The health effects of short fiber chrysotile and amphibole asbestos

Figures & data

Figure 1. Typical plot of fiber number concentration against fiber length showing the skewed nature of the lognormal distribution.

Table

The reference datasets included:
Asbestiform riebeckite (crocidolite)	South Africa, Cape Province
Asbestiform riebeckite (crocidolite)	Australia, Wittenoom
Asbestiform grunerite (amosite)	South Africa, Transvaal
Na-Ca amphiboles (Libby amphiboles)	USA, Montana
While the validation datasets data from:
Asbestiform fluoro-edenite	Biancavilla, Italy
Non-asbestiform cummingtonite-grunerite (mineral fragments)	Homestake mine, Lead, South Dakota, USA
Asbestiform anthophyllite	Ekaterinburg, Russia

Figure 2. Meta-analysis of mesothelioma risk among workers involved with brake repair (reproduced from Garabrant et al. 2016).

Table 1. In vitro studies that differentiate the effects of short versus long fiber asbestos.

Cell culture model	Asbestos type	Fiber sizes	Exposure concentrations and duration	Type of assays	Results	Publication
Rat phagocytic ascetic tumor cells	AMOSITE; CHRYSOTILE; CROCIDOLITE	Ultrafine: 3 µm Fine: 8 µm Modified: 50 µm	30 µg/ml 27–216 × 10^6 fibers/ml 72 h	DNA and protein synthesis; cytotoxicity	It was possible to demonstrate clear-cut relationships between fiber length and cytotoxicity. At the 27 × 10^6 fibers/ml dose, the short fiber fraction (3 µm) was not different than the controls.	Tilkes and Beck 1980
Mouse peritoneal macrophages	AMOSITE; CHRYSOTILE; CROCIDOLITE NOTE: Same samples as used by Tilkes and Beck (1980)	UICC: ultrafine; medium; long	100 µg/1.5 × 10^6 cells 72 h	Erythrosin uptake, the release of lactate dehydrogenase (LDH), and uptake of 3H amino acids	Long-fiber asbestos dusts proved more cytotoxic than the corresponding short fibers when used on the basis of equal mass.	Kaw et al. 1982
Hamster tracheal organ culture	CHRYSOTILE, ANTIGORITE, the amorphous analogue of chrysotile	Chrysotile: Short: 100% <5 µm 59% < 1 µm Long: 0% <5 µm 50% > 30 µm Antigorite 100% < 3 µm 47% < 0.2 µm	1, 4, and 16 mg/ml 1 h	Assessment of metaplasia	While short fibers did not stimulate cell proliferation, long chrysotile fibers induced cell distortion and sloughing. Attapulgite (palygorskite) and both “long-” and “short-“ fiber preparations of chrysotile-asbestos had similar but less marked effects. Authors reported that the results suggest that the ability of inorganic particulates to induce squamous metaplasia depends upon their fibrous geometry and not on any particular chemical or structural property unique to asbestos.	Woodworth et al. 1983
Guinea pig lung macrophages	CHRYSOTILE; CROCIDOLITE	Chrysotile: Short Munroe: 0.4 9.99 µm Long Munroe: <3–80 µm Jeffrey 1: 1–19 µm Jeffery 2: 0.1–39 µm Crocidolite: Short: 0.1–19 µm Short S. African: 0.4–29 µm Long: 3–129 µm	100 µg/10^6 cells 20 h	Cytotoxicity; phagocytic activity	Long, very thin fibers of glass, chrysotile, crocidolite, and synthetic fluoro-amphiboles were all toxic in the test system	Tilkes and Beck 1983
Rat peritoneal macrophages	AMOSITE	Amosite: Short: 0.5% >5 µm Mean ∼ 0.5 µm 1.9 × 10^7 fibers/µg UICC: 9.5% >5 µm Mean ∼ 1.5 µm 6.2 × 10^6 fibers/µg Long: 31% >5 µm Mean ∼4 µm 2.3 × 10^6 fibers/µg	200 µg/ml with 3–4 × 10^6 macrophages/ml 48 or 96 h	Collagen protein synthesis	All fiber types induced production or release of fibrogenic factor, however, short fibers were more active than long fibers. The short fiber AM sample had ∼1000 fibers/macrophage while the long fiber samples had ∼100 fibers/macrophage	Aalto and Heppleston 1984
Syrian hamster embryo cells	CHRYSOTILE	Experiment 1: Length unmilled: 9.5 ± 0.7 µm Length milled: 1.7 ± 0.1 µm Experiment 2: Length unmilled: 16.0 ± 1.7 µm Length milled: 0.95 ± 0.12 µm	Asbestos-induced Cytotoxicity and Morphological Transformation: 0.25–2 µg/cm^27 days	Cytotoxicity; morphological transformation	After milling chrysotile asbestos in a mortar and pestle, its cytotoxic potency was greatly reduced. More importantly, milling of chrysotile asbestos completely eliminated its transforming potency at all doses tested.	Hesterberg and Barrett 1984
Hamster tracheal epithelial cells	UICC CHRYSOTILE UICC CROCIDOLITE RIEBECKITE, the non-fibrous mineral analogue of crocidolite ANTIGORITE, the amorphous analogue of chrysotile	Chrysotile Short: <2 µm Long: >10 µm Riebeckite ∼1–2 µm Antigorite ∼1–2 µm	0.36–11.6 µg/cm^2CH Short, Long and antigorite 24 h	Induction of ornithine decarboxylase: ODC activity	Maximal elevation of ODC activity was observed after exposure of cells to very low concentrations (0.72 µg/cm l dish) of long fibers, whereas a 4-fold higher concentration (2.9 µg/cm^2 dish) of short fibers was required to induce similar effects. Potency of ODC induction was directly related to fiber length. Riebeckite and antigorite did not increase ODC activity significantly.	Marsh and Mossman 1988
Peritoneal macrophages	CROCIDOLITE	Short: 1.1% > 5 µm (mean 1.2 µm) 4.64 × 10^9 fibers/mg Mixed: 8.8% > 5 µm (mean 2.8 µm) 2.93 × 10^9 fibers/mg Long: 27% >5 µm (mean 5.4 µm) 1.22 × 10^9 fibers/mg	Doses of 100, 300, 500, 700, and 1000 µg of long crocidolite correspond to 1.2, 3.7, 6.1, 8.5, and 12.2 (×10^8) fibers, respectively Doses of 100, 300, 500, 700, and 1000 µg of short crocidolite correspond to 4.6, 13.9, 23.2, 32.5, and 46.4 (×10^8) fibers, respectively Up to 22 h	Cell viability; H202 pro duction; mitochondrial membrane potential	The release of reactive oxygen species was stimulated by both long and short crocidolite; however, long fibers were found to be more toxic than short fibers. The fiber dose administered per cell for the short fibers ranged from 0.9 to 9 × 10^4 fibers/cell	Goodglick and Kane 1990
Rat alveolar macrophages	AMOSITE NOTE: The same fiber samples were used by Davis et al. (1986)	Short: 100% <10 µm Long: 40% >10 µm and 15–20% >20 µm	10, 25, 50 µg/ml Approximately 10–50 fibers/cell 24 h	Cytotoxicity; cell viability: presence of TNF in macrophage supernatants	Short fibers were not effective at stimulating release of TNF at any dose while high doses (50 µg/ml) of long fibers significantly increased the release of TNF following exposure to amosite.	Dogra and Donaldson 1995
Chinese hamster ovary cells	AMOSITE NOTE: The same fiber samples were used by Davis et al. (1986)	Short: 100% <10 µm Long: 40% >10 µm and 15–20% >20 µm	10 µg/ml of long or short amosite asbestos 48 h	Chromosomal aberrations, hyperploidy, and mitotic index	The incidence of all parameters in cells treated with the short amosite fiber sample was similar to control levels. Long amosite fibers significantly increased the incidence of chromosomal aberrations and hyperploidy. Cells exposed to long amosite also demonstrated significantly less mitotic index.	Donaldson and Golyasnya 1995
Rat alveolar macrophages	AMOSITE NOTE: The same fiber samples were used by Davis et al. (1986)	Short: 100% <10 µm Long: 40% >10 µm and 15–20% >20 µm	1 mg/ml 30 m	Assay of superoxide anions	A dramatic enhancement of super oxide anion release was observed with long, but not short amosite fibers following opsonization with immunoglobulin. Unopsonized short amosite seemed to inhibit superoxide release. Opsonization short fibers induced a small superoxide anion release above background. There was no obvious toxicity to the alveolar macrophages with any small fiber treatment, as assessed by the percentage of total cellular lactate dehydrogenase released into the culture medium during incubation.	Hill et al. 1995
Primate models human alveolar macrophages	CHRYSOTILE	UICC A Rhodesian: 75% <5 µm Unmilled RG 144 (Calidria) chrysotile: 98% <5 µm Milled RG 144 (Calidria) chrysotile: 99–100% <5 µm As milling time for RG 144 increased, the particle number per unit mass increased as well.	50–100 µg 24 h	Cytotoxicity	Viability of the cells following treatment was 67% for UICC chrysotile, 47% for un-milled RG 144 chrysotile, and 24–40% for milled RG 144 chrysotile. Doses were based on equal mass, thus the more the RG 144was milled the more particles were administered per cell. With milled RG 144, based on decisions and dose, there were at least 2,500 fibers administered per cell.	Yeager et al. 1983
Human type 2 alveolar epithelial cells	AMOSITE NOTE: The same fiber samples were used by Davis et al. (1986)	Short: 100% <10 µm Long: 40% >10 µm and 15–20% >20 µm	10, 25, 50, 75, and 100 µg/ml 4 h	Epithelial cell injury assay	The long amosite sample was found to cause a rapid detachment of the epithelial cells live from their substratum compared to the short fiber treatment.	Donaldson et al. 1993
Human lung epithelial cells	AMOSITE NOTE: The same fiber samples were used by Davis et al. (1986)	Short: 100% <10 µm Long: 40% >10 µm and 15–20% >20 µm	0.35, 3.5, and 35 µg/cm^2 24 h	Measurement of PPP activity; lipid peroxidation assay; cytotoxicity; enzyme activity assay	Long amosite fibers initiated free radical reactions, significantly inhibited glucose-6-phosphate dehydrogenase (G6PD) and PPP activity, decreased intracellular glutathione and increased thiobarbituric acid reactive substances and leakage of lactase dehydrogenase more effectively than short amosite. Surface reactivity: despite the higher surface exposed, short amosite did not exhibit significant activity in the homolytic cleavage of the carbon–hydrogen bond of format, while the activity of long amosite in the same conditions was very high.	Riganti et al. 2003
Human lung epithelial cells	AMOSITE NOTE: The same fiber samples were used by Davis et al. (1986)	Short: 100% <10 µm Long: 40% >10 µm and 15–20% > 20 µm	0.35–35 μg/cm^2 for LFA and SFA 24 h	NO synthesis and NO synthase (NOS) activity	LFA is more effective than SFA in causing activation of the NO synthesis in human lung epithelial cells after a 24 h incubation.	Tomatis et al. 2010
Macrophages: Human monocytic, monomac-6 (MM6) mouse macrophage (J774A.1) cell lines BAL cells Sprague–Dawley (SD) rats	AMOSITE NOTE: The same fiber samples were used by Davis et al. (1986)	Short: 100% <10 µm Long: 40% >10 µm and 15–20% >20 µm	7.5–125 µg/ml 4, 24, and 48 h	Cytotoxicity induced in J774A.1 cells was assessed via LDH release Pro-inflammatory mediators release	The morphology in J774A.1 cells following treatment with SFA was comparable to NPCB. Cell morphology was considerably different with exposure to LFA; with increased membrane ruffling associated with phagocytosis and increased lamellipodia from cells migrating toward and along fibers. LFA exposed cells were found to “spear” themselves along fibers that were too long for successful phagocytosis, while some cells attempted to phagocytose multiple LFA fibers simultaneously	Boyles et al. 2015

Table 2. Median diameter (and range) of the alveolar macrophage population and interstitial macrophage population for never smokers, smokers and individuals with COPD (from Dewhurst et al. 2017).

	Never smokers	Smokers	COPD	p-Value
IM diameter	13.2 (4.6–15.8)	11.5 (8.7–15.8)	11.7 (7.6–16.9)	0.0035
AM diameter	17.1 (2.9–27.4)	23.7 (14.1–25.6)	23.6 (13.4–25.7)	<0.0001
p-Value	<0.0001	<0.0001	<0.0001

Figure 3. Regression curve showing the relationship of Station criteria to tumor probability (reproduced from Wylie et al. 1987).

Figure 4. Differential paradigm of short and long fiber inhalation, deposition, and response.

Figure 5. Quantification of the numbers of chrysotile fibers observed as a function of time after a 5-day exposure in the airspace, bronchial and alveolar epithelium, interstitium, lymphatics, and alveolar macrophages (reproduced from Figure 11, Bernstein et al. 2004).

Table 3. In-vitro dosing and extrapolated fiber exposure in Huang (1979).

1.77E + 07	Number fiber in 1 µg	Density 2.4
	In vitro
314	Area in cm^2 Falcon cell dish 100 mm
10	µg/cm^2 applied in vitro
1.77E + 08	Total no fiber applied per cm^2
1.00E + 05	No of cells per dish
3.18E + 02	No cells/cm^2
5.55E + 05	No fibers/cell

Table 4. In-vitro dosing and extrapolated fiber exposure in Levresse et al. (2000).

	In vitro
75	Area in cm^2 cell dish
0.5	µg/cm^2 applied
9.75E + 07	Total no fiber applied per cm^2
1.00E + 05	No of cells per dish
1.33E + 03	No cells/cm^2
7.31E + 04	No fibers/cell

Garabrant DH, Alexander DD, Miller PE, Fryzek JP, Boffetta P, Teta MJ, Hessel PA, Craven VA, Kelsh MA, Goodman M. 2016. Mesothelioma among motor vehicle mechanics: an updated review and meta-analysis. ANNHYG. 60(8):1036–1026.

 Google Scholar

Tilkes F, Beck EG. 1980. Comparison of length-dependent cytotoxicity of inhalable asbestos and man-made mineral fibres. IARC Sci Publ. (Vol. 30):475–483.

 PubMedGoogle Scholar

Kaw JL, Tilkes F, Beck EG. 1982. Reaction of cells cultured in vitro to different asbestos dusts of equal surface area but different fibre length. Br J Exp Pathol. 63(1):109–115.

 PubMedGoogle Scholar

Woodworth CD, Mossman BT, Craighead JE. 1983. Interaction of asbestos with metaplastic squamous epithelium developing in organ cultures of hamster trachea. Environ Health Perspect. 51:27–33.

 PubMed Web of Science ®Google Scholar

Tilkes F, Beck EG. 1983. Sep Macrophage functions after exposure to mineral fibers. Environ Health Perspect. 51:67–72.

 PubMed Web of Science ®Google Scholar

Aalto M, Heppleston AG. 1984. Fibrogenesis by mineral fibers: an in-vitro study of the roles of the macrophage and fiber length. Br J Exp Pathol. 65(1):91–99.

 PubMedGoogle Scholar

Hesterberg TW, Barrett JC. 1984. Dependence of asbestos- and mineral dust-induced transformation of mammalian cells in culture on fiber dimension. Cancer Res. 44(5):2170–2180.

 PubMedGoogle Scholar

Marsh JP, Mossman BT. 1988. Mechanisms of induction of ornithine decarboxylase activity in tracheal epithelial cells by asbestiform minerals. Cancer Res. 48(3):709–714.

 PubMedGoogle Scholar

Goodglick LA, Kane AB. 1990. Cytotoxicity of long and short crocidolite asbestos fibers in vitro and in vivo. Cancer Res. 50(16):5153–5163.

 PubMed Web of Science ®Google Scholar

Davis JM, Addison J, Bolton RE, Donaldson K, Jones AD, Smith T. 1986. The pathogenicity of long versus short fiber samples of amosite asbestos administered to rats by inhalation and intraperitoneal injection. Br J Exp Pathol. 67(3):415–430.

 PubMed Web of Science ®Google Scholar

Dogra S, Donaldson K. 1995. Effect of long and short fibre amosite asbestos on in vitro TNF production by rat alveolar macrophages: the modifying effect of lipopolysaccharide. Ind Health. 33(3):131–141.

 PubMedGoogle Scholar

Donaldson K, Golyasnya N. 1995. Cytogenetic and pathogenic effects of long and short amosite asbestos. J Pathol. 177(3):303–307.

 PubMedGoogle Scholar

Hill IM, Beswick PH, Donaldson K. 1995. Differential release of superoxide anions by macrophages treated with long and short fibre amosite asbestos is a consequence of differential affinity for opsonin. Occup Environ Med. 52(2):92–96.

 PubMed Web of Science ®Google Scholar

Yeager H Jr., Russo DA, Yañez M, Gerardi D, Nolan RP, Kagan E, Langer AM. 1983. Cytotoxicity of a short-fiber chrysotile asbestos for human alveolar macrophages: preliminary observations. Environ Res. 30(1):224–232.

 PubMedGoogle Scholar

Donaldson K, Miller BG, Sara E, Slight J, Brown RC. 1993. Asbestos fiber length-dependent detachment injury to alveolar epithelial cells in vitro: role of a fibronectin-binding receptor. Int J Exp Pathol. 74(3):243–250.

 PubMedGoogle Scholar

Riganti C, Aldieri E, Bergandi L, Tomatis M, Fenoglio I, Costamagna C, Fubini B, Bosia A, Ghigo D. 2003. Long and short fiber amosite asbestos alters at a different extent the redox metabolism in human lung epithelial cells. Toxicol Appl Pharmacol. 193(1):106–115.

 PubMedGoogle Scholar

Tomatis M, Turci F, Ceschino R, Riganti C, Gazzano E, Martra G, Ghigo D, Fubini B. 2010. High aspect ratio materials: role of surface chemistry vs. length in the historical “long and short amosite asbestos fibers”. Inhal Toxicol. 22(12):984–998.

 PubMed Web of Science ®Google Scholar

Boyles MS, Young L, Brown DM, MacCalman L, Cowie H, Moisala A, Smail F, Smith PJ, Proudfoot L, Windle AH, et al. 2015. Multi-walled carbon nanotube induced frustrated phagocytosis, cytotoxicity and pro-inflammatory conditions in macrophages are length dependent and greater than that of asbestos. Toxicol In Vitro. 29(7):1513–1528.

 PubMed Web of Science ®Google Scholar

Dewhurst JA, Lea S, Hardaker E, Dungwa JV, Ravi AK, Singh D. 2017. Characterisation of lung macrophage subpopulations in COPD patients and controls. Sci Rep. 7(1):7143.

 PubMedGoogle Scholar

Wylie AG, Virta RL, Segreti JM. 1987. Characterization of mineral population by index particle: implication for the Stanton hypothesis. Environ Res. 43(2):427–439.

 PubMed Web of Science ®Google Scholar

Bernstein DM, Rogers R, Smith P. 2004. The biopersistence of Brazilian chrysotile asbestos following inhalation. Inhal Toxicol. 16(11–12):745–761.

 PubMed Web of Science ®Google Scholar

Huang SL. 1979. Amosite, chrysotile and crocidolite asbestos are mutagenic in Chinese hamster lung cells. Mutat Res. 68(3):265–274.

 PubMedGoogle Scholar

Levresse V, Renier A, Levy F, Broaddus VC, Jaurand M. 2000. DNA breakage in asbestos-treated normal and transformed (TSV40) rat pleural mesothelial cells. Mutagenesis. 15(3):239–244.

 PubMedGoogle Scholar