Patterns of 8-hydroxydeoxyguanosine formation in DNA and indications of oxidative stress in rat and human pleural mesothelial cells after exposure to crocidolite asbestos

Our plain-English summary of Patterns of 8-hydroxydeoxyguanosine formation in DNA and indications of oxidative stress in rat and human pleural mesothelial cells after exposure to crocidolite asbestos, by H. Fung, Y.W. Kow, B. Van Houten, et al (Carcinogenesis, 1997).

The study

• Authors: H. Fung, Y.W. Kow, B. Van Houten, B.T. Mossman
• Journal: Carcinogenesis
• Published: 1997;18(4):825-32
• Cited by: 81 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Oxidative damage is a proposed mechanism of asbestos-induced carcinogenesis, but the detection of oxidative DNA lesions in target cells of asbestos-induced mesothelioma has not been examined. In studies here, DNA was isolated from both rat pleural mesothelial (RPM) cells and a human mesothelial cell line (MET5A) after exposure in vitro to crocidolite asbestos at various concentrations. DNA was then examined for formation of 8-hydroxydeoxyguanosine (8-OHdG) at 24, 48 and 72 h using HPLC with electrochemical detection. In addition, steady-state mRNA levels of manganese-containing superoxide dismutase (MnSOD) were assessed as an indication of oxidative stress. Whereas RPM cells showed dose-dependent and significant increases in 8-OHdG formation in response to crocidolite asbestos or iron-chelated crocidolite fibers (but not after exposure to glass beads), MET5A cells showed decreases in 8-OHdG. Both cell types exhibited elevations in message levels of MnSOD. In comparison with human MET5A cells, RPM cells exhibited increased cytotoxicity and apoptosis in response to asbestos, as documented by cell viability assays and flow cytometry analysis using propidium iodide. Results in RPM cells indicate that asbestos causes oxidative damage that may result in potentially mutagenic lesions in DNA and/or apoptosis, despite compensatory increases in expression of an antioxidant enzyme.

In plain English

This was a laboratory study of cells grown in dishes. The authors exposed rat pleural mesothelial (RPM) cells, taken from the parietal pleura of Fischer 344 rats, and a human mesothelial cell line (MET5A) made immortal with SV40 T antigen, to crocidolite asbestos at 2.5 to 10 µg/cm2 for up to 72 hours. Glass beads served as a non-fibrous comparison. They measured 8-hydroxydeoxyguanosine (8-OHdG), a marker of oxidative DNA damage, along with cell death, apoptosis (programmed cell death) and messages for the antioxidant enzyme MnSOD. In RPM cells, asbestos raised 8-OHdG to about 2-fold above control at 72 hours. Fibers treated to remove iron still raised it, while glass beads did not. In MET5A cells, 8-OHdG was about 2-fold lower than in untreated cells. Asbestos was more toxic to RPM cells, and apoptosis was more severe in RPM than MET5A cells at all time points. MnSOD messages rose in both. The authors suggested the differences may reflect immortalization of MET5A cells and/or species-specific effects, and noted that strict comparisons between the two cell types are unjustified.

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 comparing rat and human mesothelial cells exposed to asbestos. Measured outcomes were 8-OHdG in DNA, cytotoxicity, apoptosis and MnSOD mRNA.
• Who was studied: Rat pleural mesothelial (RPM) cells isolated from the parietal pleura of Fischer 344 rats, and a human pleural mesothelial cell line (MET5A) immortalized by transformation with SV40 T antigen
• Exposure: NIEHS crocidolite asbestos fibers (mean length 19 µm, mean diameter 0.25 µm) at 2.5 to 10 µg/cm2 of dish for up to 72 h. Comparisons were crocidolite treated with the iron chelators deferoxamine and ferrozine, and glass beads (1 to 4 µm diameter, 20 µg/cm2) as a non-fibrous particle control.
• Main results:
  • In human MET5A cells, 8-OHdG in asbestos-exposed cells was about 2-fold lower than in untreated cells at each time period. Decreases were statistically significant (p less than 0.05) at the highest asbestos concentration at all time points.
  • In RPM cells, both crocidolite concentrations increased 8-OHdG, with significant elevations (p less than 0.05) of about 2-fold above control values at 72 h
  • In RPM cells, iron-chelated crocidolite still significantly increased 8-OHdG; chelating iron did not reduce oxidative DNA damage
  • RPM cells exposed to glass beads at 20 µg/cm2 did not show increased 8-OHdG
  • Adding 10% serum to the medium did not change the asbestos-induced rise in 8-OHdG in RPM cells
  • In RPM cells, asbestos significantly reduced cell viability at both 2.5 and 10 µg/cm2 at all time periods, and toxicity increased over time. In MET5A cells, toxicity at 2.5 µg/cm2 at 24 h was not significant, and toxicity did not increase over time.
  • Apoptosis after asbestos exposure was significantly more severe (p less than 0.05) in RPM cells than in MET5A cells at all time points. In MET5A cells, asbestos-induced apoptosis was not significant at 72 h.
  • MnSOD mRNA levels rose in both cell types after asbestos exposure, with differing patterns over time. Glass beads did not alter MnSOD message levels.
  • In RPM cells, MnSOD mRNA was not increased at 72 h, whereas in MET5A cells MnSOD up-regulation persisted for 72 h
• The authors' conclusion: The authors concluded that 8-OHdG, a marker of oxidative DNA damage associated with mutagenesis, was significantly increased in RPM cells but decreased in human MET5A cells exposed to toxic concentrations of asbestos. They proposed that the differences may be due to alterations associated with immortalization of MET5A cells and/or species-specific effects.
• Limitations the authors state:
  • The authors stated that strict comparisons between RPM and MET5A cells are unjustified
  • The authors noted that MET5A cells may be more resistant than RPM cells to asbestos effects because of their immortalized state

Topics

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

Medical terms in this study, explained in our Lexicon: Oxidative Stress, Mesothelial Cells, Crocidolite, Carcinogenesis, DNA lesions, Mesothelioma, 8-OHdG, Superoxide.

Related studies

• Detection of 8-oxo-2'-deoxyguanosine, a marker of oxidative DNA damage, in culture medium from human mesothelial cells exposed to crocidolite asbestos — Q. Chen, J. Marsh, et al. (1996)
• Activation of p38 MAP kinase by asbestos in rat mesothelial cells is mediated by oxidative stress — W.A. Swain, K.J. O'Byrne, et al. (2004)
• Iron-dependent formation of 8-hydroxydeoxyguanosine in isolated DNA and mutagenicity in Salmonella typhimurium TA102 induced by crocidolite — S.P. Faux, P.J. Howden, et al. (1994)
• Potential role of ferritin heavy chain in oxidative stress and apoptosis in human mesothelial and mesothelioma cells: implications for asbestos-induced oncogenesis — W. Aung, S. Hasegawa, et al. (2007)
• Mechanisms of oxidative stress and alterations in gene expression by Libby six-mix in human mesothelial cells — J.M. Hillegass, A. Shukla, et al. (2010)
• Phagocytosis of crocidolite asbestos induces oxidative stress, DNA damage, and apoptosis in mesothelial cells — W. Liu, J.D. Ernst, et al. (2000)

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