Short-term inhalation and in vitro tests as predictors of fiber pathogenicity

Our plain-English summary of Short-term inhalation and in vitro tests as predictors of fiber pathogenicity, by R.T. Cullen, B.G. Miller, J.M. Davis, et al (Environmental Health Perspectives, 1997).

The study

• Authors: R.T. Cullen, B.G. Miller, J.M. Davis, D.M. Brown, K. Donaldson
• Journal: Environmental Health Perspectives
• Published: 1997;105 Suppl 5(Suppl 5):1235-40
• Cited by: 18 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

A wide range of fiber types was tested in two in vitro assays: toxicity to A549 epithelial cells, as detachment from substrate, and the production of the proinflammatory cytokine tumor necrosis factor (TNF) by rat alveolar macrophages. Three of the fibers were also studied in vivo, using short-term inhalation followed by a) bronchoalveolar lavage to assess the inflammatory response and b) measurement of cell proliferation in terminal bronchioles and alveolar ducts, using incorporation of bromodeoxyuridine (BrdU). The amount of TNF produced by macrophages in vitro depended on the fiber type, with the man-made vitreous fibers, and refractory ceramic fibers being least stimulatory and silicon carbide (SiC) whiskers providing the greatest stimulation. In the epithelial detachment assay there were dose-dependent differences in the toxicity of the various fibers, with long amosite being the most toxic. However, there was no clear relationship to known chronic pathogenicity. Fibers studied by short-term inhalation produced some inflammation, but there was no clear discrimination between the responses to code 100/475 glass fibers and the more pathogenic amosite and SiC. However, measurements of BrdU uptake into lung cells showed that amosite and SiC produced a greater reaction than code 100/475, which itself caused no more proliferation than that seen in untreated lungs. These results mirror the pathogenicity ranking of the fibers in long-term experiments. In conclusion, the only test to show potential as a predictive measure of pathogenicity was that of cell proliferation in lungs after brief inhalation exposure (BrdU assay). We believe that this assay should be validated with a wider range of fibers, doses, and time points.

In plain English

This laboratory study tested whether quick tests could predict fiber disease-causing ability. Rat lung macrophages were exposed to amosite and crocidolite asbestos, two glass microfibers, two silicon carbide (SiC) whisker types, four refractory ceramic fibers and four man-made vitreous fibers. Lung epithelial (A549) cells received the asbestos, ceramic and vitreous fibers. Rats briefly inhaled long-fiber amosite, SiC whiskers or code 100/475 glass fiber. At equal mass doses, amosite caused the most cell detachment, but adjusted to equal fiber numbers it ranked fourth. Crocidolite caused no more detachment than many man-made fibers, and the authors found no clear link to known disease-causing ability. One SiC type, SiC1, stimulated by far the most macrophage production of the inflammatory signal TNF. All three inhaled fibers caused some lung inflammation, without clear differences between them. After one day of inhalation, amosite and SiC increased cell division in the terminal bronchioles and alveolar ducts, while code 100/475 matched unexposed rats. The authors concluded that only this cell-division test showed promise and that it should be validated with more fibers, doses and time points.

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 study: two in vitro (cell culture) assays plus a short-term inhalation experiment in rats
• Who was studied: Cell cultures (the alveolar epithelial cell line A549 and alveolar macrophages taken from rat lungs) and rats. The inhalation groups used 6 rats per time point for SiC whiskers and 6 to 12 rats per time point for amosite and code 100/475. The cell-proliferation (BrdU) groups used three rats each (four for long amosite), exposed for 1 day. The work was done at the Institute of Occupational Medicine and Napier University in Edinburgh, UK.
• Exposure: Macrophage TNF test: amosite and crocidolite asbestos; code 100/475 and code 104E glass fibers; two silicon carbide whisker types (SiC1, SiC2); refractory ceramic fibers RCF1 to RCF4; and man-made vitreous fibers MMVF10 and 11 (glass insulation wools), 21 (stone wool) and 22 (slag wool). The A549 detachment results reported in Tables 3 and 4 cover only long amosite, crocidolite, RCF1 to RCF4 and MMVF10, 11, 21 and 22, at mass doses of 10, 25, 50 and 100 micrograms per well. Inhalation: SiC1 whiskers, code 100/475 glass fiber and long-fiber amosite, at 1000 (World Health Organization) fibers/ml for 7 hours a day, for 1, 3, 8 or 14 days of exposure over 3 weeks.
• Main results:
  • In the epithelial cell detachment test at mass doses, the toxicity of the fibers differed and depended on dose, with amosite the most toxic.
  • The detachment results showed no clear relationship to known pathogenicity: crocidolite caused no more detachment than many of the man-made fibers. When the results were adjusted to a common fiber number, crocidolite and amosite moved much lower in the ranking: crocidolite ranked 1st (least detachment) and long amosite 4th of the 10 fibers.
  • TNF production by rat macrophages differed significantly between fibers. SiC1 whiskers were by far the most active (1527 units per million cells, against 125 for SiC2 and 37 with no fiber). The two asbestos samples ranked next and the glass microfibers were intermediate. The other fibers (the MMVF and RCF types) produced no more TNF than control cultures.
  • After short-term inhalation, each fiber type caused some lung inflammation. Lavage measurements did not clearly separate code 100/475 glass fibers from amosite and SiC, which the authors describe as more pathogenic.
  • After 1 day of inhalation, amosite and SiC increased cell proliferation in the terminal bronchioles and alveolar ducts. Mean BrdU-positive cells per millimeter of duct were 3.14 for long amosite and 2.80 for SiC1. The mean for code 100/475 (0.71) was close to that for unexposed controls (0.74). The effects were most marked in the upper levels of the lung.
• The authors' conclusion: The authors concluded that the only test showing potential to predict fiber pathogenicity was cell proliferation in the lungs after brief inhalation (the BrdU assay). They believe this assay should be validated with a wider range of fibers, doses and time points. They also stated that in vitro toxicity assays are not appropriate models of how fibers cause disease.
• Limitations the authors state:
  • In vitro and short-term in vivo tests generally do not account for the long-term dissolution of fibers in the lung.
  • The authors did not check whether macrophages engulfed the different fiber types to different degrees, which might affect the amount of TNF produced.
  • The TNF assay was not sensitive enough to separate the man-made vitreous fibers from the refractory ceramic fibers.

Topics

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

Medical terms in this study, explained in our Lexicon: Cytokine, Necrosis, Alveolar Macrophage, Bronchoalveolar Lavage (BAL), Macrophage, Vitreous, Amosite, Inflammation.

Related studies

• Short-term crocidolite inhalation studies in mice: validation of an inhalation chamber — B. Rihn, E. Kauffer, et al. (1996)
• In vitro activity of silicon carbide whiskers in comparison to other industrial fibers using four cell culture systems — N.F. Johnson, M.D. Hoover, et al. (1992)
• Pathogenicity of a special-purpose glass microfiber (E glass) relative to another glass microfiber and amosite asbestos — R.T. Cullen, A. Searl, et al. (2000)
• Superoxide anion release by alveolar macrophages exposed to respirable industrial fibres: modifying effect of fibre opsonisation — K. Donaldson, I.M. Hill, et al. (1995)
• An in vitro/in vivo study into the short term effects of exposure to mineral fibres — A. Clouter, C.E. Houghton, et al. (1996)
• Biopersistence of rock wool in lungs after short-term inhalation in rats — Y. Kudo, Y. Aizawa (2008)

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