Asbestos fibers and interleukin-1 upregulate the formation of reactive nitrogen species in rat pleural mesothelial cells

Our plain-English summary of Asbestos fibers and interleukin-1 upregulate the formation of reactive nitrogen species in rat pleural mesothelial cells, by N. Choe, S. Tanaka, E. Kagan (American Journal of Respiratory Cell and Molecular Biology, 1998).

The study

• Authors: N. Choe, S. Tanaka, E. Kagan
• Journal: American Journal of Respiratory Cell and Molecular Biology
• Published: 1998;19(2):226-36
• Cited by: 65 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Nitric oxide radical (.NO) and peroxynitrite anion (ONOO-) have been implicated in lung inflammation and may be important in pleural injury. The present study was undertaken to determine the effects of asbestos exposure and cytokine stimulation on .NO and ONOO- production by rat pleural mesothelial cells. Accordingly, rat parietal pleural mesothelial cells were cultured for 2 to 72 h with or without 50 ng/ml of recombinant interleukin-1beta (IL-1beta) in the presence (1.05 to 8.4 microg/cm2) or absence of crocidolite or chrysotile asbestos fibers. The effects of asbestos were compared with those of carbonyl iron, a nonfibrogenic particulate. Mesothelial cell messenger RNA (mRNA) expression of the inducible form of .NO synthase (iNOS), assessed with the reverse transcription-polymerase chain reaction (RT-PCR), increased progressively from 2 to 12 h in IL-1beta-containing cultures. Nitrite (NO2-), the stable oxidation product of .NO in mesothelial cell conditioned medium, was assayed through the Griess reaction. Both types of asbestos fibers (chrysotile > crocidolite) upregulated the formation of NO2- in mesothelial cells costimulated with IL-1beta in a concentration-dependent and time-dependent fashion. In contrast, carbonyl iron did not upregulate NO2- formation in IL-1beta-stimulated cells. Both types of asbestos fibers also induced iNOS protein expression and the formation of nitrotyrosine in mesothelial cells and greatly induced the formation of nitrate (NO3-), a surrogate marker of ONOO- formation, in IL-1beta-stimulated cells. However, the effects of chrysotile were notably greater than those of crocidolite. These findings may have significance for the induction of pleural injury by asbestos fibers.

In plain English

This was a laboratory study of cultured cells, not people or living animals. The authors cultured pleural mesothelial cells scraped from the chest-wall lining of adult Fischer-344 rats. They exposed the cells to crocidolite or chrysotile asbestos fibers (1.05 to 8.4 micrograms per square centimetre), with or without the immune signalling protein interleukin-1 beta (IL-1β). Carbonyl iron, a non-fibrous particle, was the comparison. Asbestos alone did not switch on the nitric-oxide-producing enzyme iNOS or produce nitrite. With IL-1β, both fiber types raised nitrite, a marker of nitric oxide, in a dose- and time-dependent way. After 48 hours at the highest dose, nitrite measured 21.83 micromolar with IL-1β alone, 56.35 with crocidolite, 59.9 with chrysotile and 31.36 with carbonyl iron. With IL-1β, asbestos also increased nitrate, the share of iNOS-positive cells, nitrotyrosine staining, and release of LDH, an enzyme that leaks from damaged cells. Chrysotile's effects were generally greater than crocidolite's. The authors concluded that both fibers can induce reactive nitrogen species in these cells when costimulated with IL-1β, and that further studies are needed to determine the significance in living animals.

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: Laboratory (in vitro) cell culture experiment
• Who was studied: Parietal pleural mesothelial cells from adult Fischer-344 rats, grown in culture and used at passages 10 to 12. The number of rats is not stated.
• Exposure: Crocidolite or chrysotile asbestos fibers (NIEHS samples) or carbonyl iron particles at 1.05 to 8.4 micrograms per square centimetre. Cells were given these with or without recombinant IL-1β (50 ng/ml). Some experiments used interferon-gamma or TNF-alpha instead of IL-1β. Cultures ran for 2 to 72 hours.
• Main results:
  • Without IL-1β, neither crocidolite nor chrysotile (4 micrograms per square centimetre) induced iNOS mRNA expression in the mesothelial cells.
  • In IL-1β-containing cultures, cells exposed to chrysotile or crocidolite produced significantly more nitrite at 48 and 72 hours than cells given IL-1β alone.
  • A dose-response relationship for nitrite formation was shown for both crocidolite and chrysotile in IL-1β-containing cultures. Chrysotile was more effective than crocidolite at all concentrations tested.
  • Carbonyl iron particles did not increase IL-1β-induced nitrite production at any concentration tested.
  • After 48 hours at 8.4 micrograms per square centimetre, nitrite (mean, micromolar) was 21.83 with IL-1β alone, 56.35 with IL-1β plus crocidolite, 59.9 with IL-1β plus chrysotile, and 31.36 with IL-1β plus carbonyl iron.
  • In the same 48-hour experiment, LDH release (U/liter) was 62.06 with IL-1β alone, 117.72 with IL-1β plus crocidolite, 134.95 with IL-1β plus chrysotile, and 87.0 with IL-1β plus carbonyl iron. The authors reported generally good correlation between LDH release and nitrite formation.
  • The iNOS inhibitor aminoguanidine significantly reduced nitrite production in asbestos-containing cultures. Cytochalasin B, which blocks uptake of particles by cells, reduced nitrite by approximately 45% to 49%.
  • Nitrate, which the authors used as a marker of peroxynitrite, was significantly greater in IL-1β plus asbestos cultures than with IL-1β alone or IL-1β plus carbonyl iron. Chrysotile-exposed cultures had the highest nitrate content.
  • At 36 hours, the percentage of iNOS-positive cells was 7.03 with IL-1β alone, 32.54 with IL-1β plus crocidolite, 74.63 with IL-1β plus chrysotile, and 12.34 with IL-1β plus carbonyl iron.
  • Strong nitrotyrosine staining was seen in more than 90% of cells given chrysotile plus IL-1β. Staining was weaker with crocidolite plus IL-1β, and none was detected with carbonyl iron plus IL-1β.
• The authors' conclusion: The authors concluded that both chrysotile and crocidolite can induce the formation of reactive nitrogen species in cultured rat pleural mesothelial cells costimulated with IL-1β. They found it of special interest that chrysotile's effects were greater, since amphiboles had previously been suggested to have greater potential to cause pleural injury. They stated that further studies are needed to determine the in vivo significance.
• Limitations the authors state:
  • The RT-PCR technique used could not have detected subtle quantitative differences in iNOS mRNA between cultures with and without asbestos.
  • The signalling pathways involved in increased nitric oxide formation were not addressed in the study.
  • The findings come from cultured cells, and their significance in living animals still needs to be determined.

Topics

Medical terms in this study, explained in our Lexicon: Reactive Nitrogen Species (RNS), Mesothelial Cells, Peroxynitrite, Inflammation, Asbestos Exposure, Cytokine, Crocidolite, Chrysotile.

Related studies

• Contribution of reactive oxygen and nitrogen species to particulate-induced lung injury — S. Zhu, M. Manuel, et al. (1998)
• Asbestos exposure upregulates the adhesion of pleural leukocytes to pleural mesothelial cells via VCAM-1 — N. Choe, J. Zhang, et al. (1999)
• Interleukin-1-mediated release of interleukin-8 by asbestos-stimulated human pleural mesothelial cells — D.E. Griffith, E.J. Miller, et al. (1994)
• Mechanisms of asbestos-induced nitric oxide production by rat alveolar macrophages in inhalation and in vitro models — T.R. Quinlan, K.A. BeruBe, et al. (1998)
• Asbestos fibers and interferon-gamma up-regulate nitric oxide production in rat alveolar macrophages — G. Thomas, T. Ando, et al. (1994)
• Patterns of 8-hydroxydeoxyguanosine formation in DNA and indications of oxidative stress in rat and human pleural mesothelial cells after exposure to crocidolite asbestos — H. Fung, Y.W. Kow, et al. (1997)

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