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Biomedical subjects

B T Mossman

Publications and source records attributed to B T Mossman.

At least 55 records · Page 3Linked to original sources
PubMed1996

Increased neutrophil adherence to endothelial cells exposed to asbestos.

Inhalation of asbestos may activate the pulmonary endothelium to promote an inflammatory cell phenotype that participates in the development of pulmonary fibrosis. However, little is known about the effects of asbestos on endothelial cell function. Therefore, endothelial cells were exposed to chrysotile and crocidolite asbestos for up to 72 hr to investigate the effects of noncytotoxic concentrations of asbestos on cell function. Noncytolytic concentrations of chrysotile and crocidolite caused localized changes in cell morphology, resulting in activation of endothelial cells to a vacuolated, "spindle-shaped" morphology at sites of fiber deposition. The adherence of neutrophils (PMN) to control and asbestos-treated cultures was examined to determine the functional significance of this altered morphology. Chrysotile asbestos caused a time-dependent, 2- to 4-fold increase in PMN adherence that was localized to spindled endothelial cells in close contact with fibers. Monoclonal antibodies against intercellular adhesion molecule-1 (ICAM-1) inhibited this asbestos-induced PMN adherence. Solution hybridization demonstrated a 2.5-fold increase in steady-state levels of ICAM-1 mRNA in cells exposed to chrysotile asbestos for 8 hr. Exposure to crocidolite asbestos resulted in similar alteration in cell morphology and increase in PMN binding to endothelial cells, while a physically similar nonasbestos fiber, refractory ceramic fiber-1 did not cause morphologic change or increased PMN binding. These data suggest that asbestos fibers can specifically and directly activate endothelial cells, resulting in an activated cell morphology, increased ICAM-1 expression, and enhanced PMN adherence.

Animals↗
PubMed1996

Transfection of a manganese-containing superoxide dismutase gene into hamster tracheal epithelial cells ameliorates asbestos-mediated cytotoxicity.

To determine if overexpression of manganese-containing SOD (MnSOD) alters cell sensitivity to asbestos, an expression cassette containing murine MnSOD cDNA was cotransfected with pSV2neo, a plasmid conferring resistance to the antibiotic G418, into a diploid cell line of hamster tracheal epithelial (HTE) cells. Pools of G418-resistant transfectants were characterized by Southern and Northern blot analyses and enzyme activity assays. Although increases in MnSOD gene copies in individual cell pools ranged from approximately 7- to 86-fold in comparison to cells transfected with pSV2neo alone, steady-state levels of MnSOD mRNA were increased only by 1.4-to 2.3-fold. Despite modest increases in MnSOD mRNA, significant elevations in MnSOD enzyme activity were observed in pools of G418-resistant cells. MnSOD-transfected cell lines were more resistant to the cytotoxic effects of crocidolite asbestos using a sensitive colony-forming efficiency (CFE) assay. These data show that MnSOD has a direct role in cell defense against asbestos-induced cytotoxicity, an oxidant-dependent process.

Animals↗
PubMed1996

Apoptosis is observed in mesothelial cells after exposure to crocidolite asbestos.

Asbestos causes protracted, dose-dependent increases in steady-state mRNA levels of the proto-oncogenes c-fos and c-jun, and AP-1 DNA-binding activity in normal rat pleural mesothelial (RPM) cells (1). To determine the phenotypic end points of overexpression of these early response genes by asbestos, both cell proliferation and apoptosis were examined in confluent RPM cells exposed to a range of concentrations (1.25 to 10 micrograms/cm2 dish) of crocidolite asbestos for 24 and 48 h. Quantitation of RPM cells pulsed with 5-bromo-2'-deoxyuridine revealed that asbestos caused dose-dependent decreases in cells undergoing DNA synthesis. Decreases in cell proliferation were accompanied by dose-related increases in apoptosis using (1) terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (i.e., ApopTag technique), (2) 4',6-diamidino-2-phenylindole cell staining, and (3) fluorescent-activated cell sorter after incorporation of propidium iodide. Less striking but significant dose-related increases in apoptosis were observed in RPM cells exposed to H2O2 (300 microM), and no apoptosis was seen after exposure of cells to high concentrations (10 micrograms/cm2 dish) of glass beads. Our results are unique in that they demonstrate that asbestos induces apoptosis in mesothelial cells at concentrations eliciting increased expression of the proto-oncogenes c-fos and c-jun.

Animals↗
PubMed1996

Mechanisms of carcinogenesis and clinical features of asbestos-associated cancers.

Exposure to asbestos, particularly members of the amphibole subgroup (crocidolite, amosite), is associated with the development of malignant mesothelioma and lung cancer. Although management of asbestos in buildings and increased regulation of asbestos in workplace settings are viable approaches to the prevention of disease, the prognosis of asbestos-associated tumors is generally dismal. Moreover, although a vast amount of information is available on the responses of cells and tissues to fibers, understanding the pathogenesis of asbestos-associated malignancies is hampered by the complexity of and differences between various fiber types. Multiple interactions between components of cigarette smoke and asbestos may be important in the development of lung cancer. In this article, the general properties of asbestos fibers will be discussed with an emphasis on chemical and physical features implicated in tumorigenesis. We will then provide a brief overview of the clinical features and treatment of cancers associated with exposure to asbestos. Finally, we will review recent experimental data providing some insight into the cellular and molecular mechanisms of carcinogenesis by asbestos.

Asbestos↗
PubMed1995

Asbestos induces nuclear factor kappa B (NF-kappa B) DNA-binding activity and NF-kappa B-dependent gene expression in tracheal epithelial cells.

Nuclear factor kappa B (NF-kappa B) is a transcription factor regulating expression of genes intrinsic to inflammation and cell proliferation--features of asbestos-associated diseases. In studies here, crocidolite asbestos caused protracted and dose-responsive increases in proteins binding to nuclear NF-kappa B-binding DNA elements in hamster tracheal epithelial (HTE) cells. This binding was modulated by cellular glutathione levels. Antibodies recognizing p65 and p50 protein members of the NF-kappa B family revealed these proteins in two of the DNA complexes. Transient transfection assays with a construct containing six NF-kappa B-binding DNA consensus sites linked to a luciferase reporter gene indicated that asbestos induced transcriptional activation of NF-kappa B-dependent genes, an observation that was confirmed by northern blot analyses for c-myc mRNA levels in HTE cells. Studies suggest that NF-kappa B induction by asbestos is a key event in regulation of multiple genes involved in the pathogenesis of asbestos-related lung cancers.

Animals↗
PubMed1995

Transcriptional activation of the proto-oncogene c-jun by asbestos and H2O2 is directly related to increased proliferation and transformation of tracheal epithelial cells.

Asbestos causes persistent increases in c-jun mRNA and AP-1 DNA binding activity in hamster tracheal epithelial (HTE) cells, the progenitor cell type of asbestos-induced bronchogenic carcinoma. Studies here were designed to determine mechanisms of c-jun induction by asbestos and the phenotypic consequences of Jun expression in HTE cells. To examine whether asbestos or H2O2 induced transcription of c-jun, we transiently transfected HTE cells with a plasmid containing a fragment of the c-jun promoter coupled to a luciferase reporter gene. In addition, c-jun was overexpressed in cells using a full-length human c-jun construct, and effects on proliferation and transformation were examined. HTE cells transfected with the jun-luciferase construct showed increased luciferase activity when exposed to crocidolite asbestos or H2O2. These results demonstrate that asbestos and H2O2 activate AP-1-dependent gene transcription. Overexpression of c-jun led to increased proliferation and enhanced ability of HTE cells to grow in soft agar, an indication of cellular transformation. Data suggest that overexpression of c-jun may contribute to asbestos and oxidant-induced proliferation and carcinogenesis.

Animals↗
PubMed1995

Induction of c-fos and c-jun proto-oncogene expression by asbestos is ameliorated by N-acetyl-L-cysteine in mesothelial cells.

Asbestos fibers cause dose-dependent, persistent increases in mRNA levels of c-jun and c-fos proto-oncogenes in rat pleural mesothelial (RPM) cells, the progenitor cells of asbestos-induced mesothelioma (N. Heintz, Y. M. W. Janssen, and B. T. Mossman. Proc. Natl. Acad. Sci. USA, 90: 3299-3303, 1993). Here we report that addition of N-acetyl-L-cysteine decreases asbestos-mediated induction of c-fos and c-jun mRNA levels in a dose-dependent fashion. Exposure of RPM cells to asbestos causes depletion of total cellular glutathione, a response that can be abolished by pretreatment with N-acetyl-L-cysteine. Pretreatment of cells with buthionine sulfoximine, an agent which diminishes glutathione pools, increases the magnitude of induction of c-fos and c-jun mRNA by asbestos. To determine whether asbestos-induced effects on proto-oncogene expression could be attributed to extracellular generation of active oxygen species (AOS), RPM cells were exposed to H2O2 or xanthine and xanthine oxidase, a generating system of AOS. These oxidant stresses did not decrease cellular glutathione levels nor alter mRNA levels of c-fos or c-jun. However, increased mRNA levels of manganese-containing superoxide dismutase and heme oxygenase were observed, indicating that RPM cells respond to AOS by increased expression of genes encoding antioxidant enzymes. These data indicate that the signaling pathways leading to c-fos/c-jun proto-oncogene induction by asbestos are not triggered directly by formation of extracellular AOS. However, intracellular thiol levels appear to influence the expression of c-fos and c-jun, suggesting a redox-sensitive component in the signaling cascade which modulates gene expression of c-fos and c-jun by asbestos.

Acetylcysteine↗
PubMed1995

TNF alpha and increased chemokine expression in rat lung after particle exposure.

Macrophage inflammatory protein 2 (MIP-2) and CINC (Cytokine-Induced-Neutrophil-Chemoattractant) are members of the chemokine family of inflammatory and immunoregulatory cytokines. MIP-2 and CINC exhibit potent neutrophil chemotactic activity and are thought to be key mediators of inflammatory cell recruitment in response to tissue injury and infection. In the present studies, we examined the potential involvement of MIP-2 and CINC in particle-elicited inflammation in the rat lung and the role of TNF alpha in particle-induced chemokine expression. Acute intratracheal instillation exposure of F344 rats to alpha quartz or titanium dioxide was shown to markedly increase steady-state levels of MIP-2 and CINC mRNA in lung tissue; a response which was associated with a significant increase in neutrophils in the bronchoalveolar lavage fluid. Additional studies demonstrated that acute inhalation of crocidolite fibers by rats also induced increased MIP-2 and CINC expression. Since previous studies had demonstrated that TNF alpha stimulates MIP-2 and CINC expression in vitro and that particle exposure induces TNF alpha production in rat lung we examined the role of TNF alpha in alpha quartz-induced MIP-2 gene expression. We demonstrated that passive immunization of mice against TNF alpha markedly attenuated the increased lung MIP-2 mRNA seen in response to alpha quartz inhalation. Collectively, these findings suggest that the chemokines MIP-2 and CINC play a role in neutrophil recruitment to the rat lung after particle exposure and indicate that particle-induced expression of these chemokines is mediated, at least in part, by production of TNF alpha.

Animals↗
PubMed1995

Patterns of inflammation, cell proliferation, and related gene expression in lung after inhalation of chrysotile asbestos.

Biochemical and molecular markers of inflammation, cell proliferation, and pulmonary fibrosis were studied in lungs and bronchoalveolar lavage preparations from Fischer 344 rats at time periods from 3 to 20 days after inhalation of two airborne concentrations (0.18 and 8.2 mg/m3 air) of chrysotile asbestos. Additional groups of animals were examined for lung histopathology and cell proliferation with an antibody to 5-bromo-2'-deoxyuridine after exposure to asbestos for 5 and 20 days and after 20 days of exposure followed by an additional 20 days in room air. Exposure to chrysotile at the higher concentration caused protracted increases in steady-state mRNA levels of manganese-containing superoxide dismutase and elevation in glyceraldehyde-3-phosphate dehydrogenase mRNA at 3 days, but levels of mRNAs encoding copper-zinc-containing superoxide dismutase, ornithine decarboxylase, and the proto-oncogene, c-jun were not statistically elevated from levels occurring in lung homogenates from sham control rats. Differential cell counts in bronchoalveolar lavage revealed an early infiltration of neutrophils that correlated with focal areas of increased cellularity and fibrosis in rat lungs at the higher concentrations of asbestos. However, elevations in lung hydroxyproline were not observed. Significant increases in epithelial cells of the bronchi, the interstitial compartment of the lung, and mesothelial cells incorporating 5-bromo-2'-deoxyuridine, an indication of DNA synthesis, were noted in the higher chrysotile group at 5 days, but labeling in all cell compartments was comparable with that occurring in sham controls at later time points. Indicators of inflammation, increased cell proliferation, and pulmonary fibrosis were not observed in the lungs of rats exposed to the lower concentration of chrysotile. Thus, results indicate that cellular and molecular markers of inflammation and proliferation in lung are dose-related and indicative of the histopathological development of asbestosis.

Administration, Inhalation↗
PubMed1994

Increased expression of manganese-containing superoxide dismutase in rat lungs after inhalation of inflammatory and fibrogenic minerals.

Steady-state mRNA levels and immunoreactive protein for manganese-containing superoxide dismutase (MnSOD) were assayed in rat lungs after subchronic inhalation of the fibrogenic silicon dioxide, cristobalite, or preparations of titanium dioxide (TiO2) of different inflammatory and fibrogenic potential. Total and differential cell counts recoverable by bronchoalveolar lavage (BAL) were also measured to ascertain whether induction of certain antioxidant enzymes (AOE) correlated with inflammatory responses. Inhalation of cristobalite and ultra-fine TiO2, a particle causing pulmonary inflammation and fibrosis, caused dramatic increases in MnSOD mRNA levels in rat lung which correlated with increases in MnSOD immunoreactive protein. Increases in gene expression of other AOE [catalase, glutathione peroxidase (GPX), copper-zinc containing superoxide dismutase (CuZnSOD)] were less striking and did not correlate precisely with inflammatory potential of minerals. Inflammatory changes in BAL correlated directly with steady-state MnSOD mRNA levels in lung. Inhalation of TiO2-F, a noninflammatory, nonfibrogenic mineral, failed to induce MnSOD or mRNAs for other AOE. Our data suggest that particles causing inflammation and pulmonary fibrosis increase expression of AOE in lung, most notably MnSOD. Thus, elevations of MnSOD mRNA levels in lung or BAL may be predictive of lung disease.

Animals↗
PubMed1994

Carcinogenesis and related cell and tissue responses to asbestos: a review.

A review of the literature reveals that chrysotile asbestos has a mulitiplicity of effects on cells and tissues which can provide a framework for assessment of its role(s) in initiation, promotion, or as a co-carcinogen which acts in concert with chemical carcinogens found in cigarette smoke in the development of lung cancer. Several caveats important in the interpretation of these data include the general lack of dose-response protocols both for in vivo and for in vitro studies as well as the absence, in many studies, of minerals which are appropriate positive or negative controls based on epidemiological data in man. Other factors which may account for disparities in results between studies include the use of different preparations of chrysotile fibres with distinct chemical and physical compositions and different cell types and species. Whether chrysotile is an initiator of lung cancer or mesothelioma in human cells is unclear, as evidence of chromosomal aberrations in human bronchial epithelial cells are for the most part negative (Kodoma et al., 1993; see Mossman, B., this Workshop): only one study employing pleural mesothelial cells from four individuals, two of whom exhibited chromosomal abnormalities before exposure to asbestos, has documented chromosomal changes with chrysotile, crocidolite and amosite asbestos, and this study did not use a non-pathogenic dust as a negative control. Studies using human lymphocytes show chromosomal changes after exposure to latex beads or chrysotile at equal weight concentrations (Korkina et al., 1992; see Mossman, B., this Workshop). Lastly, although both chrysotile and crocidolite asbestos demonstrated dose-dependent increase in aberrant anaphases in an SV40 T antigen-transformed human mesothelial cell line (Pelin et al., 1992, see Mossman, B., this Workshop), erionite, a potent mesotheliomagenic fibre in rodents and humans, caused no aberrations in this bioassay. Several studies have been performed to determine the interactions of chrysotile with rodent cells or isolated bacterial DNA. Results in a number of bioassays have been positive, but chrysotile is less potent on a fibre number comparative basis than crocidolite and no-observed-effect-levels (NOELs) have been observed in several systems. Cell proliferation by asbestos may be a more relevant phenomenon to tumour development and promotion, and the ability of chrysotile to stimulate cell proliferation, using a number of biomarkers, has been demonstrated both in vitro and after inhalation by rats.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

Oxidant stress responses in human pleural mesothelial cells exposed to asbestos.

The generation of oxidants is a proposed mechanism of cell injury by asbestos fibers. To determine whether human pleural mesothelial cells (HMC) respond to asbestos and active oxygen species (AOS) by induction of antioxidant enzymes, cells obtained from pleural effusion were exposed to crocidolite or chrysotile asbestos or xanthine/xanthine oxidase (X/XO), a chemical-generating system of AOS. Gene expression of manganese-containing superoxide dismutase (MnSOD) and heme oxygenase (HO), endogenous enzymes involved in cell defense against oxidant stresses, was then determined. Dosage-dependent increases in steady-state mRNA levels of MnSOD and HO were observed in HMC exposed to asbestos or X/XO. However, increases in gene expression of MnSOD or HO did not occur in HMC after exposure to particulates such as polystyrene beads or riebeckite, the nonfibrous analog of crocidolite asbestos. Comparative experiments with human adult lung fibroblasts (HAL) showed less striking increases in mRNA levels of MnSOD and HO in response to asbestos, but steady-state mRNA levels for HO were increased more than fivefold in response to X/XO. To determine whether increases in mRNA levels of MnSOD were translated into protein, Western blot analyses were performed on HMC and HAL cells exposed to asbestos or X/XO. Slight increases in MnSOD immunoreactive protein were observed in HMC in response to both agents. In contrast, X/XO caused striking elevations in MnSOD protein levels in HAL cells. These results suggest that certain antioxidant enzymes are inducible in HMC after exposure to asbestos and other oxidants.

Asbestos, Crocidolite↗
PubMed1994

Dose-responsive increases in pulmonary fibrosis after inhalation of asbestos.

We focused here on steady-state mRNA levels of genes involved in antioxidant defense, i.e., manganese superoxide dismutase and copper-zinc superoxide dismutase, and in cell proliferation, i.e., ornithine decarboxylase, c-jun, and glyceraldehyde-3-phosphate-dehydrogenase in whole-lung homogenates from Fischer 344 rats at 3 h to 20 d after exposure to crocridolite asbestos. Changes in gene expression were correlated with histopathologic findings, total and differential cell counts in bronchoalveolar lavage, and levels of hydroxyproline in lung. Dosage-dependent increases in mRNA levels of antioxidant enzymes and proliferation-related genes were observed. Differential cell counts revealed a dose-related infiltration of neutrophils that preceded elevations in gene expression. Neutrophil infiltration into lung and focal lesions of fibrosis as well as increased levels of hydroxyproline were observed only at high concentrations of asbestos. These results indicate that high airborne concentrations of asbestos cause molecular changes in lung that may be related to antioxidant defense and the triggering of cell proliferation, a feature of asbestosis and lung cancer.

Administration, Inhalation↗
PubMed1994

Induction of c-fos and c-jun proto-oncogenes in target cells of the lung and pleura by carcinogenic fibers.

To study mechanisms of cell proliferation by asbestos and nonasbestos fibers, we examined the effects of these agents on the mRNA levels of c-fos and c-jun and ornithine decarboxylase (ODC) in hamster tracheal epithelial (HTE) cells and rat pleural mesothelial (RPM) cells, the progenitor cells of bronchogenic carcinoma and mesothelioma, respectively. In comparison with crocidolite asbestos, increases in c-jun mRNA were less striking in HTE cells after exposure to man-made vitreous fiber-10 (MMVF-10) or refractory ceramic fiber-1 (RCF-1). No c-fos mRNA was detected in HTE cells after exposure to particulates, but exposure of HTE cells to H2O2 caused striking increases in c-fos and c-jun, which preceded increases in ODC mRNA. Increases in ODC mRNA were also observed in HTE cells after exposure to nonasbestos fibers, whereas only crocidolite asbestos caused elevations in ODC mRNA in RPM cells. In RPM cells, crocidolite and chrysotile asbestos caused increases in mRNA levels of both c-fos and c-jun. No increases in proto-oncogene induction were observed using MMVF-10 or RCF-1 at nontoxic concentrations (< or = 5 micrograms/cm2 dish). Moreover, erionite, a fiber extremely potent in the causation of mesothelioma in humans, caused more dramatic elevations in c-fos and c-jun. Nonfibrous particles (riebeckite, polystyrene beads) did not alter proto-oncogene expression in these cell types, suggesting that the fibrous geometry of particulates is critical in the induction of c-fos and c-jun.

Animals↗
PubMed1994

Regulation of antioxidant enzymes in lung after oxidant injury.

Studies have implicated active oxygen species (AOS) in the pathogenesis of various lung diseases. Many chemical and physical agents in the environment are potent generators of AOS, including ozone, hyperoxia, mineral dusts, paraquat, etc. These agents produce AOS by different mechanisms, but frequently the lung is the primary target of toxicity, and exposure results in damage to lung tissue to varying degrees. The lung has developed defenses to AOS-mediated damage, which include antioxidant enzymes, the superoxide dismutases [copper-zinc (CuZnSOD) and manganese-containing (MnSOD)], catalase, and glutathione peroxidase (GPX). In this review, antioxidant defenses to environmental stresses in the lung as well as in isolated pulmonary cells following exposure to a number of different oxidants, are summarized. Each oxidant appears to induce a different pattern of antioxidant enzyme response in the lung, although some common trends, i.e., induction of MnSOD following oxidants inducing inflammation or pulmonary fibrosis, in responses to oxidants occur. Responses may vary between the different cell types in the lung as a function of cell-cycle or other factors. Increases in MnSOD mRNA or immunoreactive protein in response to certain oxidants may serve as a biomarker of AOS-mediated damage in the lung.

Animals↗
PubMed1994

Oxygen radicals and asbestos-mediated disease.

Asbestos fibers are potent elaborators of active oxygen species whether by reactions involving iron on the surface of the fiber, or by attempted phagocytosis of fibers by cell types resident in the lung. The link between production of active oxygen species and the pathogenesis of asbestos-mediated disease has been highlighted by studies outlined here exploring the use of antioxidant scavengers which inhibit the cytotoxic effects of asbestos both in vitro and in vivo. The use of antioxidant enzymes ameliorates the induction of certain genes necessary for cell proliferation, such as ornithine decarboxylase, implicating oxidants as causative factors in some abnormal cell replicative events. Based on these observations, antioxidant enzymes likely represent an important lung defense mechanism in response to oxidative stress. In addition, their gene expression in lung or in cells from bronchoalveolar lavage might be a valuable biomarker of chronic inflammation and pulmonary disease after inhalation of oxidants.

Administration, Inhalation↗