Asbestos conceives Fe(II)-dependent mutagenic stromal milieu through ceaseless macrophage ferroptosis and β-catenin induction in mesothelium

Our plain-English summary of Asbestos conceives Fe(II)-dependent mutagenic stromal milieu through ceaseless macrophage ferroptosis and β-catenin induction in mesothelium, by F. Ito, I. Yanatori, Y. Maeda, et al (Redox Biology, 2020).

The study

• Authors: F. Ito, I. Yanatori, Y. Maeda, K. Nimura, S. Ito, T. Hirayama, H. Nagasawa, N. Kohyama, Y. Okazaki, S. Akatsuka, S. Toyokuni
• Journal: Redox Biology
• Published: 2020;36:101616
• Cited by: 40 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Asbestos is still a social burden worldwide as a carcinogen causing malignant mesothelioma. Whereas recent studies suggest that local iron reduction is a preventive strategy against carcinogenesis, little is known regarding the cellular and molecular mechanisms surrounding excess iron. Here by differentially using high-risk and low-risk asbestos fibers (crocidolite and anthophyllite, respectively), we identified asbestos-induced mutagenic milieu for mesothelial cells. Rat and cell experiments revealed that phagocytosis of asbestos by macrophages results in their distinctive necrotic death; initially lysosome-depenent cell death and later ferroptosis, which increase intra- and extra-cellular catalytic Fe(II). DNA damage in mesothelial cells, as assessed by 8-hydroxy-2'-deoxyguanosine and γ-H2AX, increased after crocidolite exposure during regeneration accompanied by β-catenin activation. Conversely, β-catenin overexpression in mesothelial cells induced higher intracellular catalytic Fe(II) with increased G2/M cell-cycle fraction, when p16INK4A genomic loci localized more peripherally in the nucleus. Mesothelial cells after challenge of H2O2 under β-catenin overexpression presented low p16INK4A expression with a high incidence of deletion in p16INK4A locus. Thus, crocidolite generated catalytic Fe(II)-rich mutagenic environment for mesothelial cells by necrotizing macrophages with lysosomal cell death and ferroptosis. These results suggest novel molecular strategies to prevent mesothelial carcinogenesis after asbestos exposure.

Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.

In plain English

This laboratory study combined rat experiments with cell-culture work to compare a high-risk asbestos (crocidolite) with a low-risk one (anthophyllite). Male Fischer-344 rats received 10 mg of either fiber intraperitoneally; histology four weeks later showed granulomatous peritonitis with both fibers, with asbestos inside CD68-positive macrophages rather than mesothelial cells, but necrotic macrophage death, higher 4-hydroxy-2-nonenal staining and higher Berlin blue iron deposition only after crocidolite. Separately, catalytic Fe(II) measured with SiRhoNox-1 in peritoneal lavage macrophages and fluid was significantly higher after crocidolite than anthophyllite. In THP1 macrophages, crocidolite ruptured phagolysosomal membranes (anthophyllite did not), moved cathepsin B into nucleus and cytosol, and at 96 h changed ferroptosis markers (GPX4 and TfR1 decreased, ferritin increased); necrostatin-1, ferrostatin-1 and deferoxamine partially suppressed cell death. Microarray and GSEA of rat mesothelial cells showed Wnt/beta-catenin activation during regeneration; 8-OHdG and gamma-H2AX rose in mesothelial cells. Beta-catenin overexpression raised catalytic Fe(II), greatest at G2/M, when p16INK4A loci lay nearer the nuclear membrane; iron plus H2O2 with LiCl produced hemizygous p16INK4A deletion. The authors concluded crocidolite creates a catalytic Fe(II)-rich mutagenic environment for mesothelial 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: Experimental laboratory study: rat intraperitoneal asbestos injection model combined with cell-culture (macrophage and mesothelial cell line) experiments, microarray/GSEA, FISH and imaging
• Who was studied: Male Fischer-344 rats (6 weeks old, 140-150 g) given intraperitoneal asbestos; and cell lines: Met5A and LP9 human mesothelial, RAW264 mouse and THP1 human macrophage, HEK293T (Nagoya University, Japan)
• Exposure: Crocidolite (described by the authors as high-risk) versus anthophyllite (low-risk); 10 mg intraperitoneally in rats, 15-25 micrograms per square centimetre in cells, with or without added iron (ferric ammonium citrate) and H2O2
• Main results:
  • In rats, necrotic macrophage death in granuloma occurred only after crocidolite, not after anthophyllite
  • 4-hydroxy-2-nonenal-modified protein immunopositivity was significantly higher in crocidolite granuloma than in anthophyllite granuloma
  • Iron deposition (Berlin blue) in intra-granuloma macrophages increased only after crocidolite exposure
  • Catalytic Fe(II), detected with SiRhoNox-1 by flow cytometry and plate reader, was significantly higher in the peritoneal lavage macrophage fraction and in lavage fluid 4 weeks after crocidolite than after anthophyllite
  • In THP1 macrophages, crocidolite reached the phagolysosome and permeabilized its membrane at 24 h, whereas anthophyllite did not
  • At 96 h after crocidolite exposure in macrophages, Lamp2 and cathepsin B increased (lysosome-dependent cell death), GPX4 and TfR1 decreased and ferritin increased (ferroptosis), RIP1 and MLKL increased (necroptosis)
  • Necrostatin-1, ferrostatin-1 and deferoxamine partially suppressed macrophage detachment and death, while z-VAD-FMK, z-FA-FMK and pepstatin A were not effective
  • In rat mesothelial cells, gene set enrichment analysis showed Wnt/beta-catenin pathway activation, and CTNNB1 expression was significantly increased 1 week after crocidolite exposure
  • In rats, nuclear 8-OHdG in mesothelial cells on granuloma and fibrosis was seen only after crocidolite exposure; in co-culture it rose with iron dose
  • Regenerative mesothelial cells on granuloma showed DNA double-strand breaks (gamma-H2AX) after crocidolite but not after anthophyllite exposure
  • Beta-catenin overexpression in Met5A mesothelial cells significantly increased intracellular catalytic Fe(II) dose-dependently compared with mock transfection, highest at G2/M phase
  • p16INK4A loci lay centrally in the nucleus at G1 phase but near the nuclear membrane at G2/M phase; iron plus H2O2 with LiCl-induced beta-catenin stabilization was needed for significant hemizygous p16INK4A deletion in Met5A cells
  • H2O2 challenge decreased both P15 and P16 expression in surviving mesothelial cells
• The authors' conclusion: The authors concluded that crocidolite generates a catalytic Fe(II)-dependent mutagenic stromal environment through macrophage lysosome-dependent cell death and ferroptosis, and that beta-catenin activation during mesothelial regeneration brings several advantages for p16INK4A deletion; they suggested the beta-catenin pathway as a possible preventive target
• Limitations the authors state:
  • The authors could not exclude a minor contribution of other forms of regulated cell death
  • Mitochondrial injury observed after crocidolite exposure was not explained and needs further study
  • TfR1 results with the inhibitors used were inconsistent and were interpreted speculatively

Topics

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

Medical terms in this study, explained in our Lexicon: Macrophage, Mesothelium, Malignant Mesothelioma, Carcinogenesis, Crocidolite, Anthophyllite, Mesothelial Cells, Phagocytosis.

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