 3-MCPD. We aimed to investigate the cytotoxic effects of 3-MCPD and glycidol, then to demonstrate the possible epigenetic mechanisms with global and gene-specific DNA methylation in rat kidney epithelial cells (NRK-52E). IC 50 value of 3-MCPD was determined as 48 mM and 41.39 mM, whereas IC 50 value of glycidol was 1.67 mM and 1.13 mM by MTT and NRU test, respectively. Decreased global DNA methylation at the concentrations of 100 μM and 1000 μM for 3-MCPD and 100 μM and 500 μM for glycidol were observed after 48 h exposure by using 5-methylcytosine (5-mC) ELISA kit. Methylation changes were detected in promoter regions of c-myc and Rassf1a in 3-MCPD and glycidol treated NRK-52E cells by using methylation-specific PCR (MSP), whereas changes on gene expression of c-myc and Rassf1a were observed by using real-time PCR. However, e-cadherin, p16, VHL and p15 genes were unmethylated in their CpG promoter regions in response to treatment with 3-MCPD and glycidol. Alterations in DNA methylation might be key events in the toxicity of 3-MCPD and glycidol.

  13. Recovery of ovary size, follicle cell apoptosis, and HSP70 expression in fish exposed to bleached pulp mill effluent

    Energy Technology Data Exchange (ETDEWEB)

    Janz, D. M.; Weber, L. P. [Oklahoma State Univ., Stillwater, OK (United States); McMaster, M. E.; Munkittrrick, K. R. [Environment Canada, Burlington, ON (Canada); Van Der Kraak, G. [Guelph Univ., Dept. of Zoology, ON (Canada)

    2001-03-01

    Apoptosis of granulosa cells that provide hormonal support for the oocyte is the normal mechanism by which atresia ( reduced ovarian size, decreased fecundity, delayed sexual maturation, alterations in plasma sex steroid levels, etc) occurs in mammals, birds and possibly fish. The objective of this study is to determine ovarian cell apoptosis, gonadosomatic index (GSI) and heat shock protein (HSP70) expression during the growth stage of ovarian development in white sucker fish in order to compare samples of fish collected upstream and downstream of a bleached kraft pulp mill in Ontario. Fish for the study were collected in two different years, before and after the pulp mill undertook a number of improvements to eliminate the release of process chemicals. Results showed a 3.4-fold increase in ovarian cell apoptosis in growing white sucker collected four km downstream of the bleached kraft pulp mill in 1996 (before the improvements) compared to fish collected from upstream sources. The elevated ovarian cell apoptosis was associated with significant reduction in gonadosomatic index in fish collected downstream. There were no differences in ovarian cell apoptosis or gonadosomatic index between fish collected upstream and four km downstream of the mill in September 1998 (after the improvements.) Based on the results, it may be concluded that chronic stimulation of ovarian cell apoptosis by certain components of bleached kraft pulp mill effluents represents an important cellular mechanism for reducing the size of ovaries and other related reproductive responses in female fish exposed to these effluents. Although the specific effluent components are not known, the improvements undertaken between 1996 and 1998 resulted in significant enough recovery of these responses to justify the belief in a cause-effect relationship. 32 refs., 1 tab., 2 figs.

  14. Gamma-H2AX foci in cells exposed to a mixed beam of X-rays and alpha particles

    Science.gov (United States)

    2012-01-01

    Background Little is known about the cellular effects of exposure to mixed beams of high and low linear energy transfer radiation. So far, the effects of combined exposures have mainly been assessed with clonogenic survival or cytogenetic methods, and the results are contradictory. The gamma-H2AX assay has up to now not been applied in this context, and it is a promising tool for investigating the early cellular response to mixed beam irradiation. Purpose To determine the dose response and repair kinetics of gamma-H2AX ionizing radiation-induced foci in VH10 human fibroblasts exposed to mixed beams of 241Am alpha particles and X-rays. Results VH10 human fibroblasts were irradiated with each radiation type individually or both in combination at 37°C. Foci were scored for repair kinetics 0.5, 1, 3 and 24 h after irradiation (one dose per irradiation type), and for dose response at the 1 h time point. The dose response effect of mixed beam was additive, and the relative biological effectiveness for alpha particles (as compared to X-rays) was of 0.76 ± 0.52 for the total number of foci, and 2.54 ± 1.11 for large foci. The repair kinetics for total number of foci in cells exposed to mixed beam irradiation was intermediate to that of cells exposed to alpha particles and X-rays. However, for mixed beam-irradiated cells the frequency and area of large foci were initially lower than predicted and increased during the first 3 hours of repair (while the predicted number and area did not). Conclusions The repair kinetics of large foci after mixed beam exposure was significantly different from predicted based on the effect of the single dose components. The formation of large foci was delayed and they did not reach their maximum area until 1 h after irradiation. We hypothesize that the presence of low X-ray-induced damage engages the DNA repair machinery leading to a delayed DNA damage response to the more complex DNA damage induced by alpha particles. PMID:23121736

  15. Osteoblastic differentiation and stress response of human mesenchymal stem cells exposed to alternating current electric fields

    Directory of Open Access Journals (Sweden)

    Kaplan David L

    2011-01-01

    Full Text Available Abstract Background Electric fields are integral to many biological events, from maintaining cellular homeostasis to embryonic development to healing. The application of electric fields offers substantial therapeutic potential, while optimal dosing regimens and the underlying mechanisms responsible for the positive clinical impact are poorly understood. Methods The purpose of this study was to track the differentiation profile and stress response of human bone marrow derived mesenchymal stem cells (hMSCs undergoing osteogenic differentiation during exposure to a 20 mV/cm, 60 kHz electric field. Morphological and biochemical changes were imaged using endogenous two-photon excited fluorescence (TPEF and quantitatively assessed through eccentricity calculations and extraction of the redox ratio from NADH, FAD and lipofuscin contributions. Real time reverse transcriptase-polymerase chain reactions (RT-PCR were used to track osteogenic differentiation markers, namely alkaline phosphatase (ALP and collagen type 1 (col1, and stress response markers, such as heat shock protein 27 (hsp27 and heat shock protein 70 (hsp70. Comparisons of collagen deposition between the stimulated hMSCs and controls were examined through second harmonic generation (SHG imaging. Results Quantitative differences in cell morphology, as described through an eccentricity ratio, were found on days 2 and days 5 (p Conclusions Electrical stimulation is a useful tool to improve hMSC osteogenic differentiation, while heat shock proteins may reveal underlying mechanisms, and optical non-invasive imaging may be used to monitor the induced morphological and biochemical changes.

  16. Cell cycle arrest and gene expression profiling of testis in mice exposed to fluoride.

    Science.gov (United States)

    Su, Kai; Sun, Zilong; Niu, Ruiyan; Lei, Ying; Cheng, Jing; Wang, Jundong

    2017-05-01

    Exposure to fluoride results in low reproductive capacity; however, the mechanism underlying the impact of fluoride on male productive system still remains obscure. To assess the potential toxicity in testis of mice administrated with fluoride, global genome microarray and real-time PCR were performed to detect and identify the altered transcriptions. The results revealed that 763 differentially expressed genes were identified, including 330 up-regulated and 433 down-regulated genes, which were involved in spermatogenesis, apoptosis, DNA damage, DNA replication, and cell differentiation. Twelve differential expressed genes were selected to confirm the microarray results using real-time PCR, and the result kept the same tendency with that of microarray. Furthermore, compared with the control group, more apoptotic spermatogenic cells were observed in the fluoride group, and the spermatogonium were markedly increased in S phase and decreased in G2/M phase by fluoride. Our findings suggested global genome microarray provides an insight into the reproductive toxicity induced by fluoride, and several important biological clues for further investigations. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1558-1565, 2017. © 2016 Wiley Periodicals, Inc.

  17. Cigarette Smoke-Exposed Candida albicans Increased Chitin Production and Modulated Human Fibroblast Cell Responses

    Directory of Open Access Journals (Sweden)

    Humidah Alanazi

    2014-01-01

    Full Text Available The predisposition of cigarette smokers for development of respiratory and oral bacterial infections is well documented. Cigarette smoke can also contribute to yeast infection. The aim of this study was to investigate the effect of cigarette smoke condensate (CSC on C. albicans transition, chitin content, and response to environmental stress and to examine the interaction between CSC-pretreated C. albicans and normal human gingival fibroblasts. Following exposure to CSC, C. albicans transition from blastospore to hyphal form increased. CSC-pretreated yeast cells became significantly (P<0.01 sensitive to oxidation but significantly (P<0.01 resistant to both osmotic and heat stress. CSC-pretreated C. albicans expressed high levels of chitin, with 2- to 8-fold recorded under hyphal conditions. CSC-pretreated C. albicans adhered better to the gingival fibroblasts, proliferated almost three times more and adapted into hyphae, while the gingival fibroblasts recorded a significantly (P<0.01 slow growth rate but a significantly higher level of IL-1β when in contact with CSC-pretreated C. albicans. CSC was thus able to modulate both C. albicans transition through the cell wall chitin content and the interaction between C. albicans and normal human gingival fibroblasts. These findings may be relevant to fungal infections in the oral cavity in smokers.

  18. BAG3 sensitizes cancer cells exposed to DNA damaging agents via direct interaction with GRP78.

    Science.gov (United States)

    Kong, De-Hui; Zhang, Qiang; Meng, Xin; Zong, Zhi-Hong; Li, Chao; Liu, Bao-Qin; Guan, Yifu; Wang, Hua-Qin

    2013-12-01

    Bcl-2 associated athanogene 3 (BAG3) has a modular structure that contains a BAG domain, a WW domain, a proline-rich (PxxP) domain to mediate potential interactions with chaperons and other proteins that participate in more than one signal transduction. In search for novel interacting partners, the current study identified that 78kDa glucose-regulated protein (GRP78) was a novel partner interacting with BAG3. Interaction between GRP78 and BAG3 was confirmed by coimmunoprecipitation and glutathione S-transferase (GST) pulldown. We also identified that the ATPase domain of GRP78 and BAG domain of BAG3 mediated their interaction. Counterintuitive for a prosurvival protein, BAG3 was found to promote the cytotoxicity of breast cancer MCF7, thyroid cancer FRO and glioma U87 cells subjected to genotoxic stress. In addition, the current study demonstrated that BAG3 interfered with the formation of the antiapoptotic GRP78-procaspase-7 complex, which resulted in an increased genotoxic stress-induced cytotoxicity in cancer cells. Furthermore, overexpression of GRP78 significantly blocked the enhancing effects of BAG3 on activation of caspase-7 and induction of apoptosis by genotoxic stress. Overall, these results suggested that through direct interaction BAG3 could prevent the antiapoptotic effect of GRP78 upon genotoxic stress. © 2013.

  19. Cell density dependence of transformation frequencies in C3H10T1/2 cells exposed to X-rays

    Energy Technology Data Exchange (ETDEWEB)

    Bettega, D; Calzolari, P; Ottolenghi, A; Lombardi, L T [Milan Univ. (Italy). Ist. di Fisica; Rimoldi, E [Milan Univ. (Italy). Ist. di Radiologia Veterinaria

    1989-12-01

    The effects of cell density on transformation frequencies were studied in C3H10T1/2 cells exposed to 0.5 and 7 Gy of 200 kVp X-rays. Initial cell density strongly influenced transformation frequency; this decreased by a factor of between 4 and 10 when the initial seeding density was changed from 50 to 2500 cells/10 cm diameter Petri dish. The data were fitted with two equations: (a) an allometric function represented on a log-log scale by a straight line and (b) a sigmoidal function with plateaux between 50 and 250 cells/dish and above 600. The two curves are compared and their probabilities discussed. Our data indicate that the region between 50 and 250 cells/dish would be the most suitable region for dose-effect measurements. A study of the growth curves at 0.5 and 8.5 Gy shows that cell growth rates are not influenced by initial cell density. (author).

  20. Skin cell isolation and expansion for cell transplantation is limited in patients using tobacco, alcohol, or are exhibiting diabetes mellitus.

    Science.gov (United States)

    Johnen, Christa; Hartmann, Bernd; Steffen, Ingo; Bräutigam, Kirsten; Witascheck, Tom; Toman, Nidal; Küntscher, Markus V; Gerlach, Jörg C

    2006-03-01

    The aim of this exploratory study was to investigate the isolation and expansion of keratinocytes and fibroblasts from donors with certain medical histories. Biopsies were taken from donors (N=32) falling into one or more of the following categories: a history of heavy smoking and/or alcohol abuse, drug abuse, diabetes mellitus or steroid treatment. Cells from donors who did not fall into any of the above-mentioned categories were used as controls. Proliferation and growth behaviour of cells were analyzed by measurement of passage duration, absorbance (MTT-assay) and light microscopy. Donors with a specific medical history required larger biopsy areas than the control group for isolating a sufficient number of fibroblasts and keratinocytes. Times to confluence were significantly prolonged and absorbances (MTT) were significantly reduced in several donor groups when compared to control cultures. Biopsies from donors with steroid treatment, drug abuse and combined nicotine and alcohol abuse could not be established beyond passage 0 degrees or 1 degree, respectively. We conclude that isolation and expansion of skin cells from donors with certain medical histories may require larger biopsies, prolonged expansion times or may even result in failure. These findings may therefore be of clinical importance in the field of autologous skin cell transplantation.

  1. Youth access to tobacco.

    Science.gov (United States)

    Rigotti, N A

    1999-01-01

    To start smoking, young people need a supply of tobacco products. Reducing youth access to tobacco is a new approach to preventing tobacco use that has been a focus of federal, state, and local tobacco control efforts over the past decade. All 50 states ban tobacco sales to minors, but compliance is poor because laws are not enforced. Consequently, young people have little trouble obtaining tobacco products. Commercial sources of tobacco (stores and vending machines) are important for underage smokers, who often purchase their own cigarettes. Underage youths also obtain tobacco from noncommercial sources such as friends, relatives, older adolescents, and adults. Educating retailers about tobacco sales laws has not produced long-term improvement in their compliance. Active enforcement of tobacco sales laws changes retailer behavior, but whether this reduces young people's access to tobacco or their tobacco use is not clear. The effectiveness of new local, state, and federal actions that aim to reduce youth access to tobacco remains to be determined. Can enforcing tobacco sales laws reduce young people's access to tobacco? If so, will this prevent or delay the onset of their tobacco use? How will youths' sources of tobacco change as commercial sources are restricted? What are the social (noncommercial) sources of tobacco for minors and how can youths' access to tobacco from these sources be reduced? What is the impact of the new federal policies aimed at reducing youth access to tobacco? Do new state and local laws that ban youth possession or use of tobacco have a net positive or negative impact on youth attitudes, access to tobacco, or tobacco use? What is the relative effectiveness and cost-effectiveness of efforts to reduce the supply of tobacco compared to those that aim to reduce demand for tobacco? Will either work alone or are both necessary to achieve reductions in youth smoking?

  2. Young adults' interpretations of tobacco brands: implications for tobacco control.

    Science.gov (United States)

    Gendall, Philip; Hoek, Janet; Thomson, George; Edwards, Richard; Pene, Gina; Gifford, Heather; Pirikahu, Gill; McCool, Judith

    2011-10-01

    Marketers have long recognized the power and importance of branding, which creates aspirational attributes that increase products' attractiveness. Although brand imagery has traditionally been communicated via mass media, packaging's importance in promoting desirable brand-attribute associations has increased. Knowledge of how groups prone to smoking experimentation interpret tobacco branding would inform the debate over plain packaging currently occurring in many countries. We conducted 12 group discussions and four in-depth interviews with 66 young adult smokers and nonsmokers of varying ethnicities from two larger New Zealand cities and one provincial city. Participants evaluated 10 familiar and unfamiliar tobacco brands using brand personality attributes and discussed the associations they had made. Participants ascribed very different images to different brands when exposed to the packaging alone, regardless of whether they had seen or heard of the brands before. Perceptual mapping of brands and image attributes highlighted how brand positions varied from older, more traditional, and male oriented to younger, feminine, and "cool." Our findings emphasize the continuing importance of tobacco branding as a promotion tool, even when communicated only by packaging. The ease with which packaging alone enabled young people to identify brand attributes and the desirable associations these connoted illustrate how tobacco packaging functions as advertising. The results support measures such as plain packaging of tobacco products to reduce exposure to these overt behavioral cues.

  3. Repair and cell cycle response in cells exposed to environmental biohazards. Final report, January 1, 1973-December 31, 1984

    International Nuclear Information System (INIS)

    Hadden, C.T.; Billen, D.

    1986-01-01

    These studies have focussed on agents which cause damage to DNA leading to inhibition of DNA synthesis or faulty DNA replication or repair. The overall goal of this project has been to understand how environmental agents interact with the DNA of cells and how cells cope with any resulting damage. In particular we have been concerned with the nature of the repair systems involved in restoration of damaged DNA and the cellular responses to radiation or chemical damage

  4. Poly (ADP-ribose) catabolism in mammalian cells exposed to DNA-damaging agents

    International Nuclear Information System (INIS)

    Alvarez-Gonzalez, R.; Althaus, F.R.

    1989-01-01

    DNA damage inflicted by the alkylating agens N-methyl-N-nitro-N-nitrosoquanidine, or by UV stimulated the catabolism of protein-bound poly (ADP-ribose) in the chromatin of cultured hepatocytes. The stimulation was highest at the largest doses of DNA-damaging treatment. As a consequence, the half-life of ADP-ribosyl polymers may drop to less than 41 s. This rapid turnover contrasts with the slow catabolism of a constitutive fraction of polymers exhibiting a half-life of 7.7 h. These data suggest that post-incisional stimulation of poly (ADP-ribose) biosynthesis in DNA-excision repair is coupled with an adaptation of poly (ADP-ribose) catabolism in mammalian cells. (Author). 37 refs.; 3 figs

  5. Analysis of epigenetic factors in mouse embryonic neural stem cells exposed to hyperglycemia.

    Directory of Open Access Journals (Sweden)

    Sukanya Shyamasundar

    Full Text Available BACKGROUND: Maternal diabetes alters gene expression leading to neural tube defects (NTDs in the developing brain. The mechanistic pathways that deregulate the gene expression remain unknown. It is hypothesized that exposure of neural stem cells (NSCs to high glucose/hyperglycemia results in activation of epigenetic mechanisms which alter gene expression and cell fate during brain development. METHODS AND FINDINGS: NSCs were isolated from normal pregnancy and streptozotocin induced-diabetic pregnancy and cultured in physiological glucose. In order to examine hyperglycemia induced epigenetic changes in NSCs, chromatin reorganization, global histone status at lysine 9 residue of histone H3 (acetylation and trimethylation and global DNA methylation were examined and found to be altered by hyperglycemia. In NSCs, hyperglycemia increased the expression of Dcx (Doublecortin and Pafah1b1 (Platelet activating factor acetyl hydrolase, isoform 1b, subunit 1 proteins concomitant with decreased expression of four microRNAs (mmu-miR-200a, mmu-miR-200b, mmu-miR-466a-3p and mmu-miR-466 d-3p predicted to target these genes. Knockdown of specific microRNAs in NSCs resulted in increased expression of Dcx and Pafah1b1 proteins confirming target prediction and altered NSC fate by increasing the expression of neuronal and glial lineage markers. CONCLUSION/INTERPRETATION: This study revealed that hyperglycemia alters the epigenetic mechanisms in NSCs, resulting in altered expression of some development control genes which may form the basis for the NTDs. Since epigenetic changes are reversible, they may be valuable therapeutic targets in order to improve fetal outcomes in diabetic pregnancy.

  6. Live-cell imaging of dual-labeled Golgi stacks in tobacco BY-2 cells reveals similar behaviors for different cisternae during movement and brefeldin A treatment.

    Science.gov (United States)

    Madison, Stephanie L; Nebenführ, Andreas

    2011-09-01

    In plant cells, the Golgi apparatus consists of numerous stacks that, in turn, are composed of several flattened cisternae with a clear cis-to-trans polarity. During normal functioning within living cells, this unusual organelle displays a wide range of dynamic behaviors such as whole stack motility, constant membrane flux through the cisternae, and Golgi enzyme recycling through the ER. In order to further investigate various aspects of Golgi stack dynamics and integrity, we co-expressed pairs of established Golgi markers in tobacco BY-2 cells to distinguish sub-compartments of the Golgi during monensin treatments, movement, and brefeldin A (BFA)-induced disassembly. A combination of cis and trans markers revealed that Golgi stacks remain intact as they move through the cytoplasm. The Golgi stack orientation during these movements showed a slight preference for the cis side moving ahead, but trans cisternae were also found at the leading edge. During BFA treatments, the different sub-compartments of about half of the observed stacks fused with the ER sequentially; however, no consistent order could be detected. In contrast, the ionophore monensin resulted in swelling of trans cisternae while medial and particularly cis cisternae were mostly unaffected. Our results thus demonstrate a remarkable equivalence of the different cisternae with respect to movement and BFA-induced fusion with the ER. In addition, we propose that a combination of dual-label fluorescence microscopy and drug treatments can provide a simple alternative approach to the determination of protein localization to specific Golgi sub-compartments.

  7. Tobacco-Related Mortality

    Science.gov (United States)

    ... Exposure is High in Multiunit Housing Smokeless Products Electronic Cigarettes Youth Tobacco Prevention Tobacco Products Tobacco Ingredient ... 2004 [accessed 2015 Aug 17]. National Cancer Institute. Cigars: Health Effects and Trends [ PDF –2.93 MB] . ...

  8. Risks of tobacco

    Science.gov (United States)

    Secondhand smoke - risks; Cigarette smoking - risks; Smoking and smokeless tobacco - risks; Nicotine - risks ... tobacco that are known to cause cancer. HEALTH RISKS OF SMOKING OR USING SMOKELESS TOBACCO Knowing the ...

  9. Genotoxic and Cytotoxic Effects on the Immune Cells of the Freshwater Bivalve Dreissena polymorpha Exposed to the Environmental Neurotoxin BMAA.

    Science.gov (United States)

    Lepoutre, Alexandra; Milliote, Nadia; Bonnard, Marc; Palos-Ladeiro, Mélissa; Rioult, Damien; Bonnard, Isabelle; Bastien, Fanny; Faassen, Elisabeth; Geffard, Alain; Lance, Emilie

    2018-03-01

    The environmental neurotoxin β- N -Methylamino-l-alanine (BMAA) has been pointed out to be involved in human neurodegenerative diseases. This molecule is known to be bioaccumulated by bivalves. However, little data about its toxic effects on freshwater mussels is available, particularly on the hemolymphatic compartment and its hemocyte cells involved in various physiological processes such as immune defenses, digestion and excretion, tissue repair, and shell production. Here we exposed Dreissena polymorpha to dissolved BMAA, at the environmental concentration of 7.5 µg of /mussel/3 days, during 21 days followed by 14 days of depuration in clear water, with the objective of assessing the BMAA presence in the hemolymphatic compartment, as well as the impact of the hemocyte cells in terms of potential cytotoxicity, immunotoxicity, and genotoxiciy. Data showed that hemocytes were in contact with BMAA. The presence of BMAA in hemolymph did not induce significant effect on hemocytes phagocytosis activity. However, significant DNA damage on hemocytes occurred during the first week (days 3 and 8) of BMAA exposure, followed by an increase of hemocyte mortality after 2 weeks of exposure. Those effects might be an indirect consequence of the BMAA-induced oxidative stress in cells. However, DNA strand breaks and mortality did not persist during the entire exposure, despite the BMAA persistence in the hemolymph, suggesting potential induction of some DNA-repair mechanisms.

  10. NFAT regulation of cystathionine γ-lyase expression in endothelial cells is impaired in rats exposed to intermittent hypoxia.

    Science.gov (United States)

    Gonzalez Bosc, Laura V; Osmond, Jessica M; Giermakowska, Wieslawa K; Pace, Carolyn E; Riggs, Jennifer L; Jackson-Weaver, Olan; Kanagy, Nancy L

    2017-04-01

    Sleep apnea is a risk factor for cardiovascular disease, and intermittent hypoxia (IH, 20 episodes/h of 5% O 2 -5% CO 2 for 7 h/day) to mimic sleep apnea increases blood pressure and impairs hydrogen sulfide (H 2 S)-induced vasodilation in rats. The enzyme that produces H 2 S, cystathionine γ-lyase (CSE), is decreased in rat mesenteric artery endothelial cells (EC) following in vivo IH exposure. In silico analysis identified putative nuclear factor of activated T cell (NFAT) binding sites in the CSE promoter. Therefore, we hypothesized that IH exposure reduces Ca 2+ concentration ([Ca 2+ ]) activation of calcineurin/NFAT to lower CSE expression and impair vasodilation. In cultured rat aortic EC, inhibiting calcineurin with cyclosporine A reduced CSE mRNA, CSE protein, and luciferase activity driven by a full-length but not a truncated CSE promoter. In male rats exposed to sham or IH conditions for 2 wk, [Ca 2+ ] in EC in small mesenteric arteries from IH rats was lower than in EC from sham rat arteries (Δfura 2 ratio of fluorescence at 340 to 380 nm from Ca 2+ free: IH = 0.05 ± 0.02, sham = 0.17 ± 0.03, P intermittent hypoxia to mimic sleep apnea, nuclear factor of activated T cells c3 nuclear translocation and CSE expression are decreased, concomitant with decreased CSE-dependent vasodilation. Copyright © 2017 the American Physiological Society.

  11. Stress proteins and oxidative damage in a renal derived cell line exposed to inorganic mercury and lead

    International Nuclear Information System (INIS)

    Stacchiotti, Alessandra; Morandini, Fausta; Bettoni, Francesca; Schena, Ilaria; Lavazza, Antonio; Grigolato, Pier Giovanni; Apostoli, Pietro; Rezzani, Rita; Aleo, Maria Francesca

    2009-01-01

    A close link between stress protein up-regulation and oxidative damage may provide a novel therapeutic tool to counteract nephrotoxicity induced by toxic metals in the human population, mainly in children, of industrialized countries. Here we analysed the time course of the expression of several heat shock proteins, glucose-regulated proteins and metallothioneins in a rat proximal tubular cell line (NRK-52E) exposed to subcytotoxic doses of inorganic mercury and lead. Concomitantly, we used morphological and biochemical methods to evaluate metal-induced cytotoxicity and oxidative damage. In particular, as biochemical indicators of oxidative stress we detected reactive oxygen species (ROS) and nitrogen species (RNS), total glutathione (GSH) and glutathione-S-transferase (GST) activity. Our results clearly demonstrated that mercury increases ROS and RNS levels and the expressions of Hsp25 and inducible Hsp72. These findings are corroborated by evident mitochondrial damage, apoptosis or necrosis. By contrast, lead is unable to up-regulate Hsp72 but enhances Grp78 and activates nuclear Hsp25 translocation. Furthermore, lead causes endoplasmic reticulum (ER) stress, vacuolation and nucleolar segregation. Lastly, both metals stimulate the over-expression of MTs, but with a different time course. In conclusion, in NRK-52E cell line the stress response is an early and metal-induced event that correlates well with the direct oxidative damage induced by mercury. Indeed, different chaperones are involved in the specific nephrotoxic mechanism of these environmental pollutants and work together for cell survival.

  12. Activation of eNOS in endothelial cells exposed to ionizing radiation involves components of the DNA damage response pathway

    Energy Technology Data Exchange (ETDEWEB)

    Nagane, Masaki; Yasui, Hironobu; Sakai, Yuri; Yamamori, Tohru [Laboratory of Radiation Biology, Department of Environmental Veterinary Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan); Niwa, Koichi [Laboratory of Biochemistry, Department of Food and Cosmetic Science, Faculty of Bioindustry, Tokyo University of Agriculture, Abashiri 099-2493 (Japan); Hattori, Yuichi [Department of Molecular and Medical Pharmacology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); Kondo, Takashi [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); Inanami, Osamu, E-mail: inanami@vetmed.hokudai.ac.jp [Laboratory of Radiation Biology, Department of Environmental Veterinary Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan)

    2015-01-02

    Highlights: • eNOS activity is increased in BAECs exposed to X-rays. • ATM is involved in this increased eNOS activity. • HSP90 modulates the radiation-induced activation of ATM and eNOS. - Abstract: In this study, the involvement of ataxia telangiectasia mutated (ATM) kinase and heat shock protein 90 (HSP90) in endothelial nitric oxide synthase (eNOS) activation was investigated in X-irradiated bovine aortic endothelial cells. The activity of nitric oxide synthase (NOS) and the phosphorylation of serine 1179 of eNOS (eNOS-Ser1179) were significantly increased in irradiated cells. The radiation-induced increases in NOS activity and eNOS-Ser1179 phosphorylation levels were significantly reduced by treatment with either an ATM inhibitor (Ku-60019) or an HSP90 inhibitor (geldanamycin). Geldanamycin was furthermore found to suppress the radiation-induced phosphorylation of ATM-Ser1181. Our results indicate that the radiation-induced eNOS activation in bovine aortic endothelial cells is regulated by ATM and HSP90.

  13. Genotoxic and Cytotoxic Effects on the Immune Cells of the Freshwater Bivalve Dreissena polymorpha Exposed to the Environmental Neurotoxin BMAA

    Directory of Open Access Journals (Sweden)

    Alexandra Lepoutre

    2018-03-01

    Full Text Available The environmental neurotoxin β-N-Methylamino-l-alanine (BMAA has been pointed out to be involved in human neurodegenerative diseases. This molecule is known to be bioaccumulated by bivalves. However, little data about its toxic effects on freshwater mussels is available, particularly on the hemolymphatic compartment and its hemocyte cells involved in various physiological processes such as immune defenses, digestion and excretion, tissue repair, and shell production. Here we exposed Dreissena polymorpha to dissolved BMAA, at the environmental concentration of 7.5 µg of /mussel/3 days, during 21 days followed by 14 days of depuration in clear water, with the objective of assessing the BMAA presence in the hemolymphatic compartment, as well as the impact of the hemocyte cells in terms of potential cytotoxicity, immunotoxicity, and genotoxiciy. Data showed that hemocytes were in contact with BMAA. The presence of BMAA in hemolymph did not induce significant effect on hemocytes phagocytosis activity. However, significant DNA damage on hemocytes occurred during the first week (days 3 and 8 of BMAA exposure, followed by an increase of hemocyte mortality after 2 weeks of exposure. Those effects might be an indirect consequence of the BMAA-induced oxidative stress in cells. However, DNA strand breaks and mortality did not persist during the entire exposure, despite the BMAA persistence in the hemolymph, suggesting potential induction of some DNA-repair mechanisms.

  14. Genomic Profiling of a Human Leukemic Monocytic Cell-Line (THP-1 Exposed to Alpha Particle Radiation

    Directory of Open Access Journals (Sweden)

    Vinita Chauhan

    2012-01-01

    Full Text Available This study examined alpha (α- particle radiation effects on global changes in gene expression in human leukemic monocytic cells (THP-1 for the purposes of mining for candidate biomarkers that could be used for the development of a biological assessment tool. THP-1 cells were exposed to α-particle radiation at a dose range of 0 to 1.5 Gy. Twenty-four hours and three days after exposure gene expression was monitored using microarray technology. A total of 16 genes were dose responsive and classified as early onset due to their expression 24 h after exposure. Forty-eight transcripts were dose responsive and classified as late-onset as they were expressed 72 h after exposure. Among these genes, 6 genes were time and dose responsive and validated further using alternate technology. These transcripts were upregulated and associated with biological processes related to immune function, organelle stability and cell signalling/communication. This panel of genes merits further validation to determine if they are strong candidate biomarkers indicative of α-particle exposure.

  15. Attenuation of oxidative stress and inflammation by gravinol in high glucose-exposed renal tubular epithelial cells

    International Nuclear Information System (INIS)

    Kim, You Jung; Kim, Young Ae; Yokozawa, Takako

    2010-01-01

    Gravinol, a proanthocyanidin from grape seeds, has polyphenolic properties with powerful anti-oxidative effects. Although, increasing evidence strongly suggests that polyphenolic antioxidants suppress diabetic nephropathy that is causally associated with oxidative stress and inflammation, gravinol's protective action against diabetic nephropathy has not been fully explored to date. In the current study, we investigated the protective action of gravinol against oxidative stress and inflammation using the experimental diabetic nephropathy cell model, high glucose-exposed renal tubular epithelial cells. To elucidate the underlying actions of gravinol, several oxidative and inflammatory markers were estimated. Included are measurements of lipid peroxidation, total reactive species (RS), superoxide (·O 2 ), nitric oxide (NO·), and peroxynitrite (ONOO - ), as well as nuclear factor-kappa B (NF-κB) nuclear translocation. Results indicate that gravinol had a potent inhibitory action against lipid peroxidation, total RS, ·O 2 , NO·, ONOO - , the reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio and more importantly, against NF-κB nuclear translocation. We propose that gravinol's strong protective effect against high glucose-induced renal tubular epithelial cell damage attenuates diabetic nephropathy by suppressing oxidative stress and inflammation.

  16. Tobacco imagery in video games: ratings and gamer recall.

    Science.gov (United States)

    Forsyth, Susan R; Malone, Ruth E

    2016-09-01

    To assess whether tobacco content found in video games was appropriately labelled for tobacco-related content by the Entertainment and Software Ratings Board (ESRB). Sixty-five gamer participants (self-identified age range 13-50) were interviewed in-person (n=25) or online (n=40) and asked (A) to list favourite games and (B) to name games that they could recall containing tobacco content. The ESRB database was searched for all games mentioned to ascertain whether they had been assigned tobacco-related content descriptors. Games were independently assessed for tobacco content by examining user-created game wiki sites and watching YouTube videos of gameplay. Games with tobacco-related ESRB content descriptors and/or with tobacco imagery verified by researchers were considered to contain tobacco content. Games identified by participants as including tobacco but lacking verifiable tobacco content were treated as not containing tobacco content. Participants recalled playing 140 unique games, of which 118 were listed in the ESRB database. Participants explicitly recalled tobacco content in 31% (37/118) of the games, of which 94% (35/37) included independently verified tobacco content. Only 8% (9/118) of the games had received ESRB tobacco-related content descriptors, but researchers verified that 42% (50/118) contained such content; 42% (49/118) of games were rated 'M' for mature (content deemed appropriate for ages 17+). Of these, 76% (37/49) contained verified tobacco content; however, only 4% (2/49) received ESRB tobacco-related content descriptors. Gamers are exposed to tobacco imagery in many video games. The ESRB is not a reliable source for determining whether video games contain tobacco imagery. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

  17. Sodium nitrite induces acute central nervous system toxicity in guinea pigs exposed to systemic cell-free hemoglobin

    Energy Technology Data Exchange (ETDEWEB)

    Buehler, Paul W.; Butt, Omer I. [Laboratory of Biochemistry and Vascular Biology, Division of Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States); D' Agnillo, Felice, E-mail: felice.dagnillo@fda.hhs.gov [Laboratory of Biochemistry and Vascular Biology, Division of Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States)

    2011-06-10

    Highlights: {yields} Toxicological implications associated with the use of NaNO{sub 2} therapy to treat systemic cell-free Hb exposure are not well-defined. {yields} Systemic Hb exposure followed by NaNO{sub 2} infusion induces acute CNS toxicities in guinea pigs. {yields} These CNS effects were not reproduced by the infusion of cell-free Hb or NaNO{sub 2} alone. {yields} NaNO{sub 2}-mediated oxidation of cell-free Hb may play a causative role in the observed CNS changes. -- Abstract: Systemic cell-free hemoglobin (Hb) released via hemolysis disrupts vascular homeostasis, in part, through the scavenging of nitric oxide (NO). Sodium nitrite (NaNO{sub 2}) therapy can attenuate the hypertensive effects of Hb. However, the chemical reactivity of NaNO{sub 2} with Hb may enhance heme- or iron-mediated toxicities. Here, we investigate the effect of NaNO{sub 2} on the central nervous system (CNS) in guinea pigs exposed to systemic cell-free Hb. Intravascular infusion of NaNO{sub 2}, at doses sufficient to alleviate Hb-mediated blood pressure changes, reduced the expression of occludin, but not zona occludens-1 (ZO-1) or claudin-5, in cerebral tight junctions 4 h after Hb infusion. This was accompanied by increased perivascular heme oxygenase-1 expression, neuronal iron deposition, increased astrocyte and microglial activation, and reduced expression of neuron-specific nuclear protein (NeuN). These CNS changes were not observed in animals treated with Hb or NaNO{sub 2} alone. Taken together, these findings suggest that the use of nitrite salts to treat systemic Hb exposure may promote acute CNS toxicity.

  18. Differential transcriptional regulation of IL-8 expression by human airway epithelial cells exposed to diesel exhaust particles

    International Nuclear Information System (INIS)

    Tal, Tamara L.; Simmons, Steven O.; Silbajoris, Robert; Dailey, Lisa; Cho, Seung-Hyun; Ramabhadran, Ram; Linak, William; Reed, William; Bromberg, Philip A.; Samet, James M.

    2010-01-01

    Exposure to diesel exhaust particles (DEP) induces inflammatory signaling characterized by MAP kinase-mediated activation of NFkB and AP-1 in vitro and in bronchial biopsies obtained from human subjects exposed to DEP. NFkB and AP-1 activation results in the upregulation of genes involved in promoting inflammation in airway epithelial cells, a principal target of inhaled DEP. IL-8 is a proinflammatory chemokine expressed by the airway epithelium in response to environmental pollutants. The mechanism by which DEP exposure induces IL-8 expression is not well understood. In the current study, we sought to determine whether DEP with varying organic content induces IL-8 expression in lung epithelial cells, as well as, to develop a method to rapidly evaluate the upstream mechanism(s) by which DEP induces IL-8 expression. Exposure to DEP with varying organic content differentially induced IL-8 expression and IL-8 promoter activity human airway epithelial cells. Mutational analysis of the IL-8 promoter was also performed using recombinant human cell lines expressing reporters linked to the mutated promoters. Treatment with a low organic-containing DEP stimulated IL-8 expression by a mechanism that is predominantly NFkB-dependent. In contrast, exposure to high organic-containing DEP induced IL-8 expression independently of NFkB through a mechanism that requires AP-1 activity. Our study reveals that exposure to DEP of varying organic content induces proinflammatory gene expression through multiple specific mechanisms in human airway epithelial cells. The approaches used in the present study demonstrate the utility of a promoter-reporter assay ensemble for identifying transcriptional pathways activated by pollutant exposure.

  19. Mutant quantity and quality in mammalian cells (A{sub L}) exposed to cesium-137 gamma radiation: Effect of caffeine

    Energy Technology Data Exchange (ETDEWEB)

    McGuinness, S.M.; Shibuya, M.L.; Ueno, A.M. [Colorado State Univ., Fort Collins, CO (United States)] [and others

    1995-06-01

    We examined the effect of caffeine (1,3,7-trimethylxanthine) on the quantity and quality of mutations in cultured mammalian A{sub L} human-hamster hybrid cells exposed to {sup 137}Cs {gamma} radiation. At a dose (1.5 mg/ml for 16 h) that reduced the plating efficiency (PE) by 20%, caffeine was not itself a significant mutagen, but it increased by approximately twofold the slope of the dose-response curve for induction of S1{sup {minus}} mutants by {sup 137}Cs {gamma} radiation. Molecular analysis of 235 S1{sup {minus}} mutants using a series of DNA probes mapped to the human chromosome 11 in the A{sub L} hybrid cells revealed that 73 to 85% of the mutations in unexposed cells and in cells treated with caffeine alone, {sup 137}Cs {gamma} rays alone or {sup 137}Cs {gamma} rays plus caffeine were large deletions involving millions of base pairs of DNA. Most of these deletions were contiguous with the region of the MIC1 gene at 11p13 that encodes the S1 cell surface antigen. In other mutants that had suffered multiple marker loss, the deletions were intermittent along chromosome 11. These {open_quotes}complex{close_quotes} mutations were rare for {sup 137}Cs {gamma} irradiation (1/63 = 1.5%) but relatively prevalent (23-50%) for other exposure conditions. Thus caffeine appears to alter both the quantity and quality of mutations induced by {sup 137}Cs {gamma} irradiation. 62 refs., 3 figs., 3 tabs.

  20. CD28 in thymocyte development and peripheral T cell activation in mice exposed to suspended particulate matter

    International Nuclear Information System (INIS)

    Drela, Nadzieja; Zesko, Izabela; Jakubowska, Martyna; Biernacka, Marzena

    2006-01-01

    The CD28:B7 signaling pathway is very important for the activity of mature peripheral T lymphocytes and thymocyte development. The proper development of thymocytes into mature single positive CD4 + and CD8 + T cells is crucial for almost all immune functions. In naturally occurring conditions, T cells maturation in the thymus is influenced by environmental agents. The expression of CD28 and the distribution of CD28 low/high thymocytes have been examined at various stages of thymocyte development in BALB/c mice exposed to air-suspended particulate matter (ASM). Acute exposure to ASM resulted in the decrease of CD28 expression in the total thymocyte population. The increase of the percentage of CD28 low and the decrease of CD28 high thymocytes were observed, which may account for the acceleration of thymocyte development under the conditions of elevated risk resulting from the exposure of animals to environmental xenobiotics. ASM exposure resulted in the increase of the level of proliferation of lymph node T cells induced by anti-CD3 and anti-CD28 monoclonal antibodies activation despite normal expression of CD28 molecule. In contrast, the level of proliferation of spleen T cells was lowered or normal dependently of the concentration of stimuli used for activation. Results of these studies demonstrate that acute exposure of mice to ASM can result in the progression of two contrasting processes in the immune system: upregulation of thymocyte development, which contributes to the maintenance of peripheral T cell pool, and over-activation of lymph node lymphocytes, which may lead to uncontrolled immunostimulation

  1. Sodium nitrite induces acute central nervous system toxicity in guinea pigs exposed to systemic cell-free hemoglobin

    International Nuclear Information System (INIS)

    Buehler, Paul W.; Butt, Omer I.; D'Agnillo, Felice

    2011-01-01

    Highlights: → Toxicological implications associated with the use of NaNO 2 therapy to treat systemic cell-free Hb exposure are not well-defined. → Systemic Hb exposure followed by NaNO 2 infusion induces acute CNS toxicities in guinea pigs. → These CNS effects were not reproduced by the infusion of cell-free Hb or NaNO 2 alone. → NaNO 2 -mediated oxidation of cell-free Hb may play a causative role in the observed CNS changes. -- Abstract: Systemic cell-free hemoglobin (Hb) released via hemolysis disrupts vascular homeostasis, in part, through the scavenging of nitric oxide (NO). Sodium nitrite (NaNO 2 ) therapy can attenuate the hypertensive effects of Hb. However, the chemical reactivity of NaNO 2 with Hb may enhance heme- or iron-mediated toxicities. Here, we investigate the effect of NaNO 2 on the central nervous system (CNS) in guinea pigs exposed to systemic cell-free Hb. Intravascular infusion of NaNO 2 , at doses sufficient to alleviate Hb-mediated blood pressure changes, reduced the expression of occludin, but not zona occludens-1 (ZO-1) or claudin-5, in cerebral tight junctions 4 h after Hb infusion. This was accompanied by increased perivascular heme oxygenase-1 expression, neuronal iron deposition, increased astrocyte and microglial activation, and reduced expression of neuron-specific nuclear protein (NeuN). These CNS changes were not observed in animals treated with Hb or NaNO 2 alone. Taken together, these findings suggest that the use of nitrite salts to treat systemic Hb exposure may promote acute CNS toxicity.

  2. Exposure to Tobacco Advertising, Promotion Among the Adult Population in Vietnam and Its Implications for Public Health.

    Science.gov (United States)

    Huong, Le Thi Thanh; Long, Tran Khanh; Van Anh, Tran Thi; Tuyet-Hanh, Tran Thi; Giang, Kim Bao; Hai, Phan Thi; Huyen, Doan Thu; Khue, Luong Ngoc; Lam, Nguyen Tuan; Nga, Pham Quynh; Quan, Nguyen The; Linh, Tran Nu Quy; Ha, Nguyen Thanh; Van Minh, Hoang

    2017-10-01

    The Law on Tobacco Control and the Law on Advertisement prohibit the advertising of any tobacco product in Vietnam. Tobacco promotion and marketing are alsostrictly prohibited. However, the violation of tobacco adverting and promotion is still common in Vietnam. This article aims at describing the exposure to tobacco advertising and promotion among the population aged 15+ years in Vietnam based on the data from the Global Adult Tobacco Survey 2015 from the view of the community, identifying any possible associations between the exposure to tobacco advertising and promotion and other individual factors, and discussing its possible public health implications. A cross-sectional study with the nationwide scale. Secondary data from the Global Adult Tobacco Survey 2015 was explored and analyzed. Chi-square test and multivariate logistic regressions were applied in the data analysis. The most common type of adults' exposure to tobacco advertising and promotion was points of sale (8.6%); 9.8% adults exposure to one source of tobacco advertising and 4.0% of them exposed to one source of tobacco promotion. Around 13.3% of Vietnamese adults were exposed to tobacco advertisement, while 2.0% were exposed to tobacco promotion, 5.3% were exposed to both tobacco advertising and promotion, and 16.6% were exposed to tobacco advertising or promotion. Gender, educational level, age, occupation, marital status, socioeconomic status, location (urban, rural), and current smoking status were associated with the exposure to tobacco advertising, tobacco promotion, tobacco advertising and promotion, and tobacco advertising or promotion. Although there are comprehensive bans on tobacco advertising and promotion in Vietnam, adults aged 15+ years still reported their exposure to tobacco advertising and promotion. There should be a strict enforcement of the ban on tobacco advertising and promotion in Vietnam.

  3. Global gene expression changes in human urothelial cells exposed to low-level monomethylarsonous acid

    International Nuclear Information System (INIS)

    Medeiros, Matthew; Zheng, Xinghui; Novak, Petr; Wnek, Shawn M.; Chyan, Vivian; Escudero-Lourdes, Claudia; Gandolfi, A. Jay

    2012-01-01

    Highlights: ► Chronic exposure to 50 nM monomethylarsonous acid in UROtsa was investigated. ► At 3 months of exposure substantial changes were observed in gene expression. ► Notable changes occurred in mitogenic signaling, stress, immune and inflammatory responses. ► Gene expression changes correlate with phenotypic changes from previous studies. -- Abstract: Bladder cancer has been associated with chronic arsenic exposure. Monomethylarsonous acid [MMA(III)] is a metabolite of inorganic arsenic and has been shown to transform an immortalized urothelial cell line (UROtsa) at concentrations 20-fold less than arsenite. MMA(III) was used as a model arsenical to examine the mechanisms of arsenical-induced transformation of urothelium. A microarray analysis was performed to assess the transcriptional changes in UROtsa during the critical window of chronic 50 nM MMA(III) exposure that leads to transformation at 3 months of exposure. The analysis revealed only minor changes in gene expression at 1 and 2 months of exposure, contrasting with substantial changes observed at 3 months of exposure. The gene expression changes at 3 months were analyzed showing distinct alterations in biological processes and pathways such as a response to oxidative stress, enhanced cell proliferation, anti-apoptosis, MAPK signaling, as well as inflammation. Twelve genes selected as markers of these particular biological processes were used to validate the microarray and these genes showed a time-dependent changes at 1 and 2 months of exposure, with the most substantial changes occurring at 3 months of exposure. These results indicate that there is a strong association between the acquired phenotypic changes that occur with chronic MMA(III) exposure and the observed gene expression patterns that are indicative of a malignant transformation. Although the substantial changes that occur at 3 months of exposure may be a consequence of transformation, there are common occurrences of altered

  4. DNA damage in human germ cell exposed to the some food additives in vitro.

    Science.gov (United States)

    Pandir, Dilek

    2016-08-01

    The use of food additives has increased enormously in modern food technology but they have adverse effects in human healthy. The aim of this study was to investigate the DNA damage of some food additives such as citric acid (CA), benzoic acid (BA), brilliant blue (BB) and sunset yellow (SY) which were investigated in human male germ cells using comet assay. The sperm cells were incubated with different concentrations of these food additives (50, 100, 200 and 500 μg/mL) for 1 h at 32 °C. The results showed for CA, BA, BB and SY a dose dependent increase in tail DNA%, tail length and tail moment in human sperm when compared to control group. When control values were compared in the studied parameters in the treatment concentrations, SY was found to exhibit the highest level of DNA damage followed by BB > BA > CA. However, none of the food additives affected the tail DNA%, tail length and tail moment at 50 and 100 μg/mL. At 200 μg/mL of SY, the tail DNA% and tail length of sperm were 95.80 ± 0.28 and 42.56 ± 4.66, for BB the values were 95.06 ± 2.30 and 39.56 ± 3.78, whereas for BA the values were 89.05 ± 2.78 and 31.50 ± 0.71, for CA the values were 88.59 ± 6.45 and 13.59 ± 2.74, respectively. However, only the highest concentration of the used food additives significantly affected the studied parameters of sperm DNA. The present results indicate that SY and BB are more harmful than BA and CA to human sperm in vitro.

  5. Reversal of Human Papillomavirus-Specific T Cell Immune Suppression through TLR Agonist Treatment of Langerhans Cells Exposed to Human Papillomavirus Type 161

    Science.gov (United States)

    Fahey, Laura M.; Raff, Adam B.; Da Silva, Diane M.; Kast, W. Martin

    2009-01-01

    Human papillomavirus (HPV) type 16 infects the epithelial layer of cervical mucosa and is causally associated with the generation of cervical cancer. Langerhans cells (LC) are the resident antigen-presenting cells at the site of infection and therefore are responsible for initiating an immune response against HPV16. On the contrary, LC exposed to HPV16 do not induce a specific T cell immune response, which leads to the immune evasion of HPV16. Demonstrating that Toll-like receptor 7 (TLR7) and TLR8 are expressed on human LC, we hypothesized that imidazoquinolines would activate LC exposed to HPV16, leading to the induction of an HPV16-specific cell-mediated immune response. Surprisingly both phenotypic and functional hallmarks of activation are not observed when LC are exposed to HPV16 virus-like particles (VLP) and treated with imiquimod (TLR7 agonist). However, we found that LC are activated by 3M-002 (TLR8 agonist) and resiquimod (TLR8/7 agonist). LC exposed to HPV16 VLP and subsequently treated with 3M-002 or resiquimod highly up-regulate surface activation markers, secrete pro-inflammatory cytokines and chemokines, induce CCL21-directed migration, and initiate an HPV16-specific CD8+ T cell response. These data strongly indicate that 3M-002 and resiquimod are promising therapeutics for treatment of HPV-infections and HPV-induced cervical lesions. This is an author-produced version of a manuscript accepted for publication in The Journal of Immunology (The JI). The American Association of Immunologists, Inc. (AAI), publisher of The JI, holds the copyright to this manuscript. This version of the manuscript has not yet been copyedited or subjected to editorial proofreading by The JI; hence, it may differ from the final version published in The JI (online and in print). AAI (The JI) is not liable for errors or omissions in this author-produced version of the manuscript or in any version derived from it by the U.S. National Institutes of Health or any other third

  6. Live-Cell Imaging Visualizes Frequent Mitotic Skipping During Senescence-Like Growth Arrest in Mammary Carcinoma Cells Exposed to Ionizing Radiation

    International Nuclear Information System (INIS)

    Suzuki, Masatoshi; Yamauchi, Motohiro; Oka, Yasuyoshi; Suzuki, Keiji; Yamashita, Shunichi

    2012-01-01

    Purpose: Senescence-like growth arrest in human solid carcinomas is now recognized as the major outcome of radiotherapy. This study was designed to analyze cell cycle during the process of senescence-like growth arrest in mammary carcinoma cells exposed to X-rays. Methods and Materials: Fluorescent ubiquitination-based cell cycle indicators were introduced into the human mammary carcinoma cell line MCF-7. Cell cycle was sequentially monitored by live-cell imaging for up to 5 days after exposure to 10 Gy of X-rays. Results: Live-cell imaging revealed that cell cycle transition from G2 to G1 phase without mitosis, so-called mitotic skipping, was observed in 17.1% and 69.8% of G1- and G2-irradiated cells, respectively. Entry to G1 phase was confirmed by the nuclear accumulation of mKO 2 -hCdt1 as well as cyclin E, which was inversely correlated to the accumulation of G2-specific markers such as mAG-hGeminin and CENP-F. More than 90% of cells skipping mitosis were persistently arrested in G1 phase and showed positive staining for the senescent biochemical marker, which is senescence-associated ß-galactosidase, indicating induction of senescence-like growth arrest accompanied by mitotic skipping. While G2 irradiation with higher doses of X-rays induced mitotic skipping in approximately 80% of cells, transduction of short hairpin RNA (shRNA) for p53 significantly suppressed mitotic skipping, suggesting that ionizing radiation-induced mitotic skipping is associated with p53 function. Conclusions: The present study found the pathway of senescence-like growth arrest in G1 phase without mitotic entry following G2-irradiation.

  7. Live-Cell Imaging Visualizes Frequent Mitotic Skipping During Senescence-Like Growth Arrest in Mammary Carcinoma Cells Exposed to Ionizing Radiation

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Masatoshi, E-mail: msuzuki@nagasaki-u.ac.jp [Department of Radiation Medical Sciences, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki (Japan); Yamauchi, Motohiro; Oka, Yasuyoshi; Suzuki, Keiji; Yamashita, Shunichi [Department of Radiation Medical Sciences, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki (Japan)

    2012-06-01

    Purpose: Senescence-like growth arrest in human solid carcinomas is now recognized as the major outcome of radiotherapy. This study was designed to analyze cell cycle during the process of senescence-like growth arrest in mammary carcinoma cells exposed to X-rays. Methods and Materials: Fluorescent ubiquitination-based cell cycle indicators were introduced into the human mammary carcinoma cell line MCF-7. Cell cycle was sequentially monitored by live-cell imaging for up to 5 days after exposure to 10 Gy of X-rays. Results: Live-cell imaging revealed that cell cycle transition from G2 to G1 phase without mitosis, so-called mitotic skipping, was observed in 17.1% and 69.8% of G1- and G2-irradiated cells, respectively. Entry to G1 phase was confirmed by the nuclear accumulation of mKO{sub 2}-hCdt1 as well as cyclin E, which was inversely correlated to the accumulation of G2-specific markers such as mAG-hGeminin and CENP-F. More than 90% of cells skipping mitosis were persistently arrested in G1 phase and showed positive staining for the senescent biochemical marker, which is senescence-associated ss-galactosidase, indicating induction of senescence-like growth arrest accompanied by mitotic skipping. While G2 irradiation with higher doses of X-rays induced mitotic skipping in approximately 80% of cells, transduction of short hairpin RNA (shRNA) for p53 significantly suppressed mitotic skipping, suggesting that ionizing radiation-induced mitotic skipping is associated with p53 function. Conclusions: The present study found the pathway of senescence-like growth arrest in G1 phase without mitotic entry following G2-irradiation.

  8. Effect of doxorubicin on cell survival and micronuclei formation in HeLa cells exposed to different doses of gamma-radiation

    International Nuclear Information System (INIS)

    Jagetia, G.C.; Nayak, V.

    2000-01-01

    Purpose: The present study was undertaken to obtain an insight into the combined effects of doxorubicin with radiation on the cell survival and micronuclei induction in HeLa cells. Material and Methods: HeLa S3 cells were allowed to grow till they reached plateau phase, inoculated with 10 μg/ml doxorubicin hydrochloride and then exposed to 0, 0.5, 1, 2 and 3 Gy γ-radiation. Clonogenicity of cell was measured using the colony forming assay, micronuclei formation using the micronucleus assay. Results: The treatment of HeLa cells with doxorubicin (adriamycin) for 2 hours before exposure to different doses of γ-radiation resulted in a significant and dose-dependent decline in the cell survival and cell proliferation when compared to the PBS+irradiation group. Conversely, the frequency of micronuclei increased in a dose-related manner in both the PBS+irradiation and doxorubicin+irradiation groups. The pretreatment of HeLa cells with doxorubicin before irradiation to various doses of γ-rays resulted in a significant elevation in the frequency of micronuclei when compared with the concurrent PBS+irradiation group. The dose-response relationship for both PBS+irradiation and doxorubicin+irradiation groups was linear. The correlation between cell survival and micronuclei induction was also determined for PBS or doxorubicin+irradiation group, where the clonogenicity of cells declined with the increase in micronuclei formation. The correlation between cell survical and micronuclei induction was linear quadratic for both PBS+irradiation and doxorubicin+irradiation groups. Conclusion: From our study it can be concluded that combination treatment with doxorubicin and radiation increased the genotoxic effect of the either treatment given alone. (orig.) [de

  9. Mode selection in electrical activities of myocardial cell exposed to electromagnetic radiation

    International Nuclear Information System (INIS)

    Ma, Jun; Wang, Ya; Wang, Chunni; Xu, Ying; Ren, Guodong

    2017-01-01

    Highlights: • Neuronal model under electromagnetic induction and radiation is set up; • The transition of electrical activities under electromagnetic radiation is discussed; • Dynamical response of encoding of neuron is discussed for possible mechanism of heart disease. - Abstract: Based on the Fitzhugh–Nagumo neuron model, the effect of electromagnetic induction is considered and external electromagnetic radiation is imposed to detect the mode transition of electrical activities in a myocardial cell. Appropriate dynamical and functional responses can be observed in the sampled series for membrane potentials by setting different feedback modulation on the membrane potential in presence of electromagnetic radiation. The electromagnetic radiation is described by a periodical forcing on the magnetic flux, and it is found that the response frequency can keep pace with the frequency of external forcing. However, mismatch of frequency occurs by further increasing the frequency of external forcing, it could account for the information encoding of neuron. The dynamical response could be associated with the magnetization and polarization of the media, thus the outputs of membrane potential can become quiescent and/or bursting as well.

  10. In vitro safety evaluation of human nasal epithelial cell monolayers exposed to carrageenan sinus wash.

    Science.gov (United States)

    Ramezanpour, Mahnaz; Murphy, Jae; Smith, Jason L P; Vreugde, Sarah; Psaltis, Alkis James

    2017-12-01

    Carrageenans have shown to reduce the viral load in nasal secretions and lower the incidence of secondary infections in children with common cold. Despite the widespread use of carrageenans in topical applications, the effect of carrageenans on the sinonasal epithelial barrier has not been elucidated. We investigate the effect of different carrageenans on the sinonasal epithelial barrier and inflammatory response in vitro. Iota and Kappa carrageenan delivered in saline irrigation solutions applied to air-liquid interface (ALI) cultures of primary human nasal epithelial cells from chronic rhinosinusitis patients and controls. Epithelial barrier structure was assessed by measuring the transepithelial electrical resistance (TEER) and immunolocalization of F actin. Ciliary beat frequency (CBF), toxicity, and inflammatory response was studied. Kappa or Iota carrageenan in the different solutions was not toxic, did not have detrimental effects on epithelial barrier structure and CBF. Rather, application of Kappa carrageenan significantly increased TEER and suppressed interleukin 6 (IL-6) secretion in ALI cultures from CRS patients. Kappa or Iota carrageenan solution was safe and did not negatively affect epithelial barrier function. Kappa carrageenan increased TEER and decreased IL-6 production in CRS patients, indicating positive effects on epithelial barrier function in vitro. © 2017 ARS-AAOA, LLC.

  11. The structure of cortical cytoplasm in cold-treated tobacco cells: the role of the cytoskeleton and the endomembrane system

    Czech Academy of Sciences Publication Activity Database

    Schwarzerová, K.; Pokorná, J.; Petrášek, Jan; Zelenková, S.; Čapková, Věra; Janotová, I.; Opatrný, Z.

    2003-01-01

    Roč. 27, - (2003), 263ů265 ISSN 1065-6995 R&D Projects: GA AV ČR IAA5038207 Institutional research plan: CEZ:AV0Z5038910 Keywords : cytoskeletal polymers * tobacco * endomembrane system Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.092, year: 2003

  12. The structure of cortical cytoplasm in cold-treated tobacco cells: the role of the cytoskeleton and the endomembrane system

    Czech Academy of Sciences Publication Activity Database

    Schwarzerová, K.; Pokorná, J.; Petrášek, Jan; Zelenková, S.; Čapková, Věra; Janotová, I.; Opatrný, Z.

    2003-01-01

    Roč. 27, - (2003), s. 263ů265 ISSN 1065-6995 R&D Projects: GA AV ČR IAA5038207 Institutional research plan: CEZ:AV0Z5038910; CEZ:MSM 113100003 Keywords : cytoskeletal polymers * tobacco * endomembrane system Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.092, year: 2003

  13. Evaluation of buffers toxicity in tobacco cells: Homopiperazine-1,4-bis (2-ethanesulfonic acid) is a suitable buffer for plant cells studies at low pH.

    Science.gov (United States)

    Borgo, Lucélia

    2017-06-01

    Low pH is an important environmental stressor of plant root cells. Understanding the mechanisms of stress and tolerance to acidity is critical; however, there is no widely accepted pH buffer for studies of plant cells at low pH. Such a buffer might also benefit studies of Al toxicity, in which buffering at low pH is also important. The challenge is to find a buffer with minimal cellular effects. We examined the cytotoxicity and possible metabolic disturbances of four buffers that have adequate pK a values and potential use for studies in the pH range of 4.0-5.0. These were homopipes (homopiperazine-1,4-bis (2-ethanesulfonic acid); pK a1 4.4), 3,3-dimethylglutaric acid (pK a1 3.73), β-alanine (pK a1 3.70) and potassium biphthalate (pK a1 2.95; pK a2 5.41). First, tobacco BY-2 cells were grown in a rich medium containing 10 mM of each buffer or MES (2-(N-morpholino) ethanesulfonic acid) as a control, with the pH initially adjusted to 5.7. β-alanine was clearly toxic and dimethylgluturate and biphthalate were found to be cytostatic, in which no culture growth occurred but cell viability was either unaffected or decreased only after 5 days. Only homopipes allowed normal culture growth and cell viability. Homopipes (10 mM) was then tested in cell cultures with an initial pH of 4.3 ± 0.17 in minimal medium to examine whether its undissociated species (H 2 A) displayed any cellular effects and no cytotoxic effects were observed. It is possible to conclude that among tested buffers, homopipes is the most suitable for studies at low pH, and may be especially useful for aluminum toxicity experiments. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  14. Quantitative proteomics and transcriptomics addressing the estrogen receptor subtype-mediated effects in T47D breast cancer cells exposed to the phytoestrogen genistein

    NARCIS (Netherlands)

    Sotoca Covaleda, A.M.; Sollewijn Gelpke, M.D.; Boeren, S.; Ström, A.; Gustafsson, J.A.; Murk, A.J.; Rietjens, I.M.C.M.; Vervoort, J.J.M.

    2011-01-01

    The present study addresses, by transcriptomics and quantitative SILAC-based proteomics, the estrogen receptor alpha (ER) and beta (ERß)-mediated effects on gene and protein expression in T47D breast cancer cells exposed to the phytoestrogen genistein. Using the T47D human breast cancer cell line

  15. Repair and cell cycle response in cells exposed to environmental biohazards. Progress report, 1 June 1982-31 May 1983

    International Nuclear Information System (INIS)

    Billen, D.; Hadden, C.T.

    1983-01-01

    The research program has attempted to determine low environmental agents, especially ionizing radiation, interact with DNA and how the cells respond to the resulting damage. The focus has been on the spectrum of damage generated in DNA, the kinds of damage that can be repaired, and the mechanisms of repair. The effects of radioprotective agents and certain sensitizing conditions on breakage of DNA in bacterial cells by ionizing radiation, and mechanisms of enzymatic excision of pyrimidine dimers and other DNA adducts have been studied

  16. Exposing primary rat retina cell cultures to γ-rays: An in vitro model for evaluating radiation responses.

    Science.gov (United States)

    Gaddini, Lucia; Balduzzi, Maria; Campa, Alessandro; Esposito, Giuseppe; Malchiodi-Albedi, Fiorella; Patrono, Clarice; Matteucci, Andrea

    2018-01-01

    Retinal tissue can receive incidental γ-rays exposure during radiotherapy either of tumors of the eye and optic nerve or of head-and-neck tumors, and during medical diagnostic procedures. Healthy retina is therefore at risk of suffering radiation-related side effects and the knowledge of pathophysiological response of retinal cells to ionizing radiations could be useful to design possible strategies of prevention and management of radiotoxicity. In this study, we have exploited an in vitro model (primary rat retinal cell culture) to study an array of biological effects induced on retinal neurons by γ-rays. Most of the different cell types present in retinal tissue - either of the neuronal or glial lineages - are preserved in primary rat retinal cultures. Similar to the retina in situ, neuronal cells undergo in vitro a maturational development shown by the formation of polarized neuritic trees and operating synapses. Since 2 Gy is the incidental dose received by the healthy retina per fraction when the standard treatment is delivered to the brain, retina cell cultures have been exposed to 1 or 2 Gy of γ-rays at different level of neuronal differentiation in vitro: days in vitro (DIV)2 or DIV8. At DIV9, retinal cultures were analyzed in terms of viability, apoptosis and characterized by immunocytochemistry to identify alterations in neuronal differentiation. After irradiation at DIV2, MTT assay revealed an evident loss of cell viability and βIII-tubulin immunostaining highlighted a marked neuritic damage, indicating that survived neurons showed an impaired differentiation. Differentiated cultures (DIV8) appeared to be more resistant with respect to undifferentiated, DIV2 cultures, both in terms of cell viability and differentiation. Apoptosis evaluated with TUNEL assay showed that irradiation at both DIV2 and DIV8 induced a significant increase in the apoptotic rate. To further investigate the effects of γ-rays on retinal neurons, we evaluated the

  17. Different Blood Cell-Derived Transcriptome Signatures in Cows Exposed to Vaccination Pre- or Postpartum

    Science.gov (United States)

    Weikard, Rosemarie; Demasius, Wiebke; Hadlich, Frieder; Kühn, Christa

    2015-01-01

    Periparturient cows have been found to reveal immunosuppression, frequently associated with increased susceptibility to uterine and mammary infections. To improve understanding of the causes and molecular regulatory mechanisms accounting for this phenomenon around calving, we examined the effect of an antigen challenge on gene expression modulation on cows prior to (BC) or after calving (AC) using whole transcriptome sequencing (RNAseq). The transcriptome analysis of the cows’ blood identified a substantially higher number of loci affected in BC cows (2,235) in response to vaccination compared to AC cows (208) and revealed a divergent transcriptional profile specific for each group. In BC cows, a variety of loci involved in immune defense and cellular signaling processes were transcriptionally activated, whereas protein biosynthesis and posttranslational processes were tremendously impaired in response to vaccination. Furthermore, energy metabolism in the blood cells of BC cows was shifted from oxidative phosphorylation to the glycolytic system. In AC cows, the number and variety of regulated pathways involved in immunomodulation and maintenance of immnunocompetence are considerably lower after vaccination, and upregulation of arginine degradation was suggested as an immunosuppressive mechanism. Elevated transcript levels of erythrocyte-specific genes involved in gas exchange processes were a specific transcriptional signature in AC cows pointing to hematopoiesis activation. The divergent and substantially lower magnitude of transcriptional modulation in response to vaccination in AC cows provides evidence for a suppressed immune capacity of early lactating cows on the molecular level and demonstrates that an efficient immune response of cows is related to their physiological and metabolic status. PMID:26317664

  18. Monitoring protein turnover during phosphate starvation-dependent autophagic degradation using a photoconvertible fluorescent protein aggregate in tobacco BY-2 cells.

    Science.gov (United States)

    Tasaki, Maiko; Asatsuma, Satoru; Matsuoka, Ken

    2014-01-01

    We have developed a system for quantitative monitoring of autophagic degradation in transformed tobacco BY-2 cells using an aggregate-prone protein comprised of cytochrome b5 (Cyt b5) and a tetrameric red fluorescent protein (RFP). Unfortunately, this system is of limited use for monitoring the kinetics of autophagic degradation because the proteins synthesized before and after induction of autophagy cannot be distinguished. To overcome this problem, we developed a system using kikume green-red (KikGR), a photoconvertible and tetrameric fluorescent protein that changes its fluorescence from green to red upon irradiation with purple light. Using the fusion protein of Cyt b5 and KikGR together with a method for the bulk conversion of KikGR, which we had previously used to convert the Golgi-localized monomeric KikGR fusion protein, we were able to monitor both the growth and de novo formation of aggregates. Using this system, we found that tobacco cells do not cease protein synthesis under conditions of phosphate (Pi)-starvation. Induction of autophagy under Pi-starvation, but not under sugar- or nitrogen-starvation, was specifically inhibited by phosphite, which is an analog of Pi with a different oxidation number. Therefore, the mechanism by which BY-2 cells can sense Pi-starvation and induce autophagy does not involve sensing a general decrease in energy supply and a specific Pi sensor might be involved in the induction of autophagy under Pi-starvation.

  19. Production by Tobacco Transplastomic Plants of Recombinant Fungal and Bacterial Cell-Wall Degrading Enzymes to Be Used for Cellulosic Biomass Saccharification

    Directory of Open Access Journals (Sweden)

    Paolo Longoni

    2015-01-01

    Full Text Available Biofuels from renewable plant biomass are gaining momentum due to climate change related to atmospheric CO2 increase. However, the production cost of enzymes required for cellulosic biomass saccharification is a major limiting step in this process. Low-cost production of large amounts of recombinant enzymes by transgenic plants was proposed as an alternative to the conventional microbial based fermentation. A number of studies have shown that chloroplast-based gene expression offers several advantages over nuclear transformation due to efficient transcription and translation systems and high copy number of the transgene. In this study, we expressed in tobacco chloroplasts microbial genes encoding five cellulases and a polygalacturonase. Leaf extracts containing the recombinant enzymes showed the ability to degrade various cell-wall components under different conditions, singly and in combinations. In addition, our group also tested a previously described thermostable xylanase in combination with a cellulase and a polygalacturonase to study the cumulative effect on the depolymerization of a complex plant substrate. Our results demonstrate the feasibility of using transplastomic tobacco leaf extracts to convert cell-wall polysaccharides into reducing sugars, fulfilling a major prerequisite of large scale availability of a variety of cell-wall degrading enzymes for biofuel industry.

  20. Production by Tobacco Transplastomic Plants of Recombinant Fungal and Bacterial Cell-Wall Degrading Enzymes to Be Used for Cellulosic Biomass Saccharification.

    Science.gov (United States)

    Longoni, Paolo; Leelavathi, Sadhu; Doria, Enrico; Reddy, Vanga Siva; Cella, Rino

    2015-01-01

    Biofuels from renewable plant biomass are gaining momentum due to climate change related to atmospheric CO2 increase. However, the production cost of enzymes required for cellulosic biomass saccharification is a major limiting step in this process. Low-cost production of large amounts of recombinant enzymes by transgenic plants was proposed as an alternative to the conventional microbial based fermentation. A number of studies have shown that chloroplast-based gene expression offers several advantages over nuclear transformation due to efficient transcription and translation systems and high copy number of the transgene. In this study, we expressed in tobacco chloroplasts microbial genes encoding five cellulases and a polygalacturonase. Leaf extracts containing the recombinant enzymes showed the ability to degrade various cell-wall components under different conditions, singly and in combinations. In addition, our group also tested a previously described thermostable xylanase in combination with a cellulase and a polygalacturonase to study the cumulative effect on the depolymerization of a complex plant substrate. Our results demonstrate the feasibility of using transplastomic tobacco leaf extracts to convert cell-wall polysaccharides into reducing sugars, fulfilling a major prerequisite of large scale availability of a variety of cell-wall degrading enzymes for biofuel industry.

  1. Repair and cell cycle response in cells exposed to environmental biohazards. Progress report, October 1, 1976--May 31, 1979

    International Nuclear Information System (INIS)

    Billen, D.

    1979-01-01

    A wide variety of environmental agents are known which induce damage in DNA leading to an inhibition of DNA synthesis or faulty replication. Both results may cause cell death or mutation. Both bacteria and mommalian cells are being used to assess the roles of the several known DNA polymerases and other DNA metabolic enzymes and factors in DNA repair, replication and recombination. The many DNA mutants of E. coli and B. subtilis provide a genetic approach to measuring the role of individual components of the DNA repair and replicative system. Because of the advantage of controlling pools and precursors of nucleic acid synthesis we have further developed the use of permeabilized cells for such studies. In addition a series of repair studies with Bacillus subtilis have been carried on because of the unique genetic manipulation of this system which includes the ability of cells to be easily transformed by exogenous DNA. The information obtained with prokaryotes provides leads to assess the details of DNA repair and replication in mammalian systems including man

  2. Cytogenetic effects in bone marrow cells of mice exposed on the biosatellite "BION-M1"

    Science.gov (United States)

    Dorozhkina, Olga; Ivanov, Alexander

    In studies of cytogenetic damage in blood lymphocytes of astronauts, conducted in recent years, have shown an increase in the frequency of chromosomal damage bound, as believe, with influence on an organism of astronauts of space radiation (B.S. Fedorenko, G.P. Snigireva, 2004). However, in recent years published evidence that both acute and chronic stress induce chromosomal aberrations and modified genome sensitivity to mutagens of different nature, including to ionizing radiation (F.I. Ingel et al, 2005 ). This question is especially actual for space biology and medicine due to a number of specific features of space flights, when the interaction of factors more pronounced than in normal terrestrial conditions. In experiment "BION - M1" by anaphase method was determined level of chromosomal aberrations in bone marrow cells of tibia of mice. Flight duration biosatellite "BION - M1" was 30 days in Earth orbit. Euthanasia of experimental animals was carried out at intervals of 15-20 minutes by method of cervical dislocation after 12 hours from the moment of landing satellite. Level of chromosomal aberrations in vivarium-housed control mice was 1,75 ± 0,6% and 1,8 ± 0,45%, while the mitotic index 1,46 ± 0,09% and 1,53 ± 0,05%. Differences are not significant. The maintenance of animals in experiment with the onboard equipment (ground experiment) led to some increase in aberrant mitoses (2,3 ± 0,4%) and to decrease in a mitotic index (1,37 ± 0,02%). In the flight experiment "BION - M1" statistically significant increase of level of chromosomal aberrations (29,7 ± 4,18%) and a decrease in the mitotic index (0,74 ± 0,07%). Since the mouse is a suitable experimental model , also had several ground experiments on research of combined effect of irradiation and other stress factors specific to space flight, with marked tendency to increase the level of aberrant mitoses under the combined action of radiation and stress exposure group housing male mice. Statistically

  3. PI-103 and Quercetin Attenuate PI3K-AKT Signaling Pathway in T- Cell Lymphoma Exposed to Hydrogen Peroxide.

    Directory of Open Access Journals (Sweden)

    Akhilendra Kumar Maurya

    Full Text Available Phosphatidylinositol 3 kinase-protein kinase B (PI3K-AKT pathway has been considered as major drug target site due to its frequent activation in cancer. AKT regulates the activity of various targets to promote tumorigenesis and metastasis. Accumulation of reactive oxygen species (ROS has been linked to oxidative stress and regulation of signaling pathways for metabolic adaptation of tumor microenvironment. Hydrogen peroxide (H2O2 in this context is used as ROS source for oxidative stress preconditioning. Antioxidants are commonly considered to be beneficial to reduce detrimental effects of ROS and are recommended as dietary supplements. Quercetin, a ubiquitous bioactive flavonoid is a dietary component which has attracted much of interest due to its potential health-promoting effects. Present study is aimed to analyze PI3K-AKT signaling pathway in H2O2 exposed Dalton's lymphoma ascite (DLA cells. Further, regulation of PI3K-AKT pathway by quercetin as well as PI-103, an inhibitor of PI3K was analyzed. Exposure of H2O2 (1mM H2O2 for 30min to DLA cells caused ROS accumulation and resulted in increased phosphorylation of PI3K and downstream proteins PDK1 and AKT (Ser-473 and Thr-308, cell survival factors BAD and ERK1/2, as well as TNFR1. However, level of tumor suppressor PTEN was declined. Both PI-103 & quercetin suppressed the enhanced level of ROS and significantly down-regulated phosphorylation of AKT, PDK1, BAD and level of TNFR1 as well as increased the level of PTEN in H2O2 induced lymphoma cells. The overall result suggests that quercetin and PI3K inhibitor PI-103 attenuate PI3K-AKT pathway in a similar mechanism.

  4. Quantitative proteomic analysis of cabernet sauvignon grape cells exposed to thermal stresses reveals alterations in sugar and phenylpropanoid metabolism.

    Science.gov (United States)

    George, Iniga S; Pascovici, Dana; Mirzaei, Mehdi; Haynes, Paul A

    2015-09-01

    Grapes (Vitis vinifera) are a valuable fruit crop and wine production is a major industry. Global warming and expanded range of cultivation will expose grapes to more temperature stresses in future. Our study investigated protein level responses to abiotic stresses, with particular reference to proteomic changes induced by the impact of four different temperature stress regimes, including both hot and cold temperatures, on cultured grape cells. Cabernet Sauvignon cell suspension cultures grown at 26°C were subjected to 14 h of exposure to 34 and 42°C for heat stress, and 18 and 10°C for cold stress. Cells from the five temperatures were harvested in biological triplicates and label-free quantitative shotgun proteomic analysis was performed. A total of 2042 non-redundant proteins were identified from the five temperature points. Fifty-five proteins were only detected in extreme heat stress conditions (42°C) and 53 proteins were only detected at extreme cold stress conditions (10°C). Gene Ontology (GO) annotations of differentially expressed proteins provided insights into the metabolic pathways that are involved in temperature stress in grape cells. Sugar metabolism displayed switching between alternative and classical pathways during temperature stresses. Additionally, nine proteins involved in the phenylpropanoid pathway were greatly increased in abundance at extreme cold stress, and were thus found to be cold-responsive proteins. All MS data have been deposited in the ProteomeXchange with identifier PXD000977 (http://proteomecentral.proteomexchange.org/dataset/PXD000977). © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Effect of Green Tea Extract on T cell Mediated Hypersensitivity Reaction in BALB/c Mice Exposed to Gamma Irradiation

    International Nuclear Information System (INIS)

    Hashim, A.M.; Ismail Al-kadey, M.M.I.; Shabon, M.H.; Hussien, S.M.

    2010-01-01

    Gamma radiation is widely used in the treatment of malignant neoplasms. However, it deprives the host immune function which may retard tumor rejection by the immune response. The main purpose of the present study is to test the ability of green tea dry extract to restore the T cell hypersensitivity reaction in gamma irradiated BALB/c mice. It aims also to elucidate the possible mechanism of action of ionizing radiation and green tea dry extract in the immune function. Four groups of BALB/c mice, each of ten, have been used in each experiment. The first group served as a control, the second group received green tea dry extract and the third group was exposed to 2 Gy gamma irradiation, while the fourth group received green tea dry extract before and after gamma irradiation. The following parameters were determined, the contact sensitivity reaction by the mouse ear swelling response, local dendritic cell migration, local lymph node weight, lymphocyte proliferation, spleen and thymus weight with their lymphocyte count. The effect of gamma irradiation and green tea dry extract on the elicitation phase of contact sensitivity was also determined. Data from the present study showed that gamma irradiation caused a significant decrease of the mouse ear swelling response and retarded dendritic cell migration. They also showed a significant decline in the lymphocytes proliferation in lymph node draining the contact sensitizer application. Total body exposure to 2 Gy gamma irradiation induced marked decline of thymus weight and thymocyte count, while it reduced spleen weight and spleenocyte count to a lesser extent. Exposure to gamma irradiation enhanced the elicitation phase of contact sensitivity. Administration of green tea dry extract partially preserved the contact sensitivity response to oxazolone in gamma irradiated BALB/c mice. It markedly minimized the enhancement of the elicitation phase of ear swelling. In conclusion, the present study heralds a beneficial role of

  6. Changes of inflammatory cells in rat lungs exposed to diesel emissions; Diesel haiki bakuro ni yoru rat hai no ensho saibo no henka

    Energy Technology Data Exchange (ETDEWEB)

    Kato, A. [Japan Automobile Research Institute Inc., Tsukuba (Japan); Kagawa, J. [Tokyo Women`s Medical College, Tokyo (Japan)

    1998-05-01

    Study was made on the effect of exposure to diesel emissions on inflammatory cells in a rat lungs. Four kinds of exposure gases with different contents of NO2 and particulate were prepared by diluting diesel emissions. Rats were exposed to diluted diesel emissions for 24 months, and inflammatory cells were detected morphologically in light microscopic and TEM specimens. As a result, particle-laden- alveolar macrophages increased dose- and time-dependently into the submucosa of intrapulmonary bronchioles, alveolar spaces and interstitume of alveolar walls, and bronchoassociated lymphatic tissues. Mast cells infiltrated into the interspaces of epithelial cells in airways. In the submucosa of the terminal bronchioles and the interstitume of alveolar walls, some sorts of inflammatory cells such as mast cells, plasma cells, neutrophils and lymphocytes infiltrated, and some cells showed cell-to-cell contacts. However, the airways were rarely injured by infiltration of inflammatory cells except for a fibrotic change. 2 refs., 2 figs., 2 tabs.

  7. Relationship among tobacco habits, human papilloma virus (HPV) infection, p53 polymorphism/mutation and the risk of oral squamous cell carcinoma.

    Science.gov (United States)

    Chakrobarty, Bidyut; Roy, Jay Gopal; Majumdar, Sumit; Uppala, Divya

    2014-05-01

    The prevalence of oral squamous cell carcinoma (OSCC) has significantly increased over decades in several countries and human papilloma virus (HPV) has been indicated as one of the underlying causes. This suggests that HPV plays a role in the early stages of carcinogenesis but is not a requisite for the maintenance and progression of malignant state. p53 is a tumor suppressor gene that checks the cell and promotes apoptosis and cell repair that can be deactivated by mutations and a viral interaction leading to cancer and individuals with particular polymorphic variant of p53 is more susceptible to HPV-induced carcinogenesis. The present study has been carried out to detect and correlate p53 polymorphism/mutation, HPV DNA in the biopsy samples of oral cancer patients who had tobacco habits.

  8. Autophagy is activated in compression-induced cell degeneration and is mediated by reactive oxygen species in nucleus pulposus cells exposed to compression.

    Science.gov (United States)

    Ma, K-G; Shao, Z-W; Yang, S-H; Wang, J; Wang, B-C; Xiong, L-M; Wu, Q; Chen, S-F

    2013-12-01

    To determine whether autophagy contributes to the pathogenesis of degenerative disc disease (DDD) or retards the intervertebral disc (IVD) degeneration, and investigate the possible relationship between compression-induced autophagy and intracellular reactive oxygen species (ROS) in nucleus pulposus (NP) cells in vitro. The autophagosome and autophagy-related markers were used to explore the role of autophagy in rat NP cells under compressive stress, which were measured directly by electronic microscopy, monodansylcadaverine (MDC) staining, immunofluorescence, western blot, and indirectly by analyzing the impact of pharmacological inhibitors of autophagy such as 3-methyladenine (3-MA) and chloroquine (CQ). And the relationship between autophagy and apoptosis was investigated by Annexin-V/propidium iodide (PI)-fluorescein staining. In addition, ROS were measured to determine whether these factors are responsible for the development of compression-induced autophagy. Our results indicated that rat NP cells activated autophagy in response to the same strong apoptotic stimuli that triggered apoptosis by compression. Autophagy and apoptosis were interconnected and coordinated in rat NP cells exposed to compression stimuli. Compression-induced autophagy was closely related to intracellular ROS production. Enhanced degradation of damaged components of NP cells by autophagy may be a crucial survival response against mechanical overload, and extensive autophagy may trigger autophagic cell death. Regulating autophagy and reducing the generation of intracellular ROS may retard IVD degeneration. Copyright © 2013 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

  9. Comparison of initial DNA (Chromosome) damage/repair in cells exposed to heavy ion particles and X-rays

    International Nuclear Information System (INIS)

    Okayasu, Ryuichi; Okada, Maki; Noguchi, Mitsuho; Saito, Shiori; Okabe, Atsushi; Takakura, Kahoru

    2005-01-01

    We have studied cell survival and chromosome damage/repair in normal and non homologous end-joining (NHEJ) deficient human cells exposed to carbon ions (290 MeV/u, ∼70 keV/um), iron ions (500 MeV/u, ∼200 keV/um) and X-rays. In order to examine the effect of heavy ion on double strand break (DSB) repair machinery, the auto-phosphorylation of DNA-PKcs was also investigated. The important discoveries made during this period are: 200 keV/um iron irradiation induced additional molecular damage beyond that 70 keV/um carbon did. Iron irradiation not only caused an inefficient G1 chromosome repair, but also induced non-repairable DSB/chromosome damage. The auto-phosphorylation of DNA-PKcs was significantly affected by high linear energy transfer (LET) irradiation when compared to X-rays. These results indicate NHEJ machinery was markedly disturbed by high LET radiation when compared to low LET radiation. (author)

  10. Structure-related clustering of gene expression fingerprints of thp-1 cells exposed to smaller polycyclic aromatic hydrocarbons.

    Science.gov (United States)

    Wan, B; Yarbrough, J W; Schultz, T W

    2008-01-01

    This study was undertaken to test the hypothesis that structurally similar PAHs induce similar gene expression profiles. THP-1 cells were exposed to a series of 12 selected PAHs at 50 microM for 24 hours and gene expressions profiles were analyzed using both unsupervised and supervised methods. Clustering analysis of gene expression profiles revealed that the 12 tested chemicals were grouped into five clusters. Within each cluster, the gene expression profiles are more similar to each other than to the ones outside the cluster. One-methylanthracene and 1-methylfluorene were found to have the most similar profiles; dibenzothiophene and dibenzofuran were found to share common profiles with fluorine. As expression pattern comparisons were expanded, similarity in genomic fingerprint dropped off dramatically. Prediction analysis of microarrays (PAM) based on the clustering pattern generated 49 predictor genes that can be used for sample discrimination. Moreover, a significant analysis of Microarrays (SAM) identified 598 genes being modulated by tested chemicals with a variety of biological processes, such as cell cycle, metabolism, and protein binding and KEGG pathways being significantly (p < 0.05) affected. It is feasible to distinguish structurally different PAHs based on their genomic fingerprints, which are mechanism based.

  11. Nuclear translocation of Nrf2 and expression of antioxidant defence genes in THP-1 cells exposed to carbon nanotubes.

    Science.gov (United States)

    Brown, David M; Donaldson, Kenneth; Stone, Vicki

    2010-06-01

    Carbon nanotubes have a wide range of applications in various industries and their use is likely to rise in the future. Currently, a major concern is that with the increasing use and production of these materials, there may be increased health risks to exposed workers. Long (> 15 microm) straight nanotubes may undergo frustrated phagocytosis which is likely to result in reduced clearance. We examine here the effects of multiwalled carbon nanotubes of different sizes on monocytic THP-1 cells, with regard to their ability to stimulate increased expression of the HO-1 and GST genes and their ability to produce nuclear translocation of the transcription factor, Nrf2, as well as the release of several pro-inflammatory cytokines and mediators of inflammation. Our results suggest that long (50 microm) carbon nanotubes (62.5 microg/ml for 4 hours) produce increased nuclear translocation of Nrf2 and increased HO-1 gene expression compared with shorter entangled nanotubes. There was no increased gene expression for GST. The long nanotubes (NT1) caused increased release of the proinflammatory cytokine IL-1beta, an effect which was diminished by the antioxidant trolox, suggesting a role of oxidative stress in the upregulation of this cytokine. Tentatively, our study suggests that long carbon nanotubes may exert their effect in THP-1 cells in part via an oxidative stress mechanism.

  12. Role of Oxides of Nitrogen in Tobacco-Specific Nitrosamine Formation in Flue-Cured Tobacco

    Directory of Open Access Journals (Sweden)

    Nestor TB

    2014-12-01

    Full Text Available Tobacco is known to contain a class of nitrosamines known as tobacco-specific nitrosamines or TSNA. Nitrosation of naturally occurring tobacco alkaloids is commonly accepted as the mechanism of TSNA formation in tobacco. Because green and freshly harvested tobaccos are virtually free of TSNA, formation and accumulation of TSNA are generally considered to occur during the curing process. Most recent hypotheses have focused on microbial reduction of nitrate to nitrite and other oxides of nitrogen (NOcompounds that react with tobacco alkaloids to form TSNA during curing. This natural microbial process remains the prevalent hypothesis for TSNA formation in burley and other air-cured tobaccos. However, a different mechanism for the formation of TSNA in flue-cured tobacco, independent of microbial activity, is documented in this paper. It is common practice to flue-cure Virginia or blonde tobacco in bulk barns that incorporate forced air ventilation and temperature control. For the last thirty-five years, many modern bulk barns in North America generally have used liquid propane gas (LPG with direct-fired burners that exhaust combustion gases directly into the barn where the tobacco is exposed to those gases. Our studies indicate that LPG combustion by-products in the exhaust stream, namely NO, react with naturally occurring tobacco alkaloids to form TSNA. Heat exchange curing methods preclude exposure of the tobacco to combustion gases and by-products, thereby eliminating this significant source of TSNA formation, without degrading leaf quality or smoking character. Research findings from 1998 and 1999 are presented to demonstrate the role of NOgases in TSNA formation and the significance of direct-fired curing as a primary source of TSNA formation in flue-cured tobacco. Also, data from an extensive barn conversion program in 2000, which resulted in a 94% average reduction in TSNA levels in cured flue-cured leaf, are presented.

  13. Tobacco use and exposure to tobacco promoting and restraining factors among adolescents in a developing country.

    Science.gov (United States)

    Doku, D; Koivusilta, L; Raisamo, S; Rimpelä, A

    2012-08-01

    With a long history of tobacco cultivation, adolescents in Ghana are at relatively high risk of the emerging tobacco epidemic in developing countries. This study explored exposure to tobacco promoting/restraining factors and their associations with smoking and tawa (traditional smokeless tobacco) use among 13-18-year-old Ghanaians. School-based representative data were collected in 2008 (n = 1165). Prevalence rates of tobacco use, smoking and tawa use were 9.1% (11.5% boys and 6.4% girls), 6.6% (8.0% boys and 4.7% girls) and 5.7% (7.3% boys and 3.9% girls), respectively. Four percent of the respondents attended schools without a smoking ban, 66% had been taught about the harmful effects of smoking in the current school year, and 53% had been exposed to tobacco advertising. Fifty-three percent of adolescents who had tried to purchase tobacco products were not refused because of their age. Multivariate analyses found that attendance at a school where smoking was allowed, not having been taught about the harmful effects of smoking, exposure to tobacco advertising and parental smoking were positively associated with tobacco use, and knowledge that smoking is harmful to health and difficult to quit were negatively associated with tobacco use. Both smoking and tawa use were relatively low among Ghanaian adolescents. Exposure to tobacco advertising was high. There is no tobacco legislation in Ghana, but societal norms or cultural values seem to restrict smoking in schools and access to tobacco products. Copyright © 2012 The Royal Society for Public Health. Published by Elsevier Ltd. All rights reserved.

  14. HMGB1 Is Involved in IFN-α Production and TRAIL Expression by HIV-1-Exposed Plasmacytoid Dendritic Cells: Impact of the Crosstalk with NK Cells.

    Directory of Open Access Journals (Sweden)

    Héla Saïdi

    2016-02-01

    Full Text Available Plasmacytoid dendritic cells (pDCs are innate sensors of viral infections and important mediators of antiviral innate immunity through their ability to produce large amounts of IFN-α. Moreover, Toll-like receptor 7 (TLR7 and 9 (TLR9 ligands, such as HIV and CpG respectively, turn pDCs into TRAIL-expressing killer pDCs able to lyse HIV-infected CD4+ T cells. NK cells can regulate antiviral immunity by modulating pDC functions, and pDC production of IFN-α as well as cell-cell contact is required to promote NK cell functions. Impaired pDC-NK cell crosstalk was reported in the setting of HIV-1 infection, but the impact of HIV-1 on TRAIL expression and innate antiviral immunity during this crosstalk is unknown. Here, we report that low concentrations of CCR5-tropic HIV-1Ba-L promote the release of pro-inflammatory cytokines such as IFN-α, TNF-α, IFN-γ and IL-12, and CCR5-interacting chemokines (MIP-1α and MIP-1β in NK-pDCs co-cultures. At high HIV-1BaL concentrations, the addition of NK cells did not promote the release of these mediators, suggesting that once efficiently triggered by the virus, pDCs could not integrate new activating signals delivered by NK cells. However, high HIV-1BaL concentrations were required to trigger IFN-α-mediated TRAIL expression at the surface of both pDCs and NK cells during their crosstalk. Interestingly, we identified the alarmin HMGB1, released at pDC-NK cell synapse, as an essential trigger for the secretion of IFN-α and IFN-related soluble mediators during the interplay of HIV-1 exposed pDCs with NK cells. Moreover, HMGB1 was found crucial for mTRAIL translocation to the plasma membrane of both pDCs and NK cells during their crosstalk following pDC exposure to HIV-1. Data from serum analyses of circulating HMGB1, HMGB1-specific antibodies, sTRAIL and IP-10 in a cohort of 67 HIV-1+ patients argue for the in vivo relevance of these observations. Altogether, these findings identify HMGB1 as a trigger for IFN

  15. HMGB1 Is Involved in IFN-α Production and TRAIL Expression by HIV-1-Exposed Plasmacytoid Dendritic Cells: Impact of the Crosstalk with NK Cells.

    Science.gov (United States)

    Saïdi, Héla; Bras, Marlène; Formaglio, Pauline; Melki, Marie-Thérèse; Charbit, Bruno; Herbeuval, Jean-Philippe; Gougeon, Marie-Lise

    2016-02-01

    Plasmacytoid dendritic cells (pDCs) are innate sensors of viral infections and important mediators of antiviral innate immunity through their ability to produce large amounts of IFN-α. Moreover, Toll-like receptor 7 (TLR7) and 9 (TLR9) ligands, such as HIV and CpG respectively, turn pDCs into TRAIL-expressing killer pDCs able to lyse HIV-infected CD4+ T cells. NK cells can regulate antiviral immunity by modulating pDC functions, and pDC production of IFN-α as well as cell-cell contact is required to promote NK cell functions. Impaired pDC-NK cell crosstalk was reported in the setting of HIV-1 infection, but the impact of HIV-1 on TRAIL expression and innate antiviral immunity during this crosstalk is unknown. Here, we report that low concentrations of CCR5-tropic HIV-1Ba-L promote the release of pro-inflammatory cytokines such as IFN-α, TNF-α, IFN-γ and IL-12, and CCR5-interacting chemokines (MIP-1α and MIP-1β) in NK-pDCs co-cultures. At high HIV-1BaL concentrations, the addition of NK cells did not promote the release of these mediators, suggesting that once efficiently triggered by the virus, pDCs could not integrate new activating signals delivered by NK cells. However, high HIV-1BaL concentrations were required to trigger IFN-α-mediated TRAIL expression at the surface of both pDCs and NK cells during their crosstalk. Interestingly, we identified the alarmin HMGB1, released at pDC-NK cell synapse, as an essential trigger for the secretion of IFN-α and IFN-related soluble mediators during the interplay of HIV-1 exposed pDCs with NK cells. Moreover, HMGB1 was found crucial for mTRAIL translocation to the plasma membrane of both pDCs and NK cells during their crosstalk following pDC exposure to HIV-1. Data from serum analyses of circulating HMGB1, HMGB1-specific antibodies, sTRAIL and IP-10 in a cohort of 67 HIV-1+ patients argue for the in vivo relevance of these observations. Altogether, these findings identify HMGB1 as a trigger for IFN

  16. Comprehensive analysis of 5-aminolevulinic acid dehydrogenase (ALAD variants and renal cell carcinoma risk among individuals exposed to lead.

    Directory of Open Access Journals (Sweden)

    Dana M van Bemmel

    Full Text Available BACKGROUND: Epidemiologic studies are reporting associations between lead exposure and human cancers. A polymorphism in the 5-aminolevulinic acid dehydratase (ALAD gene affects lead toxicokinetics and may modify the adverse effects of lead. METHODS: The objective of this study was to evaluate single-nucleotide polymorphisms (SNPs tagging the ALAD region among renal cancer cases and controls to determine whether genetic variation alters the relationship between lead and renal cancer. Occupational exposure to lead and risk of cancer was examined in a case-control study of renal cell carcinoma (RCC. Comprehensive analysis of variation across the ALAD gene was assessed using a tagging SNP approach among 987 cases and 1298 controls. Occupational lead exposure was estimated using questionnaire-based exposure assessment and expert review. Odds ratios (OR and 95% confidence intervals (CI were calculated using logistic regression. RESULTS: The adjusted risk associated with the ALAD variant rs8177796(CT/TT was increased (OR = 1.35, 95%CI = 1.05-1.73, p-value = 0.02 when compared to the major allele, regardless of lead exposure. Joint effects of lead and ALAD rs2761016 suggest an increased RCC risk for the homozygous wild-type and heterozygous alleles ((GGOR = 2.68, 95%CI = 1.17-6.12, p = 0.01; (GAOR = 1.79, 95%CI = 1.06-3.04 with an interaction approaching significance (p(int = 0.06. No significant modification in RCC risk was observed for the functional variant rs1800435(K68N. Haplotype analysis identified a region associated with risk supporting tagging SNP results. CONCLUSION: A common genetic variation in ALAD may alter the risk of RCC overall, and among individuals occupationally exposed to lead. Further work in larger exposed populations is warranted to determine if ALAD modifies RCC risk associated with lead exposure.

  17. Linking Global Youth Tobacco Survey (GYTS) data to tobacco control policy in Turkey--2003 and 2009.

    Science.gov (United States)

    Erguder, Toker; Polat, Halil; Arpad, Ceylan; Khoury, Rule Nabil; Warren, Charles W; Lee, Juliette; Lea, Veronica

    2012-03-01

    The purpose of this paper is to use data from the Global Youth Tobacco Survey (GYTS) conducted in Turkey in 2003 and 2009 to examine changes in tobacco use and important tobacco control measures. The GYTS were conducted in grades 7-9 in 2003 and 7-10 in 2009 in Turkey. Data in this paper are limited to 13 to 15 year old students. A total of 15,957 students from 202 schools participated in 2003 and 5,054 students from 69 schools participated in 2009. The overall response rate was 92.1% in 2003 and 87.5% in 2009. Between 2003 and 2009 current cigarette smoking did not change significantly for either boys (9.4% to 10.2%) or girls (3.5% to 5.3%). Current cigarette smoking was higher among boys than girls in 2003 and in 2009. In 2009, half of students reported they had been exposed to second hand smoking (SHS) at home and 80% reported they had been exposed to SHS in public places. Three in ten students reported they had been exposed to pro-tobacco advertising in newspapers or magazines; one in ten had an object with a cigarette brand logo on it; and 7% had been offered free cigarettes by a cigarette company representative. Two-thirds of current cigarette smokers reported that they wanted to stop smoking; and almost two-thirds had been taught in school in the past year about the dangers of smoking. Passing and implementing the Law No. 4207 on Prevention of Hazards of Tobacco Products, ratifying the World Health Organization Framework Convention on Tobacco Control (WHO FCTC), raising tax on tobacco, and requiring pictorial warning labels were important steps forward for tobacco control in Turkey. However, as to the tobacco control much work yet to be accomplished including developing an effective enforcement plan for all tobacco control efforts.

  18. Tobacco use among students in Bihar (India).

    Science.gov (United States)

    Sinha, Dhirendra N; Gupta, Prakash C; Pednekar, Mangesh

    2004-01-01

    Determination of the prevalence and attitudes toward tobacco use was assessed among 13-15 years school students in Bihar (India). Schools having grade 8-10 in Bihar. A two stage cluster sample design was used. SUDAAN and the C-sample procedure in Epi-Info was used for statistical analysis. Of the 2636 respondents, 71.8% (76.5% boys, 57.2% girls) were ever tobacco users; of them 48.9% had used tobacco before 10 years of age. Current use was reported by 58.9% (Boys 61.4%, Girls 51.2%); smokeless tobacco by 55.6% (Boys 57.6%, Girls 49.2%); and smoking by 19.4% (23.0% boys, 7.8% girls). Nearly one third (29%) students were exposed to ETS inside their homes and nearly half (48%) outside their homes. Almost all students reported watching cigarette and gutka advertisements in almost all kinds of media and events. Tobacco use by parents and friends, knowledge on harmful effects of chewing tobacco, smoking and environmental smoke, and attitudes on tobacco use by others were strongly associated with student tobacco use. Current tobacco use was reported significantly more by students who received pocket money/or were earning than by students who did not receive any pocket money/or did not earn (p value for trend <0.0001). Over half of current users (56%) bought their tobacco products from stores; of these, over 3/4th (77.2%) of them despite their age, had no difficulty in procuring these products. Teaching in schools regarding harmful effects of tobacco use was non-existent (3%). This urgently requires a comprehensive prevention program in schools and the community especially targeted towards girls.

  19. Tobacco, e-cigarettes, and child health.

    Science.gov (United States)

    Peterson, Lisa A; Hecht, Stephen S

    2017-04-01

    The availability of the Children's Health Exposure Assessment Resource funded by the National Institute of Environmental Health Sciences provides new opportunities for exploring the role of tobacco smoke exposure in causing harm to children. Children of smokers are exposed to nicotine and other harmful tobacco smoke chemicals in utero as well as in their environment. This passive exposure to tobacco smoke has a variety of negative effects on children. In-utero exposure to tobacco smoke causes poor birth outcomes and influences lung, cardiovascular, and brain development, placing children at increased risk of a number of adverse health outcomes later in life, such as obesity, behavioral problems, and cardiovascular disease. Furthermore, most smokers start in their adolescence, an age of increased nicotine addiction risk. Biomarkers of tobacco exposure helps clarify the role tobacco chemicals play in influencing health both in childhood and beyond. Although electronic cigarettes (e-cigarettes) appear to be a nicotine delivery device of reduced harm, it appears to be a gateway to the use of combustible cigarette smoking in adolescents. Pediatric researchers interested in elucidating the role of tobacco smoke exposure in adverse outcomes in children should incorporate biomarkers of tobacco exposure in their studies.

  20. Strategic marketing in the UK tobacco industry.

    Science.gov (United States)

    Anderson, Susan; Hastings, Gerard; MacFadyen, Lynn

    2002-08-01

    Tobacco-industry marketing has played a central part in the global spread of tobacco use and addiction. Although the absolute size of the tobacco market has dwindled, the industry is still immensely successful, largely due to sophisticated and manipulative marketing strategies. The UK tobacco industry identifies target groups and builds enduring relationships based on careful brand management. Potential customers are exposed to brands which are likely to appeal to them most. Tobacco companies tailor their products to target markets by altering the content of tar and nicotine, and by adding flavourings to produce a distinctive taste. Marketing strategies ensure that the products are promoted heavily at the point of sale, and directed advertising and sponsorship agreements are used to increase the visibility of the brand and strengthen its image. Tobacco companies also target non-consumer organisations such as retailers and policy makers with the aim of creating the best possible business environment for tobacco sales. We review published evidence, internal-advertising-agency documents, and observational data about tobacco promotion, and discuss the use of targeted marketing strategies in the UK.

  1. Tobacco, E-Cigarettes and Child Health

    Science.gov (United States)

    Peterson, Lisa A.; Hecht, Stephen S.

    2017-01-01

    Purpose of the review The availability of the Children’s Health Exposure Assessment Resource funded by the National Institute of Environmental Health Sciences provides new opportunities for exploring the role of tobacco smoke exposure in causing harm to children. Findings Children of smokers are exposed to nicotine and other harmful tobacco smoke chemicals in utero as well as in their environment. This passive exposure to tobacco smoke has a variety of negative effects on children. In utero exposure to tobacco smoke causes poor birth outcomes and influences lung, cardiovascular and brain development, placing children at increased risk of a number of adverse health outcomes later in life such as obesity, behavioral problems and cardiovascular disease. Furthermore, most smokers start in their adolescence, an age of increased nicotine addiction risk. Biomarkers of tobacco exposure helps clarify the role tobacco chemicals play in influencing health both in childhood and beyond. While e-cigarettes appear to be a nicotine delivery device of reduced harm, it appears to be a gateway to the use of combustible cigarette smoking in adolescents. Summary Pediatric researchers interested in elucidating the role of tobacco smoke exposure in adverse outcomes in children should incorporate biomarkers of tobacco exposure in their studies. PMID:28059903

  2. Arabidopsis ASYMMETRIC LEAVES2 protein required for leaf morphogenesis consistently forms speckles during mitosis of tobacco BY-2 cells via signals in its specific sequence.

    Science.gov (United States)

    Luo, Lilan; Ando, Sayuri; Sasabe, Michiko; Machida, Chiyoko; Kurihara, Daisuke; Higashiyama, Tetsuya; Machida, Yasunori

    2012-09-01

    Leaf primordia with high division and developmental competencies are generated around the periphery of stem cells at the shoot apex. Arabidopsis ASYMMETRIC-LEAVES2 (AS2) protein plays a key role in the regulation of many genes responsible for flat symmetric leaf formation. The AS2 gene, expressed in leaf primordia, encodes a plant-specific nuclear protein containing an AS2/LOB domain with cysteine repeats (C-motif). AS2 proteins are present in speckles in and around the nucleoli, and in the nucleoplasm of some leaf epidermal cells. We used the tobacco cultured cell line BY-2 expressing the AS2-fused yellow fluorescent protein to examine subnuclear localization of AS2 in dividing cells. AS2 mainly localized to speckles (designated AS2 bodies) in cells undergoing mitosis and distributed in a pairwise manner during the separation of sets of daughter chromosomes. Few interphase cells contained AS2 bodies. Deletion analyses showed that a short stretch of the AS2 amino-terminal sequence and the C-motif play negative and positive roles, respectively, in localizing AS2 to the bodies. These results suggest that AS2 bodies function to properly distribute AS2 to daughter cells during cell division in leaf primordia; and this process is controlled at least partially by signals encoded by the AS2 sequence itself.

  3. AKR1C1 as a Biomarker for Differentiating the Biological Effects of Combustible from Non-Combustible Tobacco Products.

    Science.gov (United States)

    Woo, Sangsoon; Gao, Hong; Henderson, David; Zacharias, Wolfgang; Liu, Gang; Tran, Quynh T; Prasad, G L

    2017-05-03

    Smoking has been established as a major risk factor for developing oral squamous cell carcinoma (OSCC), but less attention has been paid to the effects of smokeless tobacco products. Our objective is to identify potential biomarkers to distinguish the biological effects of combustible tobacco products from those of non-combustible ones using oral cell lines. Normal human gingival epithelial cells (HGEC), non-metastatic (101A) and metastatic (101B) OSCC cell lines were exposed to different tobacco product preparations (TPPs) including cigarette smoke total particulate matter (TPM), whole-smoke conditioned media (WS-CM), smokeless tobacco extract in complete artificial saliva (STE), or nicotine (NIC) alone. We performed microarray-based gene expression profiling and found 3456 probe sets from 101A, 1432 probe sets from 101B, and 2717 probe sets from HGEC to be differentially expressed. Gene Set Enrichment Analysis (GSEA) revealed xenobiotic metabolism and steroid biosynthesis were the top two pathways that were upregulated by combustible but not by non-combustible TPPs. Notably, aldo-keto reductase genes, AKR1C1 and AKR1C2 , were the core genes in the top enriched pathways and were statistically upregulated more than eight-fold by combustible TPPs. Quantitative real time polymerase chain reaction (qRT-PCR) results statistically support AKR1C1 as a potential biomarker for differentiating the biological effects of combustible from non-combustible tobacco products.

  4. In vitro response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells exposed to 60Co at single fraction

    International Nuclear Information System (INIS)

    Andrade, Lidia Maria; Campos, Tarcisio Passos Ribeiro de; Leite, M.F.; Goes, A.M.

    2005-01-01

    Radiotherapy using gamma rays is a common modality of breast cancer treatment. The aim of this research is to investigate the biological response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells (PBMC) exposed in vitro to 60 Co irradiation at a single fraction of 10 Gy, 25 Gy and 50 Gy doses at 136,4 cGy.min -1 rate. Cells were irradiated at room temperature by the Theratron 80 radiotherapy system. Biological response was evaluated through cellular viability using MTT assay and nucleus damages visualized by Propidium Iodide assay and electrophoresis agarose gel after gamma irradiation. Nucleus damages induced by 60 Co irradiation were compared to damage caused by cell exposure to 10% methanol. The 50 Gy dose of irradiation did not stimulate nucleus damages at the same level as that affected by 10% methanol induction in the MDAMB-231. Further studies are necessary to understand these mechanisms in the MDAMB-231 human breast carcinoma cell line.(author)

  5. In vitro response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells exposed to {sup 60}Co at single fraction

    Energy Technology Data Exchange (ETDEWEB)

    Andrade, Lidia Maria; Campos, Tarcisio Passos Ribeiro de [Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil). Dept. de Engenharia Nuclear]. E-mail: lidia.andrade@unifenas.br; Leite, M.F. [Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil). Dept. de Fisiologia e Biofisica; Goes, A.M. [Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil). Dept. de Bioquimica e Imunologia

    2005-10-15

    Radiotherapy using gamma rays is a common modality of breast cancer treatment. The aim of this research is to investigate the biological response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells (PBMC) exposed in vitro to {sup 60} Co irradiation at a single fraction of 10 Gy, 25 Gy and 50 Gy doses at 136,4 cGy.min{sup -1} rate. Cells were irradiated at room temperature by the Theratron 80 radiotherapy system. Biological response was evaluated through cellular viability using MTT assay and nucleus damages visualized by Propidium Iodide assay and electrophoresis agarose gel after gamma irradiation. Nucleus damages induced by {sup 60} Co irradiation were compared to damage caused by cell exposure to 10% methanol. The 50 Gy dose of irradiation did not stimulate nucleus damages at the same level as that affected by 10% methanol induction in the MDAMB-231. Further studies are necessary to understand these mechanisms in the MDAMB-231 human breast carcinoma cell line.(author)

  6. Differentiated THP-1 Cells Exposed to Pathogenic and Nonpathogenic Borrelia Species Demonstrate Minimal Differences in Production of Four Inflammatory Cytokines.

    Science.gov (United States)

    Stokes, John V; Moraru, Gail M; McIntosh, Chelsea; Kummari, Evangel; Rausch, Keiko; Varela-Stokes, Andrea S

    2016-11-01

    Tick-borne borreliae include Lyme disease and relapsing fever agents, and they are transmitted primarily by ixodid (hard) and argasid (soft) tick vectors, respectively. Tick-host interactions during feeding are complex, with host immune responses influenced by biological differences in tick feeding and individual differences within and between host species. One of the first encounters for spirochetes entering vertebrate host skin is with local antigen-presenting cells, regardless of whether the tick-associated Borrelia sp. is pathogenic. In this study, we performed a basic comparison of cytokine responses in THP-1-derived macrophages after exposure to selected borreliae, including a nonpathogen. By using THP-1 cells, differentiated to macrophages, we eliminated variations in host response and reduced the system to an in vitro model to evaluate the extent to which the Borrelia spp. influence cytokine production. Differentiated THP-1 cells were exposed to four Borrelia spp., Borrelia hermsii (DAH), Borrelia burgdorferi (B31), B. burgdorferi (NC-2), or Borrelia lonestari (LS-1), or lipopolysaccharides (LPS) (activated) or media (no treatment) controls. Intracellular and secreted interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α were measured using flow cytometric and Luminex-based assays, respectively, at 6, 24, and 48 h postexposure time points. Using a general linear model ANOVA for each cytokine, treatment (all Borrelia spp. and LPS compared to no treatment) had a significant effect on secreted TNF-α only. Time point had a significant effect on intracellular IFN-γ, TNF-α and IL-6. However, we did not see significant differences in selected cytokines among Borrelia spp. Thus, in this model, we were unable to distinguish pathogenic from nonpathogenic borreliae using the limited array of selected cytokines. While unique immune profiles may be detectable in an in vitro model and may reveal predictors for pathogenicity in borreliae

  7. Uranyl nitrate-exposed rat alveolar macrophages cell death: Influence of superoxide anion and TNF α mediators

    International Nuclear Information System (INIS)

    Orona, N.S.; Tasat, D.R.

    2012-01-01

    Uranium compounds are widely used in the nuclear fuel cycle, military and many other diverse industrial processes. Health risks associated with uranium exposure include nephrotoxicity, cancer, respiratory, and immune disorders. Macrophages present in body tissues are the main cell type involved in the internalization of uranium particles. To better understand the pathological effects associated with depleted uranium (DU) inhalation, we examined the metabolic activity, phagocytosis, genotoxicity and inflammation on DU-exposed rat alveolar macrophages (12.5–200 μM). Stability and dissolution of DU could differ depending on the dissolvent and in turn alter its biological action. We dissolved DU in sodium bicarbonate (NaHCO 3 100 mM) and in what we consider a more physiological vehicle resembling human internal media: sodium chloride (NaCl 0.9%). We demonstrate that uranyl nitrate in NaCl solubilizes, enters the cell, and elicits its cytotoxic effect similarly to when it is diluted in NaHCO 3 . We show that irrespective of the dissolvent employed, uranyl nitrate impairs cell metabolism, and at low doses induces both phagocytosis and generation of superoxide anion (O 2 − ). At high doses it provokes the secretion of TNFα and through all the range of doses tested, apoptosis. We herein suggest that at DU low doses O 2 − may act as the principal mediator of DNA damage while at higher doses the signaling pathway mediated by O 2 − may be blocked, prevailing damage to DNA by the TNFα route. The study of macrophage functions after uranyl nitrate treatment could provide insights into the pathophysiology of uranium‐related diseases. -- Highlights: ► Uranyl nitrate effect on cultured macrophages is linked to the doses and independent of its solubility. ► At low doses uranyl nitrate induces generation of superoxide anion. ► At high doses uranyl nitrate provokes secretion of TNFα. ► Uranyl nitrate induces apoptosis through all the range of doses tested.