Asbestos inhalation induces tyrosine nitration associated with extracellular signal-regulated kinase 1/2 activation in the rat lung

Our plain-English summary of Asbestos inhalation induces tyrosine nitration associated with extracellular signal-regulated kinase 1/2 activation in the rat lung, by A. Iwagaki, N. Choe, Y. Li, et al (American Journal of Respiratory Cell and Molecular Biology, 2003).

The study

• Authors: A. Iwagaki, N. Choe, Y. Li, D.R. Hemenway, E. Kagan
• Journal: American Journal of Respiratory Cell and Molecular Biology
• Published: 2003;28(1):51-60
• Cited by: 18 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Nitration of proteins by peroxynitrite (ONOO-) has been shown to critically alter protein function in vitro. We have shown previously that asbestos inhalation induced nitrotyrosine formation, a marker of ONOO- production, in the rat lung. To determine whether asbestos-induced protein nitration may affect mitogen-activated protein kinase (MAPK) signaling pathways, lung lysates from crocidolite and chrysotile asbestos-exposed rats and from sham-exposed rats were immunoprecipitated with anti-nitrotyrosine antibody, and captured proteins were subjected to Western blotting with anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibodies. Both types of asbestos inhalation induced significantly greater phosphorylation of ERK1/2 in rat lung lysates than was noted after sham exposure. Phosphorylated ERK proteins co-immunoprecipitated with nitrotyrosine. Moreover, in MAPK functional assays using Elk-1 substrate, immunoprecipitated phospho-ERK1/2 in lung lysates from both crocidolite-exposed and chrysotile-exposed rats demonstrated significantly greater phosphorylation of Elk-1 than was noted after sham exposure. Asbestos inhalation also induced ERK phosphorylation in bronchoalveolar lavage cells. Lung sections from rats exposed to crocidolite or chrysotile (but not from sham-exposed rats nor from rats exposed to "inert" carbonyl iron particles) demonstrated strong immunoreactivity for nitrotyrosine and phospho-ERK1/2 in alveolar macrophages and bronchiolar epithelium. These findings suggest that asbestos fibers may activate the ERK signaling pathway by generating ONOO- or other nitrating species that induce tyrosine nitration and phosphorylation of critical signaling molecules.

In plain English

In this animal experiment, three groups of adult male Fischer-344 rats breathed crocidolite asbestos, chrysotile asbestos, or filtered room air (sham) for 6 hours a day, 5 days a week, for 2 weeks. Analyses used 3 rats per group. Archival lungs from rats exposed for 5 hours to chrysotile, carbonyl iron particles (an "inert" control) or room air were also stained. Lungs from both asbestos groups contained two nitrated proteins (44 and 42 kD) not seen in sham rats. Total ERK, a cell-signaling protein, did not differ between groups. Its activated (phosphorylated) form was significantly greater after either fiber than after sham exposure, in lung tissue and lavage cells; the fibers did not differ significantly on this measure. ERK activity in lung tissue, measured on its target Elk-1, was also greater than in sham rats. Activated ERK was captured together with nitrated proteins. Most alveolar macrophages in asbestos-exposed lungs stained strongly for nitrotyrosine, while sham and carbonyl iron lungs showed only occasional staining. The authors concluded ERK phosphorylation was associated with tyrosine nitration. They could not tell whether both occurred in the same cells, and did not study the mechanism.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Animal experiment: inhalation exposure of rats, followed by laboratory analysis of lung tissue and lavage cells (Western blotting, immunoprecipitation, kinase activity assays, immunohistochemistry)
• Who was studied: Adult male Fischer-344 rats in three exposure groups. Figure legends give 3 rats per exposure group. The authors also used archival lung specimens from Tulane University: three groups of three rats exposed for 5 hours to chrysotile, carbonyl iron particles or room air.
• Exposure: UICC crocidolite asbestos, NIEHS chrysotile asbestos, or filtered room air (sham), by whole-body inhalation for 6 hours a day, 5 days a week, over 2 weeks. Rats were killed 14 days after exposure ended. The archival rats had 5-hour exposures to chrysotile, carbonyl iron or room air.
• Main results:
  • Two nitrated protein bands, of 44 and 42 kD, were detected in lung lysates from crocidolite-exposed and chrysotile-exposed rats but were not evident in lysates from sham-exposed rats.
  • Total amounts of ERK proteins did not differ between exposure groups, in either whole lung or lavage cell lysates.
  • Phosphorylation of ERK1/2 was significantly greater after crocidolite or chrysotile exposure than after sham exposure, in both lung lysates and lavage cell lysates.
  • For ERK phosphorylation, no statistically significant differences were noted between the effects of chrysotile and crocidolite inhalation.
  • ERK activity (measured as phosphorylation of the Elk-1 substrate) in lung lysates was significantly greater in both asbestos-exposed groups than in sham-exposed rats.
  • Phosphorylated ERK1/2 proteins were captured together with nitrated proteins (anti-nitrotyrosine immunoprecipitates), with significantly greater phosphorylation after both types of asbestos than after sham exposure.
  • In lung sections from crocidolite-exposed rats, the majority of alveolar macrophages and many bronchiolar epithelial cells showed intense staining for nitrotyrosine. A similar pattern was seen after 5-hour or 2-week chrysotile exposure. Only occasional alveolar macrophages and isolated bronchial epithelial cells stained in sham-exposed and carbonyl iron-exposed rats.
  • Staining for phosphorylated ERK1/2 was seen in bronchiolar epithelial cells and many alveolar macrophages of asbestos-exposed rats, but was rare in sham-exposed or carbonyl iron-exposed rats.
• The authors' conclusion: The authors concluded that both serpentine (chrysotile) and amphibole (crocidolite) asbestos fibers induced protracted ERK phosphorylation in the rat lung and upregulated Elk-1 phosphorylation by ERK. They stated that ERK phosphorylation was associated with tyrosine nitration. They also suggested that these effects were unique to inhaled asbestos fibers, compared with sham and carbonyl iron exposure.
• Limitations the authors state:
  • Whether phosphorylated ERK and nitrated proteins were located in the same lung cells could not be answered, because dual labeling within individual cells was difficult to interpret.
  • The study did not address the mechanism by which asbestos-induced tyrosine nitration might have caused ERK activation.
  • Only a limited amount of protein could be extracted from lavage cells, so these cells were used only for Western blots of phosphorylated ERK, not for immunoprecipitation or kinase activity assays.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Chrysotile, Crocidolite.

Medical terms in this study, explained in our Lexicon: Tyrosine Nitration, Peroxynitrite, Crocidolite, Chrysotile, Bronchoalveolar Lavage (BAL), Alveolar Macrophage, Epithelium.

Related studies

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• Persistent localization of activated extracellular signal-regulated kinases (ERK1/2) is epithelial cell-specific in an inhalation model of asbestosis — A.B. Cummins, C. Palmer, et al. (2003)
• Increased phosphorylated extracellular signal-regulated kinase immunoreactivity associated with proliferative and morphologic lung alterations after chrysotile asbestos inhalation in mice — R.F. Robledo, S.A. Buder-Hoffmann, et al. (2000)
• Role of extracellular signal-regulated protein kinases in apoptosis by asbestos and H2O2 — L.A. Jiménez, C. Zanella, et al. (1997)
• Asbestos-mediated CREB phosphorylation is regulated by protein kinase A and extracellular signal-regulated kinases 1/2 — C.A. Barlow, T.F. Barrett, et al. (2007)
• An extracellular signal-regulated kinase 2 survival pathway mediates resistance of human mesothelioma cells to asbestos-induced injury — A. Shukla, T.F. Barrett, et al. (2011)

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