The crocidolite fibres interaction with human mesothelial cells as investigated by combining electron microscopy, atomic force and scanning near-field optical microscopy

Our plain-English summary of The crocidolite fibres interaction with human mesothelial cells as investigated by combining electron microscopy, atomic force and scanning near-field optical microscopy, by L. Andolfi, E. Trevisan, M. Zweyer, et al (Journal of Microscopy, 2013).

The study

• Authors: L. Andolfi, E. Trevisan, M. Zweyer, S. Prato, B. Troian, F. Vita, V. Borelli, M.R. Soranzo, M. Melato, G. Zabucchi
• Journal: Journal of Microscopy
• Published: 2013;249(3):173-83
• Cited by: 10 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

In this study, we have performed a morphological analysis of crocidolite fibres interaction with mesothelial cells (MET5A) by combining conventional electron microscopy with atomic force (AFM) and scanning near-field optical microscopy (SNOM). After 6-h exposure at a crocidolite dose of 5 μg cm(-2), 90% of MET5A cells interact with fibres that under these conditions have a low cytotoxic effect. SEM images point out that fibres can be either engulfed by the cells that lose their typical morphology or they can accumulate over or partially inside the cells, which preserve their typical spread morphology. By using AFM we are able to directly visualize the entry-site of nanometric-sized fibres at the plasma membrane of the spread mesothelial cells. More importantly, the crocidolite fibres that are observed to penetrate the plasma membrane in SNOM topography can be simultaneously followed beneath the cell surface in the SNOM optical images. The analysis of SNOM data demonstrates the entrance of crocidolite fibres in proximity of nuclear compartment, as observed also in the TEM images. Our findings indicate that the combination of conventional electron microscopy with novel nanoscopic techniques can be considered a promising approach to achieve a comprehensive morphological description of the interaction between asbestos fibres and mesothelial cells that represents the early event in fibre pathogenesis.

© 2013 The Authors Journal of Microscopy © 2013 Royal Microscopical Society.

In plain English

In this laboratory study, human mesothelial cells of the MET5A cell line were grown in dishes and exposed to standard crocidolite asbestos fibres at 5 micrograms per square centimetre for 3, 6 and 12 hours. Unexposed cells served as controls. No people or animals were studied. The authors combined electron microscopy with atomic force microscopy and scanning near-field optical microscopy (SNOM) to look at how fibres met the cells. Over 80% of cells were viable after 12 hours, and 90% after 6 hours, the time point the imaging focused on. Fibres were attached to more than 60% of cells at 3 hours and 80% at 6 hours. Most cells kept their flat shape, with fibres lying on them or partly inside. In about 5% of treated cells, a fibre went through the whole cell, which became rounded. Atomic force images showed where fibres entered the cell membrane. SNOM images let the authors follow fibres beneath the surface, some near the nucleus. Electron microscopy showed fibres inside or close to the nucleus. The authors concluded that combining these methods gives a comprehensive picture of how the fibres interact with the 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: Laboratory (in vitro) cell culture study using microscopy (SEM, TEM, AFM and SNOM) and a cell viability assay
• Who was studied: Human mesothelial cells of the MET5A cell line grown on coated glass coverslips. Unexposed cells were treated the same way and used as controls.
• Exposure: Crocidolite Asbestos UICC Standard fibres at 5 micrograms per square centimetre, for 3, 6 and 12 hours. The morphological analysis focused on 6 hours.
• Main results:
  • Over 80% of exposed cells were viable after 12 hours. 90% were viable after 6 hours.
  • By SEM, crocidolite fibres were attached to more than 60% of cells after 3 hours and to 80% after 6 hours. The share did not increase with longer exposure.
  • Two cell appearances were seen. In most cells, fibres lay over the membrane or were partly inside a cell that kept its flat shape. In about 5% of treated cells, a fibre went through the whole cell body and the cell became globular.
  • AFM showed a fibre about 30 nm thick lying on the membrane and another about 100 nm thick penetrating below it. On another cell, an entering fibre was about 600 nm thick.
  • In SNOM optical images, the part of a fibre inside the cell looked different from the part outside, which allowed the fibre's route to be followed beneath the plasma membrane.
  • Fibres that SNOM images showed penetrating near the nucleus were 15 to 30 micrometres long and 100 to 300 nm thick. The authors wrote that the images suggest the fibre enters the cell near the nuclear area.
  • TEM images showed crocidolite fibres inside or close to the nucleus of the mesothelial cells.
• The authors' conclusion: The authors concluded that combining SEM, TEM, AFM and SNOM gives a comprehensive morphological description of how crocidolite fibres interact with mesothelial cells. They also concluded that SNOM can tell fibres inside the cell from fibres outside it in flat mesothelial cells.
• Limitations the authors state:
  • With SEM, the fibre entry site at the membrane was sometimes hard to recognise, likely because of the metal coating SEM requires.
  • During AFM imaging, fibres weakly bound to the membrane could interfere with the scanning tip.
  • TEM sectioning does not keep fibres intact and gives only a few sections of these thin cells, so TEM provides only a partial view.

Topics

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

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

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