DNA breakage in asbestos-treated normal and transformed (TSV40) rat pleural mesothelial cells

Our plain-English summary of DNA breakage in asbestos-treated normal and transformed (TSV40) rat pleural mesothelial cells, by V. Levresse, A. Renier, F. Levy, et al (Mutagenesis, 2000).

The study

• Authors: V. Levresse, A. Renier, F. Levy, V.C. Broaddus, M. Jaurand
• Journal: Mutagenesis
• Published: 2000;15(3):239-44
• Cited by: 31 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Asbestos has been shown to induce cell cycle arrest, DNA repair and some abnormalities consistent with DNA damage but not DNA breakage. The purpose of the study was to investigate DNA breakage in asbestos-exposed rat pleural mesothelial cells (RPMC). RPMC were compared with their transformed counterparts, RPMC-TSV40 (i.e. p53-inactivated by infection with a retroviral recombinant encoding the SV40 large T antigen), as in the latter cells the cell cycle does not arrest and DNA repair is deficient due to ineffective p53-dependent cell cycle control. RPMC and RPMC-TSV40 were exposed to chrysotile and crocidolite asbestos and also to camptothecin for comparison. The presence of DNA breakage was determined using the single cell gel (Comet) assay with alkaline electrophoresis and quantified by measuring comet tail length (TL) and the percentage of total DNA in the tail and calculating tail moment (TM). We found that comets were generated by both types of asbestos in RPMC and in RPMC-TSV40 as well as by camptothecin in RPMC. On a per weight basis, chrysotile induced more abnormalities in comet parameters than did crocidolite. The comet TL and TM increased with fibre concentration, although less so with crocidolite than with chrysotile. When exposed to chrysotile at similar concentrations, RPMC consistently showed more abnormal comet parameters than did RPMC-TSV40. We concluded that asbestos causes DNA breakage and suggest that some of the DNA breakage measured was due to repair mechanisms in the normal RPMC.

In plain English

This laboratory study tested whether asbestos breaks the DNA of rat pleural mesothelial cells grown in culture. The authors compared normal cells with modified cells in which p53 was inactivated, so their growth cycle does not pause and DNA repair is deficient. For 24 hours, normal cells received chrysotile at 0.5, 2, 5 or 10 µg/cm2, modified cells received chrysotile at 2, 5 or 10 µg/cm2, and both cell types received UICC crocidolite at 2 or 10 µg/cm2. DNA breakage was measured with the comet assay. In normal cells, chrysotile significantly increased breakage at every concentration, including 0.5 µg/cm2. UICC crocidolite was much less effective and differed from untreated normal cells only at 10 µg/cm2. A second sample, NIEHS crocidolite, differed at both 2 and 10 µg/cm2. With chrysotile, modified cells showed smaller changes than normal cells at all doses tested. With UICC crocidolite at 10 µg/cm2, the two cell types did not differ significantly. The authors concluded that asbestos causes DNA breakage and suggested that some breakage in normal cells came from the cells' own repair. They noted that the method may have difficulty analysing heavily damaged cells.

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: in vitro (cell culture) experiment using the alkaline comet assay to measure DNA breakage
• Who was studied: Cultured normal rat pleural mesothelial cells (RPMC) from Sprague–Dawley rats, and RPMC-TSV40 cells (RPMC made to express SV40 large T antigen, with p53 inactivated); 150 cells analysed per series
• Exposure: Normal RPMC: UICC Rhodesian chrysotile at 0.5, 2, 5 or 10 µg/cm2 or UICC crocidolite at 2 or 10 µg/cm2 for 24 h. RPMC-TSV40: chrysotile at 2, 5 and 10 µg/cm2 and UICC crocidolite at 2 and 10 µg/cm2. In some experiments NIEHS crocidolite was used. Camptothecin (5 µg/ml) served as a reference agent in normal RPMC.
• Main results:
  • In untreated normal RPMC, at least 85% of cells had unbroken DNA
  • In normal RPMC, chrysotile increased tail moment even at the lowest concentration, 0.5 µg/cm2
  • In normal RPMC, tail moment distribution differed significantly from untreated cultures at all chrysotile concentrations
  • In normal RPMC, the increase in mean tail moment with chrysotile levelled off (plateaued) from 2 µg/cm2
  • In normal RPMC, UICC crocidolite produced greater comets than in untreated cells only at 10 µg/cm2
  • In normal RPMC, at concentrations the authors describe as having equivalent fibre numbers, mean tail moment was 5.3 with chrysotile 0.5 µg/cm2 versus 1.1 with UICC crocidolite 2 µg/cm2
  • In normal RPMC, NIEHS crocidolite produced comets, but with lower values than UICC crocidolite
  • In normal RPMC, tail moment distribution differed statistically from untreated cells in all groups exposed in this comparison: 2 and 10 µg/cm2 NIEHS crocidolite and 10 µg/cm2 UICC crocidolite
  • In RPMC-TSV40 cells, chrysotile-induced changes were smaller than in normal RPMC at all doses
  • With UICC crocidolite at 10 µg/cm2, comet parameters in RPMC-TSV40 cells were not significantly different from those in normal RPMC
• The authors' conclusion: The authors concluded that asbestos causes DNA breakage and suggested that some of the DNA breakage measured in normal RPMC was due to repair mechanisms. They also offered an alternative explanation for the lower breakage in RPMC-TSV40 cells: these cells may be less sensitive to chrysotile.
• Limitations the authors state:
  • The plateau at higher chrysotile doses may reflect a limitation of the method, because comets from heavily damaged cells may be difficult to analyse
  • The contribution of DNA repair could not be tested directly by removing the fibres, because the fibres are taken up into the cells

Topics

Medical terms in this study, explained in our Lexicon: Mesothelial Cells, SV40, Chrysotile, Crocidolite.

Related studies

• Effects of asbestos on initiation of DNA damage, induction of DNA-strand breaks, P53-expression and apoptosis in primary, SV40-transformed and malignant human mesothelial cells — B. Burmeister, T. Schwerdtle, et al. (2004)
• Effect of simian virus large T antigen expression on cell cycle control and apoptosis in rat pleural mesothelial cells exposed to DNA damaging agents — V. Levresse, S. Moritz, et al. (1998)
• Synthesis of poly(ADP-ribose) in asbestos treated rat pleural mesothelial cells in culture — H.Y. Dong, A. Buard, et al. (1995)
• Phagocytosis of crocidolite asbestos induces oxidative stress, DNA damage, and apoptosis in mesothelial cells — W. Liu, J.D. Ernst, et al. (2000)
• In vitro sensitivity of normal human mesothelial and malignant mesothelioma cell lines to four new chemotherapeutic agents — T. Ollikainen, A. Knuuttila, et al. (2000)
• Modulation of DNA single-strand breaks by intracellular glutathione in human lung cells exposed to asbestos fibers — A. Puhakka, T. Ollikainen, et al. (2002)

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