Effects of Chrysotile Exposure in Human Bronchial Epithelial Cells: Insights into the Pathogenic Mechanisms of Asbestos-Related Diseases

Our plain-English summary of Effects of Chrysotile Exposure in Human Bronchial Epithelial Cells: Insights into the Pathogenic Mechanisms of Asbestos-Related Diseases, by G.R. Gulino, M. Polimeni, M. Prato, et al (Environmental Health Perspectives, 2016).

The study

• Authors: G.R. Gulino, M. Polimeni, M. Prato, E. Gazzano, J. Kopecka, S. Colombatto, D. Ghigo, E. Aldieri
• Journal: Environmental Health Perspectives
• Published: 2016;124(6):776-84
• Cited by: 20 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Chrysotile asbestos accounts for > 90% of the asbestos used worldwide, and exposure is associated with asbestosis (asbestos-related fibrosis) and other malignancies; however, the molecular mechanisms involved are not fully understood. A common pathogenic mechanism for these malignancies is represented by epithelial-mesenchymal transition (EMT), through which epithelial cells undergo a morphological transformation to assume a mesenchymal phenotype. In the present work, we propose that chrysotile asbestos induces EMT through a mechanism involving a signaling pathway mediated by tranforming growth factor beta (TGF-β).

Objectives: We investigated the role of chrysotile asbestos in inducing EMT in order to elucidate the molecular mechanisms involved in this event.

Methods: Human bronchial epithelial cells (BEAS-2B) were incubated with 1 μg/cm2 chrysotile asbestos for ≤ 72 hr, and several markers of EMT were investigated. Experiments with specific inhibitors for TGF-β, glycogen synthase kinase-3β (GSK-3β), and Akt were performed to confirm their involvement in asbestos-induced EMT. Real-time polymerase chain reaction (PCR), Western blotting, and gelatin zymography were performed to detect mRNA and protein level changes for these markers.

Results: Chrysotile asbestos activated a TGF-β-mediated signaling pathway, implicating the contributions of Akt, GSK-3β, and SNAIL-1. The activation of this pathway in BEAS-2B cells was associated with a decrease in epithelial markers (E-cadherin and β-catenin) and an increase in mesenchymal markers (α-smooth muscle actin, vimentin, metalloproteinases, and fibronectin).

Conclusions: Our findings suggest that chrysotile asbestos induces EMT, a common event in asbestos-related diseases, at least in part by eliciting the TGF-β-mediated Akt/GSK-3β/SNAIL-1 pathway.

Citation: Gulino GR, Polimeni M, Prato M, Gazzano E, Kopecka J, Colombatto S, Ghigo D, Aldieri E. 2016. Effects of chrysotile exposure in human bronchial epithelial cells: insights into the pathogenic mechanisms of asbestos-related diseases. Environ Health Perspect 124:776-784; http://dx.doi.org/10.1289/ehp.1409627.

In plain English

This was a laboratory study of human lung cells grown in dishes. No people or animals were studied. The authors exposed immortalized human bronchial epithelial cells (BEAS-2B) to UICC chrysotile asbestos at 1 μg/cm2 for up to 72 hours, and in other tests to UICC crocidolite at 5 μg/cm2 for 7 days. They looked for epithelial–mesenchymal transition (EMT), a change in which lining cells take on a stretched, fibroblast-like form. After chrysotile, the cells became spindle-shaped. They had less E-cadherin and beta-catenin and more alpha-smooth muscle actin, vimentin and fibronectin, and they secreted more of the enzymes MMP-2 and MMP-9. Crocidolite, and chrysotile in two other cell lines, gave similar marker changes. Chrysotile raised reactive oxygen species and the signaling protein TGF-beta. At 72 hours TGF-beta measured 244.75 pg/mg protein in treated cells versus 164.77 in untreated cells. Blocking TGF-beta, GSK-3beta or Akt partly reversed the changes. The authors concluded that chrysotile can trigger EMT in these cells at least partly through TGF-beta and the Akt/GSK-3beta/SNAIL-1 pathway, and that other mechanisms may yet be found.

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: Cultured human cell lines, not people: immortalized human bronchial epithelial cells (BEAS-2B, the main model), plus human bronchial epithelial cells (NuLi-1) and human lung adenocarcinoma alveolar epithelial cells (A549) for confirmation
• Exposure: UICC chrysotile and UICC crocidolite, sonicated before use. BEAS-2B cells received 1 μg/cm2 chrysotile for up to 72 hours, or 5 μg/cm2 crocidolite for 7 days. NuLi-1 cells received 1 μg/cm2 chrysotile for 96 hours. A549 cells received 5 μg/cm2 chrysotile for 96 hours. In some experiments, inhibitors of TGF-beta, GSK-3beta and Akt were added.
• Main results:
  • After 72 hours of chrysotile, BEAS-2B cells took on a spindle-shaped, fibroblast-like shape.
  • Chrysotile-exposed BEAS-2B cells had significantly lower levels of the epithelial proteins E-cadherin and beta-catenin than untreated cells.
  • Mesenchymal markers (alpha-smooth muscle actin, vimentin, fibronectin) were significantly increased in chrysotile-exposed BEAS-2B cells, and these cells secreted more MMP-2 and MMP-9 than untreated cells.
  • In BEAS-2B cells exposed to 5 μg/cm2 crocidolite for 7 days, E-cadherin and beta-catenin were also significantly decreased.
  • In NuLi-1 and A549 cells, chrysotile also significantly decreased E-cadherin and beta-catenin.
  • In BEAS-2B cells, chrysotile caused significant production of reactive oxygen species (ROS) after 30 minutes.
  • In BEAS-2B cells, TGF-beta rose significantly after 1 hour of chrysotile and was no longer significantly different from controls at 6 hours.
  • At 72 hours, TGF-beta was significantly higher in chrysotile-treated BEAS-2B cells (244.75, standard error 11.75, pg/mg protein) than in control cells (164.77, standard error 12.62, pg/mg protein), with p less than 0.05 over three independent experiments.
  • Adding a TGF-beta-blocking antibody together with chrysotile partially restored the marker protein levels in BEAS-2B cells.
  • Over 72 hours, a GSK-3beta inhibitor partially reversed the chrysotile-induced rise of SNAIL-1 in the cell nucleus and the fall in E-cadherin.
  • Over 72 hours, an Akt inhibitor partially restored nuclear SNAIL-1 and E-cadherin levels in chrysotile-exposed BEAS-2B cells.
• The authors' conclusion: The authors concluded that their findings suggest chrysotile can trigger EMT in BEAS-2B cells, at least in part, through TGF-beta and its intracellular effectors Akt/GSK-3beta/SNAIL-1. They described this as one of several possible mechanisms.
• Limitations the authors state:
  • Blocking the pathway did not fully reverse the changes. The authors explained this by TGF-beta being only one of the possible mediators of asbestos-induced EMT in these cells.
  • The authors stated that their findings describe just one of the possible molecular mechanisms and that additional mechanisms may be found.

Topics

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

Medical terms in this study, explained in our Lexicon: Chrysotile, Fibrosis.

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• Epithelial to Mesenchymal Transition in Human Mesothelial Cells Exposed to Asbestos Fibers: Role of TGF-β as Mediator of Malignant Mesothelioma Development or Metastasis via EMT Event — S. Turini, L. Bergandi, et al. (2019)
• Chrysotile Causes Human Bronchial Epithelial Cell Apoptosis in Response to the Fas-Mediated Apoptosis Pathway — M. Zhang, J. Chen, et al. (2017)
• Differentially expressed genes in asbestos-induced tumorigenic human bronchial epithelial cells: implication for mechanism — Y.L. Zhao, C.Q. Piao, et al. (2000)
• Malignant transformation of immortalized human bronchial epithelial cells by asbestos fibers — T.K. Hei, L.J. Wu, et al. (1997)
• Karyotype analysis of tumorigenic human bronchial epithelial cells transformed by chrysolite asbestos using chemically induced premature chromosome condensation technique — M. Suzuki, C.Q. Piao, et al. (2001)

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