Experimental sensitization to subtilisin. I. Production of immediate- and late-onset pulmonary reactions.

Four experiments were undertaken to explore the nature of pulmonary sensitivity which may result from inhalation of subtilisin aerosols. In the first study, immediate- and late-onset responses were identified. Groups of guinea pigs were exposed to atmospheres containing 0.15 to 15 mg/m3 bacterial subtilisin for 15 min/day on each of 5 consecutive days. Following an exposure-free period, reexposure to subtilisin at 1.9 mg/m3 for 20 min resulted in pulmonary responses characterized as "rapid shallow breathing." In some cases, reactions were severe and resulted in respiratory anaphylaxis; in one case, it was fatal. These responses usually occurred during the inhalation challenge or within 1 hr of challenge. However, late-onset reactions, maximum at 3-6 hr postchallenge, were occasionally observed. Results from the second series of experiments indicated that pulmonary responses were most prevalent in animals initially exposed to high concentrations of subtilisin (greater than 0.15 mg/m3) whereas no responses were observed in animals which had been exposed to 8.3 X 10(-3) or 41 X 10(-3) mg/m3 subtilisin. A third study indicated that exposure to 1.9 mg/m3 subtilisin for 20 min could result in pulmonary hypersensitivity. Additionally, when animals received subtilisin exposure on Days 1-5, pulmonary responses were equally severe whether elicited by inhalation challenge on Day 10 or on Day 17. Lastly, long-term exposure of animals to low levels of the enzyme did not result in any cases of pulmonary sensitivity even though the total cumulative exposure received by these animals when administered over a short period of time regularly induced sensitivity. The identification of a "threshold" concentration for sensitization in this animal model implies that it should be possible to recommend a safe industrial exposure level to subtilisin and thus prevent sensitization of exposed workers.

Immunotoxicologic evaluation of the respiratory system: animal models for immediate- and delayed-onset pulmonary hypersensitivity.

A wide variety of industrial materials is known to cause allergic pulmonary reactions. Respiratory symptoms may occur either immediately upon exposure to the agent (immediate-onset response), or several hours later (delayed-onset reaction). In order to determine both the mechanism of response, and a safe exposure level which would prevent development of pulmonary sensitization, animal models are currently being developed for both types of sensitivity responses. In the models, emphasis is placed on simulating conditions present in the industrial environment. For example, (1) exposure to agents is via the inhalation route, (2) animals are unrestrained and unanesthetized both during exposure and during elicitation of response, and (3) reactive chemicals, as opposed to hapten-protein conjugates, are used for exposure. In the model for immediate-onset sensitivity, concentration-response relationships have been observed between the concentration of agent inhaled and the percentage of animals becoming sensitized. Agents employed were bacterial subtilisin and toluene diisocyanate. Identification of "no observable effect" threshold concentrations implies that exposure levels can be proposed for industrial chemicals which will not result in sensitization of workers. Further development of the animal models, followed by calibration of the models to humans, should allow immunotoxicologic evaluation of chemicals for their respiratory sensitizing ability and recommendation of threshold-limit values (TLVs) which would prevent sensitization.