Silicon

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian germ cell study: gene mutation

Remarks:

Type of genotoxicity: gene mutation

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, meets generally accepted scientific principles, acceptable for assessment.

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

publication

Title:

Pulmonary chemokine and mutagenic responses in rats after subchronic inhalation of amorphous and crystalline silica.

Author:

Johnston, C. J., K. E. Driscoll, et al.

Year:

2000

Bibliographic source:

Toxicol Sci.56(2): 405-13.

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

In a subchronic inhalation study, rats were exposed 6h/d, 5 d/wk to 50 mg SiO2/m3 of hydrophilic pyrogenic silica (Aerosil® 200, MMAD 0.81 µm) or cristobalite (MMAD 1.3 μm) 3 mg/m3 for up to 13 weeks. After the exposure alveolar type II cells were isolated and HPRT mutations frequency was evaluated in vitro. Apoptosis was evaluated by TUNEL staining in histological lung sections.

GLP compliance:

no

Type of assay:

other: HPRT mutations

Test material

Test material information

Constituent 1

Reference substance name:

amorphous silica

IUPAC Name:

amorphous silica

Constituent 2

Reference substance name:

crystalline silica

IUPAC Name:

crystalline silica

Details on test material:

- crystalline silica: cristobalite, mass median aerodynamic diameter 1.3 μm; amorphous silica :hydrophilic pyrogenic Aerosil® 200 Degussa, mass median aerodynamic diameter 0.81 μm

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 200-250 g

Administration / exposure

Route of administration:

inhalation: dust

Vehicle:

no vehicle

Details on exposure:

TYPE OF INHALATION EXPOSURE: whole body

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Aerosols were generated by a screw-feed mechanism in combination with a venturi -type dust feeder

Controls were exposed to clean air.

Duration of treatment / exposure:

for up to 13 weeks

Frequency of treatment:

6 hours/day, 5 days/week

Post exposure period:

HPRT mutation frequences in alveolar type 2 cells and lung cell apoptosis were evaluated after 13 weeks of exposure

Doses / concentrations

Remarks:

Doses / Concentrations:
3 mg/m3 (crystalline silica); 50 mg/m3 (amorphous silica)
Basis:

No. of animals per sex per dose:

4 rats/treatment

Control animals:

yes, sham-exposed

Positive control(s):

quartz

Examinations

Tissues and cell types examined:

alveolar type II cells

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: exposure concentration was selected to cause significant inflammatory reaction in lungs
TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): see above
DETAILS OF SLIDE PREPARATION: after the termination of exposure, rat alveolar type II cells were isolated by standard methods and freshly isolated alveolar type II cells were seeded into culture flasks and the cells were allowed to attach overnight. The unattached cells were washed away and the cultures were fed every other day with a medium containing 6TG to select for mutation in HPRT gene. After 14-21 days in culture the cells were fixed and immunostained with an antibody to cytokeratins 8, 18, 19 and 6TG resistant cytokeratin staining colonies of >50 cells were counted.
METHOD OF ANALYSIS: Mutation frequaences were counted as (number of colonies/treatment)/(plating efficiency)/10E6 cells =mutants/10E6 cells

Statistics:

Dunnett's test

Results and discussion

Test results

Sex:

male

Genotoxicity:

negative

Toxicity:

yes

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Endpoints studied included mutation in the hprt gene of isolated alveolar cells in an ex vivo assay, changes in bronchoalveolar lavage (BAL) fluid markers of cellular and biochemical lung injury and inflammation, expression of mRNA for the chemokine MIP-2, and detection of apoptosis (TUNEL method). After 13 weeks of exposure, the percentage of lavage neutrophils, MIP-2 expression, and lactate dehydrogenase levels as an indicator of cytotoxicity were increased in both silicas. Histopathology of the lungs showed elevated levels of neutrophils and macrophages in lungs and TUNEL staining revealed increased apoptosis. Increased hprt mutation frequency in alveolar epithelial cells was detected with crystalline silica, but not with amorphous silica .

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
Repeated inhalation exposure to synthetic amorphous silica induced lung inflammation but not genotoxicity in rat lungs whereas crystalline silica caused a positive genotoxic response.

Executive summary:

Johnston et al. (2000) conducted a 90-day subchronic inhalation toxicity study with amorphous and crystalline silica. The exposure pattern followed OECD test 413 guidance 'Subchronic inhalation toxicity: 90-day study'. The GLP status was not mentioned. Male Fischer-344 rats (200–250 g) were exposed for 6 h/day, on 5 days/wk, for up to 13 weeks to 3 mg/m³crystalline (cristobalite, mass median aerodynamic diameter 1.3 μm) or 50 mg/m³amorphous silica (hydrophilic pyrogenic Aerosil® 200 Degussa, mass median aerodynamic diameter 0.81 μm). The genotoxic effects on the lung were characterized 13 weeks of exposure. Endpoints included mutation in thehprtgene of isolated alveolar cells in anex vivoassay, changes in bronchoalveolar lavage (BAL) fluid markers of cellular and biochemical lung injury and inflammation, expression of mRNA for the chemokine MIP-2, and detection ofapoptosis. After 13 weeks of exposure, the percentage of lavage neutrophils, MIP-2 expression, and lactate dehydrogenase levels as an indicator of cytotoxicity were increased in both silicas. All parameters remained increased for crystalline silica and decreased rapidly for amorphous silica during the 8-month recovery period. Increasedhprtmutation frequency in alveolar epithelial cells was detected with crystalline silica, but not with amorphous silica. Increased TUNEL staining indicative for apoptosis was seen in macrophages and terminal bronchiolar epithelial cells mainly after exposure to amorphous silica. Lung burdens of silica were 819 and 882 μg for crystalline and amorphous silica, respectively. In summary, genotoxic effects in alveolar epithelial cells occurred only after crystalline but not amorphous silica exposure, despite a high degree of inflammatory response after subchronic exposure to both types of silica. The authors suggest that the additional factors to inflammation, such as biopersistence of particles and direct or direct cytotoxicity to target cells, are important determinants of secondary genotoxic events.