Free Radical Biology & Medicine

Our medical research library holds 33 studies published in Free Radical Biology & Medicine (1990–2019). Each entry links to the study's record on PubMed and, where there is one, to the publisher's page and a free full text. 33 have an abstract, the authors' own summary as PubMed records it: open "Abstract" to read it.

About the journal

• Publisher: Elsevier Science
• Published: 1987 to the present, United States
• Language: English
• ISSN: 0891-5849 (print), 1873-4596 (electronic)
• Merged from: Advances in Free Radical Biology and Medicine
• Merged from: Journal of Free Radicals in Biology & Medicine
• Its record in the NLM Catalog

2010s

• Iron addiction with ferroptosis-resistance in asbestos-induced mesothelial carcinogenesis: Toward the era of mesothelioma prevention. S. Toyokuni. 2019;133:206-215. Review PubMed · Publisher

  Abstract

  Cancer is the primary cause of human mortality in most countries. This tendency has increased as various medical therapeutics have advanced, which suggests that we cannot escape carcinogenesis, although the final outcome may be modified by exposomes and statistics. Cancer is classified by its cellular differentiation. Mesothelial cells are distinct in that they line somatic cavities, facilitating the smooth movement of organs, but are not exposed to the external environment. Malignant mesothelioma, or simply mesothelioma, develops either in the pleural, peritoneal or pericardial cavities, or in the tunica vaginalis testes. Mesothelioma has been a relatively rare cancer but is socially important due to its association with asbestos exposure, caused by modern industrial development. The major pathogenic mechanisms include oxidative stress either via catalytic reactions against the asbestos surface or frustrated phagocytosis of macrophages, and specific adsorption of hemoglobin and histones by asbestos fibers in the presence of phagocytic activity of mesothelial cells. Multiwall carbon nanotubes of ~50 nm-diameter, additionally adsorbing transferrin, are similarly carcinogenic to mesothelial cells in rodents and were thus classified as Group 2B carcinogens. Genetic alterations found in human and rat mesothelioma notably contain changes found in other excess iron-induced carcinogenesis models. Phlebotomy and iron chelation therapies have been successful in the prevention of mesothelioma in rats. Alternatively, loading of oxidative stress by non-thermal plasma to mesothelioma cells causes ferroptosis. Therefore, carcinogenesis by foreign fibrous inorganic materials may overlap the uncovered molecular mechanisms of birth of life and its evolution.

  Copyright © 2018. Published by Elsevier Inc.

• Iron from a geochemical viewpoint. Understanding toxicity/pathogenicity mechanisms in iron-bearing minerals with a special attention to mineral fibers. A.F. Gualtieri, G.B. Andreozzi, M. Tomatis, et al. 2019;133:21-37. Review PubMed · Publisher

  Abstract

  Iron and its role as soul of life on Earth is addressed in this review as iron is one of the most abundant elements of our universe, forms the core of our planet and that of telluric (i.e., Earth-like) planets, is a major element of the Earth's crust and is hosted in an endless number of mineral phases, both crystalline and amorphous. To study iron at an atomic level inside the bulk of mineral phases or at its surface, where it is more reactive, both spectroscopy and diffraction experimental methods can be used, taking advantage of nearly the whole spectrum of electromagnetic waves. These methods can be successfully combined to microscopy to simultaneously provide chemical (e.g. iron mapping) and morphological information on mineral particles, and shed light on the interaction of mineral surfaces with organic matter. This review describes the crystal chemistry of iron-bearing minerals of importance for the environment and human health, with special attention to iron in toxic minerals, and the experimental methods used for their study. Special attention is devoted to the Fenton-like chain reaction involving Fe2+ in the formation of highly reactive hydroxyl radicals. The final part of this review deals with release and adsorption of iron in biological fluids, coordinative and oxidative state of iron and in vitro reactivity. To disclose the very mechanisms of carcinogenesis induced by iron-bearing toxic mineral particles, crystal chemistry and surface chemistry are fundamental for a multidisciplinary approach which should involve geo-bio-scientists, toxicologists and medical doctors.

  Copyright © 2018 Elsevier Inc. All rights reserved.

• Modulation of reactive oxygen species via ERK and STAT3 dependent signalling are involved in the response of mesothelioma cells to exemestane. B. Nuvoli, E. Camera, A. Mastrofrancesco, et al. 2018;115:266-277. PubMed · Publisher

  Abstract

  Pleural mesothelioma is a deadly form of cancer. The prognosis is extremely poor due to the limited treatment modalities. Uptake of asbestos fibres, the leading cause of mesothelioma, lead to the accumulation of reactive-oxygen-species (ROS). Interestingly, increasing ROS production by using ROS-generating drugs may offer a strategy to selectively trigger cell death. Exemestane, an aromatase inhibitor, has previously shown anti-tumor properties in mesothelioma preclinical models suggesting a role of G protein-coupled receptor 30 (GPR30) in the drug response. As exemestane, in addition to blocking estrogen biosynthesis, generates ROS that are able to arrest the growth of breast cancer, we explored the role of ROS, antioxidant defense system, and ROS-induced signalling pathways in mesothelioma cells during exemestane response. Here we report that exemestane treatment reduced cell proliferation with an increase in ROS production and reduction of cyclic adenosine monophosphate (cAMP) levels in MSTO-H211, Ist-Mes1, Ist-Mes2 and MPP89 exemestane-sensitive mesothelioma cell lines, but not in NCI-H2452 exemestane-insensitive mesothelioma cells. Exemestane induced a significant antioxidant response in NCI-H2452 cells, as highlighted by an increase in γ-glutamylcysteine levels, catalase (Cat), superoxide-dismutase and (SOD) and glutathione-peroxidase (GSH-Px) activity and nuclear factor E2-related factor 2 (Nrf2) activation, responsible for drug insensitivity. Conversely, exemestane elevated ROS levels along with increased ERK phosphorylation and a reduction of p-STA3 in exemestane-sensitive mesothelioma cells. ROS generation was the crucial event of exemestane action because ROS inhibitor N-acetyl-L-cysteine (NAC) abrogated p-ERK and p-STAT3 modulation and cellular death. Exemestane also modulates ERK and STAT3 signalling via GPR30. Results indicate an essential role of ROS in the antiproliferative action of exemestane in mesothelioma cells. It is likely that the additional oxidative insults induced by exemestane results in the lethal effects of mesothelioma cells by increasing ROS production. As such, manipulating ROS levels with exemestane seems to be a feasible strategy to selectively kill mesothelioma cells with less toxicity to normal cells by regulating ERK and STAT3 activity.

  Copyright © 2017 Elsevier Inc. All rights reserved.

• Non-thermal plasma induces a stress response in mesothelioma cells resulting in increased endocytosis, lysosome biogenesis and autophagy. L. Shi, F. Ito, Y. Wang, et al. 2017;108:904-917. PubMed · Publisher

  Abstract

  Non-thermal plasma (NTP) is a potential new therapeutic modality for cancer. However, its mechanism of action remains unclear. Herein, we studied the effect of NTP on mesothelioma cells and fibroblasts to understand its anti-proliferative efficacy. Interestingly, NTP demonstrated greater selective anti-proliferative activity against mesothelioma cells relative to fibroblasts than cisplatin, which is used for mesothelioma treatment. The anti-proliferative effect of NTP was enhanced by pre-incubation with the cellular iron donor, ferric ammonium citrate (FAC), and inhibited by iron chelation using desferrioxamine (DFO). Three oxidative stress probes (CM-H2DCFDA, MitoSOX and C11-BODIPY) demonstrated reactive oxygen species (ROS) generation by NTP, which was inhibited by DFO. Moreover, NTP decreased transferrin receptor-1 and increased ferritin-H and -L chain expression that was correlated with decreased iron-regulatory protein expression and RNA-binding activity. This regulation was potentially due to increased intracellular iron in lysosomes, which was demonstrated via the Fe(II)-selective probe, HMRhoNox-M, and was consistent with autophagic-induction. Immunofluorescence using LysoTracker and Pepstatin A probes demonstrated increased cellular lysosome content, which was confirmed by elevated LAMP1 expression. The enhanced lysosomal biogenesis after NTP could be due to the observed increase in fluid-phase endocytosis and early endosome formation. These results suggest NTP acts as a stressor, which results in increased endocytosis, lysosome content and autophagy. In fact, NTP rapidly increased autophagosome formation, as judged by increased LC3B-II expression, which co-localized with LAMP1, indicating autophagolysosome formation. Autophagic-induction by NTP was confirmed using electron microscopy. In summary, NTP acts as a cellular stressor to rapidly induce fluid-phase endocytosis, lysosome biogenesis and autophagy.

  Copyright © 2017 Elsevier Inc. All rights reserved.

• Rheostatic CD44 isoform expression and its association with oxidative stress in human malignant mesothelioma. S.H. Chew, Y. Okazaki, S. Akatsuka, et al. 2017;106:91-99. PubMed · Publisher

  Abstract

  CD44 exists as a standard (CD44s) isoform and different variant isoforms (CD44v) due to alternative splicing. While the complex nature of these different isoforms has not been fully elucidated, CD44v expression has been shown to exert oncogenic effects by promoting tumor progression, metastasis and resistance of tumor cells to chemotherapy. One of the CD44v isoforms, CD44v8-10, was recently shown to protect cancer cells from oxidative stress by increasing the synthesis of glutathione (GSH). However, data regarding CD44 isoform expression in malignant mesothelioma (MM) are still lacking. Here, we show that most of the MM cell lines express both the CD44s and CD44v isoforms, in contrast to non-tumorigenic mesothelial cells, which express only CD44s. Moreover, we show here that these MM cell lines are positive for CD44 variable exon 9, with CD44v8-10 among the variant isoforms expressed. The expression of CD44 variable exon 9 was found to be statistically associated with NF2 inactivation, a common occurrence in MM. Knockdown of CD44 reduced the protein level of xCT, a cystine transporter, and increased oxidative stress. However, an increase in GSH was also observed and was associated with enhanced chemoresistance in CD44-knockdown cells. Increased GSH was mediated by the Nrf2/AP-1-induced upregulation of GCLC, a subunit of the enzyme catalyzing GSH synthesis. Our results thus suggest that the response to CD44 depletion is cell type-dependent and, in cases such as MM cells, compensatory pathway(s) might be activated rheostatically to account for the loss of CD44 and counteract enhanced oxidative stress.

  Copyright © 2017 Elsevier Inc. All rights reserved.

• Mitochondrial catalase overexpressed transgenic mice are protected against lung fibrosis in part via preventing alveolar epithelial cell mitochondrial DNA damage. S.J. Kim, P. Cheresh, R.P. Jablonski, et al. 2016;101:482-490. PubMed · Publisher · Free full text

  Abstract

  Rationale: Alveolar epithelial cell (AEC) injury and mitochondrial dysfunction are important in the development of lung fibrosis. Our group has shown that in the asbestos exposed lung, the generation of mitochondrial reactive oxygen species (ROS) in AEC mediate mitochondrial DNA (mtDNA) damage and apoptosis which are necessary for lung fibrosis. These data suggest that mitochondrial-targeted antioxidants should ameliorate asbestos-induced lung.

  Objective: To determine whether transgenic mice that express mitochondrial-targeted catalase (MCAT) have reduced lung fibrosis following exposure to asbestos or bleomycin and, if so, whether this occurs in association with reduced AEC mtDNA damage and apoptosis.

  Methods: Crocidolite asbestos (100µg/50µL), TiO2 (negative control), bleomycin (0.025 units/50µL), or PBS was instilled intratracheally in 8-10 week-old wild-type (WT - C57Bl/6J) or MCAT mice. The lungs were harvested at 21d. Lung fibrosis was quantified by collagen levels (Sircol) and lung fibrosis scores. AEC apoptosis was assessed by cleaved caspase-3 (CC-3)/Surfactant protein C (SFTPC) immunohistochemistry (IHC) and semi-quantitative analysis. AEC (primary AT2 cells from WT and MCAT mice and MLE-12 cells) mtDNA damage was assessed by a quantitative PCR-based assay, apoptosis was assessed by DNA fragmentation, and ROS production was assessed by a Mito-Sox assay.

  Results: Compared to WT, crocidolite-exposed MCAT mice exhibit reduced pulmonary fibrosis as measured by lung collagen levels and lung fibrosis score. The protective effects in MCAT mice were accompanied by reduced AEC mtDNA damage and apoptosis. Similar findings were noted following bleomycin exposure. Euk-134, a mitochondrial SOD/catalase mimetic, attenuated MLE-12 cell DNA damage and apoptosis. Finally, compared to WT, asbestos-induced MCAT AT2 cell ROS production was reduced.

  Conclusions: Our finding that MCAT mice have reduced pulmonary fibrosis, AEC mtDNA damage and apoptosis following exposure to asbestos or bleomycin suggests an important role for AEC mitochondrial H2O2-induced mtDNA damage in promoting lung fibrosis. We reason that strategies aimed at limiting AEC mtDNA damage arising from excess mitochondrial H2O2 production may be a novel therapeutic target for mitigating pulmonary fibrosis.

  Published by Elsevier Inc.

• Malignant mesothelioma as an oxidative stress-induced cancer: An update. S.H. Chew, S. Toyokuni. 2015;86:166-78. Review PubMed · Publisher

  Abstract

  Malignant mesothelioma (MM) is a relatively rare cancer that occurs almost exclusively following respiratory exposure to asbestos in humans. Its pathogenesis is closely associated with iron overload and oxidative stress in mesothelial cells. On fiber exposure, mesothelial cells accumulate fibers simultaneously with iron, which either performs physical scissor function or catalyzes free radical generation, leading to oxidative DNA damage such as strand breaks and base modifications, followed by activation of intracellular signaling pathways. Chrysotile, per se without iron, causes massive hemolysis and further adsorbs hemoglobin. Exposure to indigestible foreign materials also induces chronic inflammation, involving consistent generation of free radicals and subsequent activation of NALP3 inflammasomes in macrophages. All of these contribute to mesothelial carcinogenesis. Genomic alterations most frequently involve homozygous deletion of INK4A/4B, and other pathways such as Hippo and TGF-β pathways are also affected in MM. Recently, analyses of familial MM sorted out BAP1 as a novel responsible tumor suppressor gene, whose function is not fully elucidated. Five-year survival of mesothelioma is still ~8%, and this cancer is increasing worldwide. Connective tissue growth factor, a secretory protein creating a vicious cycle mediated by β-catenin, has been recognized as a hopeful target for therapy, especially in sarcomatoid subtype. Recent research outcomes related to microRNAs and cancer stem cells also offer additional novel targets for the treatment of MM. Iron reduction as chemoprevention of mesothelioma is helpful at least in an animal preclinical study. Integrated approaches to fiber-induced oxidative stress would be necessary to overcome this currently fatal disease.

  Copyright © 2015 Elsevier Inc. All rights reserved.

• AKT/mTOR and c-Jun N-terminal kinase signaling pathways are required for chrysotile asbestos-induced autophagy. Z. Lin, T. Liu, D.W. Kamp, et al. 2014;72:296-307. PubMed · Publisher · Free full text

  Abstract

  Chrysotile asbestos is closely associated with excess mortality from pulmonary diseases such as lung cancer, mesothelioma, and asbestosis. Although multiple mechanisms in which chrysotile asbestos fibers induce pulmonary disease have been identified, the role of autophagy in human lung epithelial cells has not been examined. In this study, we evaluated whether chrysotile asbestos induces autophagy in A549 human lung epithelial cells and then analyzed the possible underlying molecular mechanism. Chrysotile asbestos induced autophagy in A549 cells based on a series of biochemical and microscopic autophagy markers. We observed that asbestos increased expression of A549 cell microtubule-associated protein 1 light chain 3 (LC3-II), an autophagy marker, in conjunction with dephosphorylation of phospho-AKT, phospho-mTOR, and phospho-p70S6K. Notably, AKT1/AKT2 double-knockout murine embryonic fibroblasts (MEFs) had negligible asbestos-induced LC3-II expression, supporting a crucial role for AKT signaling. Chrysotile asbestos also led to the phosphorylation/activation of Jun N-terminal kinase (JNK) and p38 MAPK. Pharmacologic inhibition of JNK, but not p38 MAPK, dramatically inhibited the protein expression of LC3-II. Moreover, JNK2(-/-) MEFs but not JNK1(-/-) MEFs blocked LC3-II levels induced by chrysotile asbestos. In addition, N-acetylcysteine, an antioxidant, attenuated chrysotile asbestos-induced dephosphorylation of P-AKT and completely abolished phosphorylation/activation of JNK. Finally, we demonstrated that chrysotile asbestos-induced apoptosis was not affected by the presence of the autophagy inhibitor 3-methyladenine or autophagy-related gene 5 siRNA, indicating that the chrysotile asbestos-induced autophagy may be adaptive rather than prosurvival. Our findings demonstrate that AKT/mTOR and JNK2 signaling pathways are required for chrysotile asbestos-induced autophagy. These data provide a mechanistic basis for possible future clinical applications targeting these signaling pathways in the management of asbestos-induced lung disease.

  Copyright © 2014 Elsevier Inc. All rights reserved.

• Indoleamine-2,3-dioxygenase elevated in tumor-initiating cells is suppressed by mitocans. M. Stapelberg, R. Zobalova, M.N. Nguyen, et al. 2014;67:41-50. PubMed · Publisher

  Abstract

  Tumor-initiating cells (TICs) often survive therapy and give rise to second-line tumors. We tested the plausibility of sphere cultures as models of TICs. Microarray data and microRNA data analysis confirmed the validity of spheres as models of TICs for breast and prostate cancer as well as mesothelioma cell lines. Microarray data analysis revealed the Trp pathway as the only pathway upregulated significantly in all types of studied TICs, with increased levels of indoleamine-2,3-dioxygenase-1 (IDO1), the rate-limiting enzyme of Trp metabolism along the kynurenine pathway. All types of TICs also expressed higher levels of the Trp uptake system consisting of CD98 and LAT1 with functional consequences. IDO1 expression was regulated via both transcriptional and posttranscriptional mechanisms, depending on the cancer type. Serial transplantation of TICs in mice resulted in gradually increased IDO1. Mitocans, represented by α-tocopheryl succinate and mitochondrially targeted vitamin E succinate (MitoVES), suppressed IDO1 in TICs. MitoVES suppressed IDO1 in TICs with functional mitochondrial complex II, involving transcriptional and posttranscriptional mechanisms. IDO1 increase and its suppression by VE analogues were replicated in TICs from primary human glioblastomas. Our work indicates that IDO1 is increased in TICs and that mitocans suppress the protein.

  © 2013 Elsevier Inc. All rights reserved.

• Allele-specific effects of ecSOD on asbestos-induced fibroproliferative lung disease in mice. S. Jun, C.L. Fattman, B.J. Kim, et al. 2011;50(10):1288-96. PubMed · Publisher · Free full text

  Abstract

  Previous work by others suggests that there is a strain-dependent variation in the susceptibility to inflammatory lung injury in mice. Specifically, the 129/J mice appear to be more resistant to asbestos-induced pulmonary fibrosis than the C57BL/6 strain. A separate line of evidence suggests that extracellular superoxide dismutase (ecSOD) may play an important role in protecting the lung from such injuries. We have recently reported that the 129/J strain of mice has an ecSOD genotype and phenotype distinctly different from those of the C57BL/6 mice. In order to identify ecSOD as a potential "asbestos-injury resistance" gene, we bred congenic mice, on the C57BL/6 background, carrying the wild type (sod3wt) or the 129/J (sod3129) allele for ecSOD. This allowed us to examine the role of ecSOD polymorphism in susceptibility to lung injury in an otherwise identical genetic background. Interestingly, asbestos treatment induces a significant (~40%) increase in plasma ecSOD activity in the sod3129 mice, but not in the sod3wt mice. Asbestos administration results in a loss of ecSOD activity and protein from lung tissue of both congenic strains, but the lung ecSOD activity remains significantly higher in sod3129 mice. As expected, asbestos treatment results in a significant recovery of ecSOD protein in bronchoalveolar lavage fluid (BALF). The BALF of sod3129 mice also have significantly lower levels of proteins and inflammatory cells, especially neutrophils, accompanied by a significantly lower extent of lung injury, as measured by a pathology index score or hydroxyproline content. Immunohistochemistry reveals a significant loss of ecSOD from the tips of the respiratory epithelial cells in response to asbestos treatment and that the loss of immunodetectable ecSOD is compensated for by enzyme expression by infiltrating cells, especially in the sod3wt mice. Our studies thus identify ecSOD as an important anti-inflammatory gene, responsible for most, if not all of the resistance to asbestos-induced lung injury reported for the 129/J strain of mice. The data further suggest allele-specific differences in the regulation of ecSOD expression. These congenic mice therefore represent a very useful model to study the role of this enzyme in all inflammatory diseases. Polymorphisms in human ecSOD have also been reported and it appears logical to assume that such variations may have a profound effect on disease susceptibility.

  Copyright © 2011 Elsevier Inc. All rights reserved.

2000s

• Beta-carotene metabolites enhance inflammation-induced oxidative DNA damage in lung epithelial cells. Y.G. van Helden, J. Keijer, A.M. Knaapen, et al. 2009;46(2):299-304. PubMed · Publisher

  Abstract

  beta-Carotene (BC) intake has been shown to enhance lung cancer risk in smokers and asbestos-exposed subjects (according to the ATBC and CARET studies), but the mechanism behind this procarcinogenic effect of BC is unclear. Both smoking and asbestos exposure induce an influx of inflammatory neutrophils into the airways, which results in an increased production of reactive oxygen species and formation of promutagenic DNA lesions. Therefore, the aim of our study was to investigate the effects of BC and its metabolites (BCM) on neutrophil-induced genotoxicity. We observed that the BCM vitamin A (Vit A) and retinoic acid (RA) inhibited the H(2)O(2)-utilizing enzyme myeloperoxidase (MPO), which is released by neutrophils, thereby reducing H(2)O(2) conversion. Moreover, BC and BCM were able to increase (.)OH formation from H(2)O(2) in the Fenton reaction (determined by electron spin resonance spectroscopy). Addition of Vit A and RA to lung epithelial cells that were co-incubated with activated neutrophils resulted in a significant increase in the level of oxidized purines assessed by the formamidopyrimidine DNA glycosylase-modified comet assay. These data indicate that BCM can enhance neutrophil-induced genotoxicity by inhibition of MPO in combination with subsequent increased formation of hydroxyl radicals.

• Increased sensitivity to asbestos-induced lung injury in mice lacking extracellular superoxide dismutase. C.L. Fattman, R.J. Tan, J.M. Tobolewski, et al. 2006;40(4):601-7. PubMed · Publisher · Free full text

  Abstract

  Asbestosis is a chronic form of interstitial lung disease characterized by inflammation and fibrosis that results from the inhalation of asbestos fibers. Although the pathogenesis of asbestosis is poorly understood, reactive oxygen species may mediate the progression of this disease. The antioxidant enzyme extracellular superoxide dismutase (EC-SOD) can protect the lung against a variety of insults; however, its role in asbestosis is unknown. To determine if EC-SOD plays a direct role in protecting the lung from asbestos-induced injury, intratracheal injections of crocidolite were given to wild-type and ec-sod-null mice. Bronchoalveolar lavage fluid (BALF) from asbestos-treated ec-sod-null mice at 24 h, 14 days, or 28 days posttreatment showed increased inflammation and total BALF protein content compared to that of wild-type mice. In addition, lungs from ec-sod-null mice showed increased hydroxyproline content compared to those of wild-type mice, indicating a greater fibrotic response. Finally, lungs from ec-sod-null mice showed greater oxidative damage, as assessed by nitrotyrosine content compared to those of their wild-type counterparts. These results indicate that depletion of EC-SOD from the lung increases oxidative stress and injury in response to asbestos.

• Interactions of exogenous or evoked agents and particles: the role of reactive oxygen species. A. Churg. 2003;34(10):1230-5. Review PubMed · Publisher

  Abstract

  Humans are commonly exposed to combinations of particles (occupational or environmental) and exogenous agents such as ozone and cigarette smoke that generate reactive oxygen species (ROS). Particles also evoke production of ROS from inflammatory cells and of mediators such as TNF-alpha that operate through ROS-related mechanisms. The interactions of particles and ROS-generating agents have been little explored. Adhesion of particles to the surface of pulmonary epithelial cells is increased by exposure to cigarette smoke, ozone, and TNF-alpha. Cigarette smoke and ozone increase the uptake of particles by epithelial cells, and both adhesion and uptake can be decreased by scavengers of ROS. Increased adhesion and uptake probably lead to increased levels of inflammatory and fibrogenic mediator production, and cigarette smoke definitely increases whole lung particle retention and enhances the fibrogenic effects of asbestos. In experimental models, the combination of particles plus ozone, cigarette smoke, or reagent hydrogen peroxide augments the inflammatory response to particles, increases cell proliferation, and leads to liberation of increased levels of chemoattractant mediators as well as vascular mediators such as endothelin. The small airways appear to be particular targets of coexposure to smoke or ozone and particles, a phenomenon that may produce chronic airflow obstruction.

• Multiple roles of oxidants in the pathogenesis of asbestos-induced diseases. A. Shukla, M. Gulumian, T.K. Hei, et al. 2003;34(9):1117-29. Review PubMed · Publisher

  Abstract

  Exposure to asbestos causes cellular damage, leading to asbestosis, bronchogenic carcinoma, and mesothelioma in humans. The pathogenesis of asbestos-related diseases is complicated and still poorly understood. Studies on animal models and cell cultures have indicated that asbestos fibers generate reactive oxygen and nitrogen species (ROS/RNS) and cause oxidation and/or nitrosylation of proteins and DNA. The ionic state of iron and its ability to be mobilized determine the oxidant-inducing potential of pathogenic iron-containing asbestos types. In addition to their capacity to damage macromolecules, oxidants play important roles in the initiation of numerous signal transduction pathways that are linked to apoptosis, inflammation, and proliferation. There is strong evidence supporting the premise that oxidants contribute to asbestos-induced lung injury; thus, strategies for reducing oxidant stress to pulmonary cells may attenuate the deleterious effects of asbestos.

• Crocidolite asbestos inhibits pentose phosphate oxidative pathway and glucose 6-phosphate dehydrogenase activity in human lung epithelial cells. C. Riganti, E. Aldieri, L. Bergandi, et al. 2002;32(9):938-49. PubMed · Publisher

  Abstract

  The cytotoxicity of asbestos has been related to its ability to increase the production of reactive oxygen species (ROS), via the iron-catalyzed reduction of oxygen and/or the activation of NADPH oxidase. The pentose phosphate pathway (PPP) is generally activated by the cell exposure to oxidant molecules. Contrary to our expectations, asbestos (crocidolite) fibers caused a dose- and time-dependent inhibition of PPP and decreased its activation by an oxidative stress in human lung epithelial cells A549. In parallel, the intracellular activity of the PPP rate-limiting enzyme, glucose 6-phosphate dehydrogenase (G6PD), was significantly diminished by crocidolite exposure. This inhibition was selective, as the activity of other PPP and glycolysis enzymes was not modified, and was not attributable to a decreased expression of G6PD. On the opposite, the incubation with glass fibers MMVF10 did not modify PPP and G6PD activity. PPP and G6PD inhibition did not correlate with the increased nitric oxide (NO) production elicited by crocidolite in A549 cells. Experiments with the purified enzyme suggest that crocidolite inhibits G6PD by directly interacting with the protein. We propose here a new mechanism of asbestos-evoked oxidative stress, wherein fibers increase the intracellular ROS levels also by inhibiting the main antioxidant pathway of the cell.

• Dual role of inducible nitric oxide synthase in acute asbestos-induced lung injury. M. Dörger, A.M. Allmeling, R. Kiefmann, et al. 2002;33(4):491-501. PubMed · Publisher

  Abstract

  Reactive oxygen and nitrogen species have been implicated in the pathogenesis of asbestos fibers-associated pulmonary diseases. By comparing the responses of inducible nitric oxide synthase (iNOS) knockout and wild-type mice we investigated the consequences of iNOS expression for the development of the inflammatory response and tissue injury upon intratracheal instillation of asbestos fibers. Exposure to asbestos fibers resulted in an increased iNOS mRNA and protein expression in the lungs from wild-type mice. Moreover, iNOS knockout mice exhibited an exceeded pulmonary expression and production of TNF-alpha as well as a higher influx of neutrophils into the alveolar space than wild-type mice. In contrast, iNOS knockout animals displayed an attenuated oxidant-related tissue injury reflected in a decrease in protein leakage and LDH release into the alveolar space as well as weaker nitrotyrosine staining of lung tissue compared to wild-type mice. Data presented here indicate that iNOS-derived NO exerts a dichotomous role in acute asbestos-induced lung injury in that iNOS deficiency resulted in an exacerbated inflammatory response but improved oxidant-promoted lung tissue damage.

• Particle-induced oxidative damage is ameliorated by pulmonary antioxidants. L.L. Greenwell, T. Moreno, T.P. Jones, et al. 2002;32(9):898-905. PubMed · Publisher

  Abstract

  This investigation focuses on the application of an in vitro assay in elucidating the role of lung lining fluid antioxidants in the protection against inhaled particles, and to investigate the source of bioreactivity in urban PM10 collections from South Wales. The Plasmid Assay is an in vitro method of assessing and comparing the oxidative bioreactivity of inhalable particles. This method has provided the basis of limited toxicological studies into various inhaled xenobiotics including asbestos, and more recently PM10. Carbon Black M120 and Diesel Exhaust Particles (DEP) were tested as PM10 surrogates, DEP displaying the greatest oxidative bioreactivity. Both urban PM2.5 (fine fraction) and PM2.5-10 (coarse fraction) (Cardiff, S. Wales, UK) caused significant damage, the coarse fraction displaying higher oxidative capacity. The soluble components were found to be responsible for most of the bioreactivity in both PM sizes. Low molecular components of fresh lung lavage were found to offer most antioxidant protection, and surrogate Epithelial Lining Fluid (sELF) showed significant amelioration of DNA damage by the coarse fraction but less effect against the fine. Overall, the coarse, soluble fraction of PM10 is a great source of oxidative bioreactivity, but natural pulmonary low molecular weight antioxidants can significantly ameliorate its effects.

• Iron inhibits the nitric oxide synthesis elicited by asbestos in murine macrophages. E. Aldieri, D. Ghigo, M. Tomatis, et al. 2001;31(3):412-7. PubMed · Publisher

  Abstract

  Crocidolite fibers stimulated nitric oxide synthase (NOS) activity and expression in glial and alveolar murine macrophages: this effect was inhibited by iron supplementation and enhanced by iron chelation. We suggest that in these cells crocidolite stimulates NOS expression by decreasing the iron bioavailability and activating an iron-sensitive transcription factor.

• Asbestos and cigarette smoke cause increased DNA strand breaks and necrosis in bronchiolar epithelial cells in vivo. M. Jung, W.P. Davis, D.J. Taatjes, et al. 2000;28(8):1295-9. PubMed · Publisher

  Abstract

  Coexposures to asbestos and cigarette smoke cause increased risks of lung cancer in asbestos workers. Although these carcinogens cause DNA damage to epithelial cells in vitro via generation of reactive oxygen species (ROS), it is unclear whether they cause injury to bronchiolar epithelial cells (i.e., the target cells of lung cancers in vivo). We exposed rats to amosite asbestos, cigarette smoke, and the two agents in combination for 1, 2, and 14 d. Numbers of cells exhibiting DNA strand breaks in comparison to sham rats were then evaluated in lungs using the terminal deoxynucleotidyl transferase (TDT)-mediated dUTP-biotin nick end labeling (TUNEL) method and by transmission electron microscopy (TEM). Increases in TUNEL-positive, necrotic epithelial cells occurred after exposure to asbestos alone and in an additive fashion after smoke and asbestos in combination. These results indicate that DNA strand breakage and necrosis are prominent mechanisms of injury by asbestos fibers and cigarette smoke in vivo to epithelial cells of the respiratory tract, thus validating in vitro observations from a number of laboratories.

1990s

• Cigarette smoke augments asbestos-induced alveolar epithelial cell injury: role of free radicals. D.W. Kamp, M.J. Greenberger, J.S. Sbalchierro, et al. 1998;25(6):728-39. PubMed · Publisher

  Abstract

  Cigarette smoke augments asbestos-induced bronchogenic carcinoma by mechanisms that are not established. Alveolar epithelial cell (AEC) injury due to oxidant-induced DNA damage and depletion of glutathione (GSH) and adenosine triphosphate (ATP) may be one important mechanism. We previously showed that amosite asbestos-induces hydroxyl radical production and DNA damage to cultured AEC and that phytic acid, an iron chelator, is protective. We hypothesized that whole cigarette smoke extracts (CSE) augment amosite asbestos-induced AEC injury by generating iron-induced free radicals that damage DNA and reduce cellular GSH and ATP levels. Asbestos or CSE each caused dose-dependent toxicity to AEC (WI-26 and rat alveolar type I-like cells) as assessed by 51chromium release. The combination of asbestos (5 microg/cm2) and CSE (0.O1-0.1%) caused synergistic injury whereas higher doses of each agent primarily had an additive toxic effect. Asbestos (5 microg/cm2) augmented CSE-induced (0.01-1.0%) AEC DNA damage over a 4 h exposure period as assessed by an alkaline unwinding, ethidium bromide fluorometric technique. These effects were synergistic in A549 cells and additive in WI-26 cells. Asbestos (5 microg/cm2) and CSE (0.5-1.0%) reduced A549 and WI-26 cell GSH levels as assessed spectrophotometrically and ATP levels as assessed by luciferin/luciferase chemiluminescence but a synergistic interaction was not detected. Phytic acid (500 microM) and catalase (100 microg/ml) each attenuated A549 cell DNA damage and depletion of ATP caused by asbestos and CSE. However, neither agent attenuated WI-26 cell DNA damage nor the reductions in GSH levels in WI-26 and A549 cells exposed to asbestos and CSE. We conclude that CSE enhance asbestos-induced DNA damage in cultured alveolar epithelial cells. These data provide additional support that asbestos and cigarette smoke are genotoxic to relevant target cells in the lung and that iron-induced free radicals may in part cause these effects.

• GRP78, HSP72/73, and cJun stress protein levels in lung epithelial cells exposed to asbestos, cadmium, or H2O2. C.R. Timblin, Y.M. Janssen, J.L. Goldberg, et al. 1998;24(4):632-42. PubMed · Publisher

  Abstract

  Occupational exposure to crocidolite asbestos is associated with the development of nonmalignant and malignant pulmonary disease. Considerable evidence indicates that the mechanisms of asbestos-induced toxicity involve the production of active oxygen species (AOS). Production of AOS in excess of cellular defenses creates an environment of oxidative stress and stimulates the expression of a number of different genes whose products may be involved in mediating responses from oxidant injury. To further investigate the mechanisms of asbestos-induced pathogenicity, we have examined by Western blot analyses the induction of the stress response proteins GRP78 and HSP72/73 in rat lung epithelial cells (RLE) exposed to crocidolite asbestos. In comparative studies, we also examined GRP78, HSP72/73, and cJun expression in RLE cells exposed to equitoxic concentrations of cadmium chloride (CdCl2) and hydrogen peroxide (H2O2). Our results demonstrate that asbestos and H2O2 do not alter GRP78 or HSP72/73 protein levels in RLE cells, but do increase levels of cJun protein. Increases by asbestos and H2O2 were not accompanied by alterations in cellular glutathione levels in this cell type, but asbestos caused elevations in protein levels of manganese-containing superoxide dismutase (MnSOD), an indirect indicator of oxidant stress. In contrast, exposure of cells to CdCl2 led to no changes in MnSOD protein levels, but increases in GRP78, HSP72/73, and cJun proteins as well as significant increases in oxidized and reduced thiol pools. Results suggest that environmental agents causing oxidative injury to lung epithelium elicit different patterns of stress responses.

• In vivo evidence of free radical formation after asbestos instillation: an ESR spin trapping investigation. A.J. Ghio, M.B. Kadiiska, Q.H. Xiang, et al. 1998;24(1):11-7. PubMed · Publisher

  Abstract

  It has been postulated that the in vivo toxicity of asbestos results from its catalysis of free radical generation. We examined in vivo radical production using electron spin resonance (ESR) coupled with the spin trap alpha-(4-pyridyl-1-oxide)-N-t-butylnitrone (4-POBN); 180 day-old rats were intratracheally instilled with either 500 microg crocidolite or saline. Twenty-four hours later, histologic examination revealed a neutrophilic inflammatory response. ESR spectroscopy of the chloroform extract from lungs exposed to asbestos gave a spectrum consistent with a carbon-centered radical adduct, while those spectra from lungs instilled with saline revealed a much weaker signal. This same radical formation persisted and, even one month after instillation, could be detected in the lungs of rats exposed to asbestos. The 4-POBN adducts detected by ESR are very similar to, if not identical with, ethyl and pentyl radical adducts, providing evidence of in vivo lipid peroxidation resulting from asbestos exposure. We conclude that, after instillation of crocidolite in the rat, ESR analysis of lung tissue demonstrates in vivo free radical production.

• Mechanisms of asbestos-induced nitric oxide production by rat alveolar macrophages in inhalation and in vitro models. T.R. Quinlan, K.A. BeruBe, M.P. Hacker, et al. 1998;24(5):778-88. PubMed · Publisher

  Abstract

  To evaluate the contribution of reactive nitrogen species to inflammation by asbestos, Fischer 344 rats were exposed to crocidolite or chrysotile asbestos by inhalation to determine whether increases occurred in nitric oxide (NO.) metabolites from alveolar macrophages (AMs). AMs from animals inhaling asbestos showed significant elevations (p < .05) in nitrite/nitrate levels which were ameliorated by NG-monomethyl-L-arginine (NMMA), an inhibitor of inducible nitric oxide synthase (iNOS) activity. Temporal patterns of NO. generation from AMs correlated with neutrophil influx in bronchoalveolar lavage samples after asbestos inhalation or bleomycin instillation, another model of pulmonary fibrosis. To determine the molecular mechanisms and specificity of iNOS promoter activation by asbestos, RAW 264.7 cells, a murine macrophage-like line, and AMs isolated from control rats were exposed to crocidolite asbestos in vitro. These cells showed increases in steady-state levels of iNOS mRNA in response to asbestos and more dramatic increases in both iNOS mRNA and immunoreactive protein after addition of lipopolysaccharide (LPS). After transfection of an iNOS promoter/luciferase reporter construct, RAW 264.7 cells exposed to LPS, crocidolite asbestos and its nonfibrous analog, riebeckite, revealed increases in luciferase activity whereas cristobalite silica had no effects. Studies suggest that NO. generation may be important in cell injury and inflammation by asbestos.

• A study of the synergistic interaction of asbestos fibers with cigarette tar extracts for the generation of hydroxyl radicals in aqueous buffer solution. A. Valavanidis, H. Balomenou, I. Macropoulou, et al. 1996;20(6):853-8. PubMed · Publisher

  Abstract

  Several models attempt to explain the synergistic increase in lung cancer among workers exposed to asbestos fibers, who were smokers at the same time. It is known that reactive oxygen species (ROS) are important mediators in asbestos-induced diseases, especially cancer. We studied quantitatively the formation of ROS (by spin trapping with DMPO) in aqueous buffer suspensions containing crocidolite (UICC), chrysotile (UICC and commercial, long fibers) alone, and in combination with aqueous cigarette tar extracts. It was observed that asbestos and cigarette tar act in a cooperative or synergistic way in the generation of hydroxyl radical spin adducts. Grinding of asbestos fibers and addition of EDTA (iron chelator) enhanced the intensity of the ESR signal. This enhancement progressed with time, probably due to the reaction of the extracted iron with the slow released hydrogen peroxide from tar extracts. It was observed a fivefold increase in the ESR signal (for crocidolite and aqueous tar extracts) in the formation of hydroxyl radicals via an iron-catalyzed Fenton reaction. These experimental results are suggest to be strong evidence to the fact that lung cancer has been found in asbestos workers exposed to high concentrations of fibers in the working environment who were smokers, and only rarely in nonsmokers.

• Manganese superoxide dismutase in human pleural mesothelioma cell lines. V.L. Kinnula, P. Pietarinen-Runtti, K. Raivio, et al. 1996;21(4):527-32. PubMed · Publisher

  Abstract

  Mesothelioma is a malignant pleural or intraperitoneal tumor attributable to asbestos exposure in more than 80% of the cases. Manganese superoxide dismutase (MnSOD), a mitochondrial superoxide radical scavenging enzyme, is low in most tumors but is known to be induced by asbestos fibers and certain cytokines. Induction of MnSOD may be associated in asbestos-related pulmonary diseases in vivo. We investigated here MnSOD specific activity and MnSOD mRNA level using healthy human lung tissue, SV40-transformed human pleural mesothelial cells (Met5A), and six human malignant mesothelioma cell line cells. Total SOD (CuZnSOD + MnSOD) and MnSOD activities were 20.0 +/- 4.8 U/mg protein and 3.2 +/- 1.2 U/mg protein in healthy human lung tissue, and 25.6 +/- 10.7 U/mg and 3.8 +/- 1.0 U/mg in Met5A cells, respectively. In four mesothelioma cell lines MnSOD activity was significantly elevated, the highest activity (30.1 +/- 8.2 U/mg) was almost 10-fold compared to the activity in Met5A cells. The steady state mRNA level of MnSOD was low in Met5A cells and markedly higher in all mesothelioma cell lines roughly in proportion with enzyme activities. Cytotoxicity experiments, which were conducted in four cell lines, indicated that cells containing high MnSOD mRNA level and activity were resistant to the mitochondrial superoxide-producing agent menadione. In conclusion, our results suggest that human mesothelioma may express high levels of MnSOD, which is associated with high oxidant resistance of these cells.

• Protective effect of natural flavonoids on rat peritoneal macrophages injury caused by asbestos fibers. V.A. Kostyuk, A.I. Potapovich, S.D. Speransky, et al. 1996;21(4):487-93. PubMed · Publisher

  Abstract

  Exposure of macrophages to asbestos fibers resulted in enhancement of the production of oxygen radicals, determined by a lucigenin enhanced chemiluminescence (LEC) assay, a formation of thiobarbituric acid reactive substances (TBARS), a LDH release into the incubation mixture, and a rapid lysis of the cells. Rutin (Rut) and quercetin (Qr) were effective in inhibiting LEC, TBARS formation, and reducing peritoneal macrophages injury caused by asbestos. The concentrations pre-treatment of antioxidants that were required to prevent the injury of peritoneal macrophages caused by asbestos by 50% (IC50) were 90 microM and 290 microM for Qr and Rut, respectively. Both flavonoids were found to be oxidized during exposure of peritoneal macrophages to asbestos and the oxidation was SOD sensitive. The efficacy of flavonoids as antioxidant agents as well as superoxide ion scavengers was also evaluated using appropriate model systems, and both quercetin and rutin were found to be effective in scavenging O2.-. These findings indicate that flavonoids are able to prevent the respiratory burst in rat peritoneal macrophages exposed to asbestos at the stage of activated oxygen species generation, mainly as superoxide scavengers. On the basis of this study it was concluded that natural flavonoids quercetin and rutin would be promising drug candidates for a prophylactic asbestos-induced disease.

• Transfection of a manganese-containing superoxide dismutase gene into hamster tracheal epithelial cells ameliorates asbestos-mediated cytotoxicity. B.T. Mossman, P. Surinrut, B.T. Brinton, et al. 1996;21(2):125-31. PubMed · Publisher

  Abstract

  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.

• Neutrophil and asbestos fiber-induced cytotoxicity in cultured human mesothelial and bronchial epithelial cells. V.L. Kinnula, K.O. Raivio, K. Linnainmaa, et al. 1995;18(3):391-9. PubMed · Publisher

  Abstract

  This study investigates reactive oxygen species generation and oxidant-related cytotoxicity induced by amosite asbestos fibers and polymorphonuclear leucocytes (PMNs) in human mesothelial cells and human bronchial epithelial cells in vitro. Transformed human pleural mesothelial cells (MET 5A) and bronchial epithelial cells (BEAS 2B) were treated with amosite (2 micrograms/cm2) for 48 h. After 24 h of incubation, the cells were exposed for 1 h to nonactivated or amosite (50 micrograms) activated PMNs, washed, and incubated for another 23 h. Reactive oxygen species generation by the PMNs and the target cells was measured by chemiluminescence. Cell injury was assessed by cellular adenine nucleotide depletion, extracellular release of nucleotides, and lactate dehydrogenase (LDH). Amosite-activated (but also to a lesser degree nonactivated) PMNs released substantial amounts of reactive oxygen metabolites, whereas the chemiluminescence of amosite-exposed mesothelial cells and epithelial cells did not differ from the background. Amosite treatment (48 h) of the target cells did not change intracellular adenine nucleotides (ATP, ADP, AMP) or nucleotide catabolite products (xanthine, hypoxanthine, and uric acid). When the target cells were exposed to nonactivated PMNs, significant adenine nucleotide depletion and nucleotide catabolite accumulation was observed in mesothelial cells only. In separate experiments, when the target cells were exposed to amosite-activated PMNs, the target cell injury was further potentiated compared with the amosite treatment alone or exposure to nonactivated PMNs. In conclusion, this study suggests the importance of inflammatory cell-derived free radicals in the development of amosite-induced mesothelial cell injury.

• Cytotoxicity of oxidants and asbestos fibers in cultured human mesothelial cells. V.L. Kinnula, K. Aalto, K.O. Raivio, et al. 1994;16(2):169-76. PubMed · Publisher

  Abstract

  The authors investigated the mechanisms caused by oxidants (superoxide and hydrogen peroxide) and asbestos (amosite) fibers in human mesothelial cells. Immortalized human pleural mesothelial cells (MET 5A) were exposed in vitro to one of the following: hypoxanthine (100-200 microM) plus xanthine oxidase (10-20 mU/ml) as a superoxide-generating system, H2O2 (50 microM-5 mM); or amosite (1-100 micrograms/cm2). Cellular adenine nucleotide depletion, DNA single strand breaks, extracellular release of nucleotides, and their catabolites and lactate dehydrogenase (LDH) were assessed as markers of cell damage after 4-6 h exposure to the oxidants or fibers. The effect of intracellular antioxidant enzymes and exogenous antioxidants on cell damage were investigated during oxidant and amosite exposure. Superoxide radical and H2O2 exposure resulted in the depletion of adenine nucleotides, accumulation of the products of nucleotide catabolism, induction of DNA single strand breaks, and extracellular LDH release. Amosite exposure did not cause nucleotide depletion or induction of DNA single strand breaks. Inactivation of the intracellular antioxidant enzymes glutathione reductase or catalase augmented cell damage during H2O2 exposure but not during amosite exposure.

• Reactive oxygen species, chromosome mutation, and cancer: possible role of clastogenic factors in carcinogenesis. I. Emerit. 1994;16(1):99-109. Review PubMed · Publisher

  Abstract

  Superoxide radicals may induce genotoxic effects by indirect action mechanisms, implicating the formation of more long-lived, secondary clastogenic material called chromosome breakage factors or clastogenic factors (CF). CF are produced via the intermediacy of superoxide, and stimulate further superoxide production by competent cells. This results in a selfsustaining and longlasting process of clastogenesis, which may exceed the DNA repair system and ultimately lead to cancer. An increased cancer risk is indeed observed in conditions accompanied by CF formation. These include irradiated persons, asbestos workers, patients with chronic inflammatory diseases, HIV-infected persons, and the congenital breakage syndromes ataxia telangiectasia, Bloom's syndrome, and Fanconi's anemia. Because reactive oxygen species (ROS) are implicated in CF formation and CF action, antioxidants may be protective as anticlastogens and consequently as anticarcinogens. In persons at high risk because of their occupation, life style or place of residence, the presence of CF may represent an indication for chemoprevention of cancer by antioxidants. CF can be useful as biochemical markers and intermediate endpoints for the evaluation of promising drugs. They are therefore not only of interest as a mechanism by which ROS may exert genotoxic effects, but also have practical implications.

• The role of free radicals in asbestos-induced diseases. D.W. Kamp, P. Graceffa, W.A. Pryor, et al. 1992;12(4):293-315. Review PubMed · Publisher

  Abstract

  Asbestos exposure causes pulmonary fibrosis and malignant neoplasms by mechanisms that remain uncertain. In this review, we explore the evidence supporting the hypothesis that free radicals and other reactive oxygen species (ROS) are an important mechanism by which asbestos mediates tissue damage. There appears to be at least two principal mechanisms by which asbestos can induce ROS production; one operates in cell-free systems and the other involves mediation by phagocytic cells. Asbestos and other synthetic mineral fibers can generate free radicals in cell-free systems containing atmospheric oxygen. In particular, the hydroxyl radical often appears to be involved, and the iron content of the fibers has an important role in the generation of this reactive radical. However, asbestos also appears to catalyze electron transfer reactions that do not require iron. Iron chelators either inhibit or augment asbestos-catalyzed generation of the hydroxyl radical and/or pathological changes, depending on the chelator and the nature of the asbestos sample used. The second principal mechanism for asbestos-induced ROS generation involves the activation of phagocytic cells. A variety of mineral fibers have been shown to augment the release of reactive oxygen intermediates from phagocytic cells such as neutrophils and alveolar macrophages. The molecular mechanisms involved are unclear but may involve incomplete phagocytosis with subsequent oxidant release, stimulation of the phospholipase C pathway, and/or IgG-fragment receptor activation. Reactive oxygen species are important mediators of asbestos-induced toxicity to a number of pulmonary cells including alveolar macrophages, epithelial cells, mesothelial cells, and endothelial cells. Reactive oxygen species may contribute to the well-known synergistic effects of asbestos and cigarette smoke on the lung, and the reasons for this synergy are discussed. We conclude that there is strong evidence supporting the premise that reactive oxygen species and/or free radicals contribute to asbestos-induced and cigarette smoke/asbestos-induced lung injury and that strategies aimed at reducing the oxidant stress on pulmonary cells may attenuate the deleterious effects of asbestos.

• Enhanced lipid peroxidation in lung lavage of rats after inhalation of asbestos. J.M. Petruska, K.O. Leslie, B.T. Mossman. 1991;11(4):425-32. PubMed · Publisher

  Abstract

  Exposure of phagocytic cells to asbestos in vitro results in an augmented production of reactive oxygen metabolites and increased peroxidation of lipids. The aim of this investigation was to assess the extent of lipid peroxidation both in cells and fluid obtained from bronchoalveolar lavage (BAL), and in lungs of rats exposed to crocidolite asbestos or titanium dioxide (TiO2), a nonfibrous particulate control. In comparison to sham and TiO2-exposed rats, the BAL fluid and cells of crocidolite-exposed animals contained significantly elevated levels of malondialdehyde (MDA), a breakdown product of lipid peroxidation detected using high-pressure liquid chromatography (HPLC). In contrast, no significant differences in MDA were detected in lavaged lung tissue from these animals. Inhalation of crocidolite caused an early inflammatory response characterized by elevated numbers of polymorphonuclear leukocytes and lymphocytes, as well as enhanced total protein in BAL. Pulmonary fibrosis and increased lung hydroxyproline also were observed after 20 days of exposure. Exposure to TiO2 did not cause inflammation, pulmonary fibrosis, or elevated amounts of hydroxyproline in the lung. Our results show that exposure to the fibrogenic and inflammatory mineral, crocidolite, results in an enhanced lipid peroxidation in BAL cells and fluid not observed after inhalation of the particulate TiO2. These novel observations suggest that MDA in BAL may be useful as a biomarker of exposure to inhaled asbestos or other oxidants.

• Detection of lipid peroxidation in lung and in bronchoalveolar lavage cells and fluid. J.M. Petruska, S.H. Wong, F.W. Sunderman, et al. 1990;9(1):51-8. PubMed · Publisher

  Abstract

  Inhalation of toxic materials such as asbestos, silica, 100% oxygen, ozone, or nitrogen dioxide may lead to an increased production of reactive oxygen metabolites which may initiate lipid peroxidation. Measurement of lipid peroxidation in cells and fluid obtained by bronchoalveolar lavage (BAL), as well as in lung tissue, may aid in monitoring the development and extent of pulmonary damage after inhalation of a toxic substance. In this study, we employed a sensitive assay for detection of malondialdehyde (MDA), a breakdown product of lipid peroxidation. By separation of the adduct with thiobarbituric acid, using a reverse phase high pressure liquid chromatographic technique, we accurately and sensitively measured the content of MDA in BAL cells, lavage fluid, and lavaged lung tissue homogenates of rats. The amounts of sample required for detection of MDA were small enough possibly to be applied to use with human specimens; in addition, recovery of added MDA was acceptable with all types of samples. Inclusion of a metal chelator in the preparation of samples appeared necessary to prevent metal-catalyzed propagation of lipid peroxidation during the assay. Overall, the method described here using samples from rats may be applicable to detecting lipid peroxidation in BAL samples from humans.

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