Assessment of respiratory hypersensitivity in guinea pigs sensitized to toluene diisocyanate: A comparison of sensitization protocols

This study used a combined intradermal injection/repeated inhalation sensitization protocol in the guinea pig with subsequent inhalation challenge with toluene diisocyanate (TDI) and the homologous TDI-protein conjugate, immunoglobulin G(1) (IgG(1))-antibody analysis, and histopathological examination of the lung as a method to evaluate the potential of TDI to induce respiratory allergy in this animal model. In order to distinguish specific from nonspecific respiratory response, guinea pigs were subjected to an additional acetylcholine (ACh) bronchoprovocation assay, 1 day before and after the challenge with TDI. For comparison, groups of guinea pigs were sensitized to TDI by combined single intradermal injection (0.3% TDI in 100 mu l corn oil) and repeated inhalation exposure (3 h/day for 5 consecutive days) to 0, 3.8, 11, 26, 46, and 51 mg TDI/m(3) air. One group of animals was sensitized by intradermal injection only. Naive and polyisocyanate resin (50 and 250 mg dust/m(3) air) sensitized animals served as controls. Three weeks after the first encounter with the inducing agent, animals were challenged with the free TDI (similar to 0.5 mg/m(3)), and 1 wk later with the TDI-protein conjugate. Animals induced with the polyisocyanate resin were similarly challenged with the free chemical (similar to 50 mg/m(3)) and TDI- as well as resin-protein conjugates. The inhalation challenge with TDI failed to elicit pulmonary responses, while during the TDI-protein conjugate challenge characteristic changes in breathing patterns occurred. Animals sensitized to higher concentrations of TDI by inhalation displayed increased responsiveness to ACh and an influx of eosinophilic granulocytes in airways or lung-associated lymph nodes (LALN). Guinea pigs of the TDI resin groups did not respond to any challenge, nor was there any influx of eosinophils in airways or LALN; that is, they were indistinguishable from naive controls. IgG, antibodies were observed in all groups receiving TDI or TDI resin. In summary, it could be demonstrated that the intradermal injection, in addition to the 5 x 3 h inhalation exposures, did not increase markedly the sensitivity of this animal model. However, the outcome of test appeared to be less dependent on the exposure concentration used for sensitization of the animals when the combined protocol was used. Therefore, it is believed that the combined induction protocol improves the robustness of this animal model. Accordingly, in order to classify a low-molecular-weight substance as a respiratory sensitizer, a triad of responses should be considered: (1) positive respiratory response upon challenge with the hapten, and if negative, also challenge with the conjugate of the hapten, (2) an influx of eosinophilic granulocytes, and (3) increased specific IgG(1) response.