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Toxicological information

Specific investigations: other studies

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Administrative data

Endpoint:
mechanistic studies
Type of information:
experimental study
Adequacy of study:
other information
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well-conducted study that meets basic scientific principles, no details on test substance identity

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Neuronal modulation of lung injury induced by polymeric hexamethylene diisocyanate in mice
Author:
Lee, C.-T.; Poovey, H. G.; Rando, R. J. and Hoyle, G. W.
Year:
2007
Bibliographic source:
Toxicology and Applied Pharmacology 224: 19 - 28
Reference Type:
publication
Title:
Pulmonary Toxicity of Polymeric Hexamthylene Diisocyanate Aerosols in Mice
Author:
Lee, C.-T.; Friedman, M.; Poovey, H. G., Ie, S. R.; Rando, R. J., and Hoyle G. W.
Year:
2003
Bibliographic source:
Toxicology and Applied Pharmacology 188: 154 - 164

Materials and methods

Principles of method if other than guideline:
The role of sensory nerves in modulating lung injury following inhalation of the test subastance was assessed in genetically manipulated mice with altered innervation of the lung.
Type of method:
in vivo
Endpoint addressed:
acute toxicity: inhalation

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test substance (as cited in study report): 1,6-Hexamethylene diisocyanate biuret trimer (HDI-BT),
no further details on test substance and therefore analogy to registered substance can not be unequivically confirmed.
- Analytical purity: no data
- Physical state: liquid
- Impurities: residual monomeric HDI: no data

Test animals

Species:
mouse
Strain:
other: Tacr1 and CCSP-NGF (transgenic) and NGFR (knock-out)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- CCSP-NGF: transgenic mice with increased airway innervation by tachykinin-containing nerves (reference is given)
- NGFR (knock-out): knowckout mice with a mutation in the low-affinity nerve growth factor receptor gene; source: Kuo-Fen Lee, Massachusetts Institute of Technology, Cambridge.
Both strains were maintained on a C57BL76 background. Tacr1 transgenic mice were generated by microinjection of a CCSP-Tacr1 DNA construct into fertilized B6SJLF2 mouse eggs. Mice that developed from injected eggs were screened for the presence of the CCSP-Tacr1 transgene by dot blot of tail biopsy DNA with a rat CCSP probe. These mice were maintained on a B6SJLF2 background.

- Housing: in microisolator cages in a specific pathogen-free rodent facility (accredited by the Association for Assessment and Accreditation of Laboratory Animal Care.

Administration / exposure

Route of administration:
inhalation: aerosol
Vehicle:
acetone
Details on exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 0.6 x 0.9 x 1.2 m stainless-steel horizontal laminar flow exposure chamber.
- Exposure chamber volume: 648 L
- Method of holding animals in test chamber: in individual compartments of stainless-steel wire cages
- Source and rate of air: nebulizers were operated at a constant flow rate of 23.3 L/min dried compressed air, which allowed uniform aerosol size generation.
- System of generating particulates/aerosols: aerosols of the test substance were generated by nebulizing the test substance dissolved in acetone with DeVilbiss model 40 nebulizers.
- Method of particle size determination: with an Andersen 2000 cascade impactor

TEST ATMOSPHERE
- Brief description of analytical method used: samples were collected onto 37 mm glass-fiber filters saturated with a solution of 2 mg/mL 9-N-methyl-aminomethyl-anthracene (MAMA) with 3% tributyl phosphate for 30 min with an air-sampling pump.
- Samples taken from breathing zone: yes; 10 times during the 5 h exposure

VEHICLE
- Composition of vehicle (if applicable): acetone
- Concentration of test material in vehicle (if applicable): no data
Exposure of mice to a concentration of 2690 ppm acetone, a concentration equivalent to that received in the test substance exposure, had no effect on any of the indicators of lung injury that were measured.

TEST ATMOSPHERE
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): 1.42 µm/1.26
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Exposure levels were monitored by collecting samples form the chamber atmosphere throughout the exposures. Test substance levels were assessed by HPLC analysis following derivatization of samples with 9-N-methyl-aminomethyl-anthracene .
Duration of treatment / exposure:
5 h
Frequency of treatment:
single exposure
Post exposure period:
3.75 d (= 90 h)
Doses / concentrations
Remarks:
Doses / Concentrations:
10 and 25.5 mg/m³ air
Basis:
nominal conc.
No. of animals per sex per dose:
no data
Control animals:
yes, concurrent vehicle
Details on study design:
Mice were exposed to aerosol of the test substance for 5 h. Respiratory parameters after exposure were measured by barometric plethysmography in conscious unrestrained mice. Mice were euthanized and lavaged at mulitple time points after exposure. Cells recovered from the lavage fluid were counted. Protein concentration in the lavage fluid was measured. Cellular proliferation was assessed by BrdU incorporation. For RNase protection assay, mice were euthanized 0, 18 and 90 h after exposure and lung RNA was isolated. Furthermore, cAMP accumulation was measured in lung fragments in response to treatment.

Examinations

Examinations:
evaluation at 0, 18, 42 and 90 h after end of exposure

Results and discussion

Details on results:
NGFR knockout mice exhibited significantly more, and CCSP-NGF transgenic mice exhibited significantly less injury and inflammation compared with wild-type mice, indicative of a protective effect of nociceptive nerves on the lung following inhalation of the test substance. Transgenic mice overexpressing the tachykinin 1 receptor (Tacr1) in lung epithelial cells also showed less severe injury and inflammation compared with wild-type mice after test substance exposure, establishing a role for released tachykinins acting through Tacr1 mediating at least part of the protective effect. Treatment of lung fragments from Tacr1 transgenic mice with the Tacr1 ligand substance P resulted in increased cAMP accumulation, suggesting this compound as a possible signaling mediator of protective effects on the lung following nocicptive nerve stimulation. The results indicate that sensory nerves acting through Tacr1 can exert protective or anit-inflammatory effects in the lung following exposure.

Any other information on results incl. tables

Breathing parameters: For most parameters, no significant differences were observed among the unexposed CCSP-NGF transgenic, wild-type, and NGFR knockout mice. The one exception was that NGFR knowkout mice demonstrated a statistically significant 11 - 18% increase in enhanced pause (Penh) over wild-type and CCSP-NGF transgenic mice. NGFR knockout mice continued to exhibit test substance induced alterations 42 h after exposure when CCSP-NGF transgenic and wild-type mice did not show alterations anymore.

Applicant's summary and conclusion