Long-term response of rats to single intratracheal exposure of Libby amphibole or amosite

Our plain-English summary of Long-term response of rats to single intratracheal exposure of Libby amphibole or amosite, by J.M. Cyphert, D.J. Padilla-Carlin, M.C. Schladweiler, et al (Journal of Toxicology and Environmental Health. Part a, 2012).

The study

• Authors: J.M. Cyphert, D.J. Padilla-Carlin, M.C. Schladweiler, J.H. Shannahan, A. Nyska, U.P. Kodavanti, S.H. Gavett
• Journal: Journal of Toxicology and Environmental Health. Part a
• Published: 2012;75(3):183-200
• Cited by: 19 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

In former mine workers and residents of Libby, Montana, exposure to amphibole-contaminated vermiculite has been associated with increased incidences of asbestosis and mesothelioma. In this study, long-term effects of Libby amphibole (LA) exposure were investigated relative to the well-characterized amosite asbestos in a rat model. Rat-respirable fractions of LA and amosite (aerodynamic diameter≤2.5 μm) were prepared by water elutriation. Male F344 rats were exposed to a single dose of either saline, amosite (0.65 mg/rat), or LA (0.65 or 6.5 mg/rat) by intratracheal (IT) instillation. One year after exposure, asbestos-exposed rats displayed chronic pulmonary inflammation and fibrosis. Two years postexposure, lung inflammation and fibrosis progressed in a time- and dose-dependent manner in LA-exposed rats, although the severity of inflammation and fibrosis was smaller in magnitude than in animals exposed to amosite. In contrast, gene expression of the fibrosis markers Col 1A2 and Col 3A1 was significantly greater in LA-exposed compared to amosite-exposed rats. There was no apparent evidence of preneoplastic changes in any of the asbestos-exposed groups. However, all asbestos-exposed rats demonstrated a significant increase in the expression of epidermal growth factor receptor (EGFR) 2 yr after instillation. In addition, only LA-exposed rats showed significant elevation in mesothelin (Msln) and Wilms' tumor gene (WT1) expression, suggesting possible induction of tumor pathways. These results demonstrate that a single IT exposure to LA is sufficient to induce significant fibrogenic, but not carcinogenic, effects up to 2 yr after exposure that differ both in quality and magnitude from those elicited by amosite administration at the same mass dose in F344 rats. Data showed that LA was on a mass basis less potent than amosite.

In plain English

In this animal experiment, male Fischer 344 rats got a single dose into the windpipe of one of three things: saline, amosite (0.65 mg per rat), or Libby amphibole (0.65 or 6.5 mg per rat). Libby amphibole is the asbestos that contaminated vermiculite mined near Libby, Montana. Only fibers small enough for rats to breathe were used. Lungs were examined at 1 year (8 rats per group) and at about 22 months, which the article reports as 2 years (11 to 15 surviving rats per group). Both fibers caused long-lasting lung inflammation and fibrosis (scarring) that increased over time. With Libby amphibole, the higher dose caused more. At the same dose by weight, amosite caused more fibrosis than Libby amphibole. No lung tumors, mesotheliomas or precancerous changes were seen in any group. Some genes linked to mesothelioma were more active at 2 years: EGFR in all asbestos groups, and mesothelin and WT1 only in the low-dose Libby amphibole group. The authors noted that the study ended early because many rats died, which might explain why no cancer was 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: Animal experiment: single intratracheal (into the windpipe) instillation in rats, with lungs examined 1 and 2 years later
• Who was studied: Healthy male Fischer 344 rats, 6–8 weeks old, studied at a U.S. EPA laboratory in Research Triangle Park, North Carolina, in four groups. There were 8 rats per group at 1 year. 24 rats per group were assigned to the 2-year necropsy, but unscheduled deaths (mainly leukemia and various tumors typical of this strain) reduced this to 11–15 rats per group. The final necropsy was at about 22 months. Study dates are not given.
• Exposure: A single dose of saline (negative control), Libby amphibole at 0.65 or 6.5 mg per rat, or amosite (positive control) at 0.65 mg per rat. Only rat-respirable fractions (aerodynamic diameter 2.5 μm or less), separated by water elutriation, were used. Libby amphibole is described as a mixture of amphibole types, primarily tremolite, winchite and richterite.
• Main results:
  • Inflammatory cells counted in fluid washed from the lungs (total cells, macrophages, neutrophils, lymphocytes) did not differ significantly among the four groups at 1 or 2 years.
  • At 1 year, albumin in fluid washed from the lungs was significantly increased in all asbestos-treated groups. The other lung-injury markers were at control levels.
  • Fibrosis increased with dose and time in rats given Libby amphibole, but was significantly greater in rats given amosite. Average fibrosis scores (0 = none, 3 = moderate) at 1 year and 2 years were: saline 0 and 0; low-dose Libby amphibole 0.25 and 0.67; high-dose Libby amphibole 0.88 and 1.88; amosite 1.50 and 3.0.
  • Collagen gene expression did not change at 1 year. At 2 years it trended upward in all asbestos groups, but only Col 1A2 and Col 3A1 in the low-dose Libby amphibole group were significantly higher than in saline-treated rats (6 rats per group tested).
  • No preneoplastic changes, lung tumors or mesotheliomas were seen under the microscope at 1 or 2 years in rats given Libby amphibole. No carcinogenic changes were seen in rats given amosite either.
  • Mesothelioma-associated gene markers (6 rats per group tested): at 1 year, WT1 expression was threefold higher in the low-dose Libby amphibole group. At 2 years, EGFR was significantly increased in all asbestos-exposed groups (2.5-fold compared with saline), while Msln and WT1 were significantly increased only in the low-dose Libby amphibole group.
  • Serum mesothelin was unchanged at 1 year. At 2 years it was significantly elevated in 43% of animals from all groups, including saline controls, compared with 1-year levels. The authors pointed to spontaneous testicular tumors common in aged Fischer 344 rats: 19 of the 20 rats analyzed had enlarged testes and/or visible testicular tumors.
  • Fibrosis scores correlated best with the number of fibers about 10 μm or longer: R2 = .9677 at 1 year and .9632 at 2 years. The authors noted that the amosite fibers were longer than the Libby amphibole fibers.
• The authors' conclusion: The authors concluded that a single dose of Libby amphibole caused progressive chronic lung inflammation and significant dose-dependent fibrosis, but no histological evidence of carcinogenic effects in the lungs. Amosite also caused no marked carcinogenic effects in the lungs. At equal mass, amosite caused more inflammation and fibrosis, which the authors suggested means amosite was more toxic than Libby amphibole. They said fiber length may explain the difference.
• Limitations the authors state:
  • The study ended at about 22 months, when more than 50% of the saline control rats had died. Earlier studies ran up to the animals' lifetime (about 2.5 years). The authors said this shorter study might explain why no cancer was detected.
  • The study used fewer animals than earlier rat studies. Based on tumor rates in one such earlier amosite study, the authors estimated that fewer than 1 of 24 amosite-exposed rats would be expected to develop a lung tumor and/or mesothelioma.
  • Pleural tissues were not examined in detail. The authors said a closer examination might reveal mesotheliomas.
  • Spontaneous, strain-related tumors raised serum mesothelin in all groups. The authors said this may hide any increase caused by lung or pleural mesothelioma. The testicular tumors were not examined pathologically.
  • The gene-expression changes may not reflect protein levels in the tissue. They may instead reflect delayed responses or age-related variation between animals.
  • Dosing into the windpipe can cause fibers to clump, which does not happen with inhalation. The authors reported that no visible clumping was seen in lung tissue.

Mines named here

Asbestos mines this study is about. Each mine's page says who ran it, when it worked, and what it produced.

• Libby vermiculite mine, Montana

All asbestos mines

Topics

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

Medical terms in this study, explained in our Lexicon: Libby amphiboles, Amosite, Amphibole Group, vermiculite, Mesothelioma, Respirable Fraction, Inflammation, Fibrosis.

Related studies

• Comparative long-term toxicity of Libby amphibole and amosite asbestos in rats after single or multiple intratracheal exposures — J.M. Cyphert, D.J. Carlin, et al. (2015)
• Pulmonary inflammatory and fibrotic responses in Fischer 344 rats after intratracheal instillation exposure to Libby amphibole — D.J. Padilla-Carlin, M.C. Schladweiler, et al. (2011)
• Persistent effects of Libby amphibole and amosite asbestos following subchronic inhalation in rats — S.H. Gavett, C.U. Parkinson, et al. (2016)
• Sumas Mountain chrysotile induces greater lung fibrosis in Fischer344 rats than Libby amphibole, El Dorado tremolite, and Ontario ferroactinolite — J.M. Cyphert, A. Nyska, et al. (2012)
• Evaluation of anti-nuclear antibodies and kidney pathology in Lewis rats following exposure to Libby amphibole asbestos — K.D. Salazar, C.B. Copeland, et al. (2013)
• Vascular and thrombogenic effects of pulmonary exposure to Libby amphibole — J.H. Shannahan, M.C. Schladweiler, et al. (2012)

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