IL-17F, rather than IL-17A, underlies airway inflammation in a steroid-insensitive toluene diisocyanate-induced asthma model.

Steroid insensitivity constitutes a major problem for asthma management. Toluene diisocyanate (TDI) is one of the leading allergens of asthma that induces both T-helper Th2 and Th17 responses, and is often associated with poor responsiveness to steroid treatment in the clinic.We sought to evaluate the effects of inhaled and systemic steroids on a TDI-induced asthma model and to find how interleukin (IL)-17A and IL-17F function in this model. BALB/c mice were exposed to TDI for generating an asthma model and were treated with inhaled fluticasone propionate, systemic prednisone, anti-IL-17A, anti-IL-17F, recombinant IL-17A or IL-17F.Both fluticasone propionate and prednisone showed no effects on TDI-induced airway hyperresponsiveness (AHR), bronchial neutrophilia and eosinophilia, and epithelial goblet cell metaplasia. TDI-induced Th2 and Th17 signatures were not suppressed by fluticasone propionate or prednisone. Treatment with anti-IL-17A after TDI exposure led to increased AHR, aggravated mucus production and airway eosinophil recruitment, accompanied by amplified Th2 responses, whereas anti-IL-17F ameliorated TDI-induced AHR and airway neutrophilia, with decreased Th17 responses. Recombinant IL-17A and IL-17F showed opposite effects to the monoclonal antibodies.IL-17A and IL-17F exert distinct biological effects during airway inflammation of a TDI-induced asthma model, which is unresponsive to both inhaled and systemic steroids.

Critique of the ACGIH 2016 derivation of toluene diisocyanate Threshold Limit Values.

In 2016, the American Conference of Governmental Industrial Hygienists (ACGIH) lowered the 8-hr Threshold Limit Value - time-weighted average (TLV-TWA) for toluene diisocyanate (TDI) from 5 ppb to 1 ppb, and the 15-min short-term exposure limit (STEL) from 20 ppb to 5 ppb. We evaluated ACGIH's basis for lowering these values. It is our opinion that the ACGIH's evaluation of the evidence for occupational asthma and respiratory effects from TDI exposure does not fully integrate the results of all the available human and animal studies. We found that some studies reported occupational asthma cases at TWAs less than 5 ppb, but these cases were likely caused by peak exposures above 20 ppb. Advances in industrial hygiene have reduced peak exposures and the incidence of upset conditions, such as spills and accidents, in modern TDI facilities. Taken together, the human evidence indicates that adherence to the previous 8-hr TLV-TWA and 15-min STEL (5 ppb and 20 ppb, respectively) prevents most, if not all, cases of occupational asthma, and eliminates or reduces the risk of lung function decrements and other respiratory effects. While limited, the animal literature supports the human evidence and indicates that TDI-induced asthma is a threshold phenomenon. We conclude that ACGIH's decision to lower the TLV-TWA and STEL values for TDI is not adequately supported.

Hypothesis-based weight-of-evidence evaluation of the human carcinogenicity of toluene diisocyanate.

Humans are exposed to toluene diisocyanate (TDI) primarily through inhalation in workplaces where TDI is produced or used. It is classified as a possible human carcinogen, based primarily on increased tumor incidences in rodents treated with TDI by oral gavage. We used the hypothesis-based weight-of-evidence (HBWoE) method to evaluate whether the available data support the hypothesis that TDI is a human carcinogen. The epidemiology data are not sufficiently robust to support TDI as a human carcinogen; the few positive associations are more likely attributable to alternative explanations than causation. The experimental animal studies indicate that inhalation exposure to TDI does not induce tumors in rats or mice. Tumors observed after oral gavage exposure are most likely due to the conversion of approximately 5% of the administered TDI to toluene diamine (TDA), a known rodent tumorigen. This contention is supported by the observations that TDA is rapidly formed from TDI during in vitro genotoxicity assays, the spectra of responses to TDA and TDI in these assays and in oral bioassays are essentially the same, and TDI is not genotoxic in rodents or humans in vivo after inhalation exposure, when TDA is not formed to a biologically significant degree. We conclude that the weight of the evidence indicates that the conversion of TDI to TDA does not occur in mammalian species under physiological exposure conditions (i.e. inhalation), but is necessary for carcinogenesis to occur. Thus, a causal association between TDI exposure and carcinogenic effects is not plausible in humans.

Micronuclei, hemoglobin adducts and respiratory tract irritation in mice after inhalation of toluene diisocyanate (TDI) and 4,4'-methylenediphenyl diisocyanate (MDI).

Toluene diisocyanate (TDI) and 4,4'-methylenediphenyl diisocyanate (MDI), used in the production of polyurethane foam, are well known for their irritating and sensitizing properties. Contradictory results have been obtained on their genotoxicity. We investigated the genotoxicity and protein binding of inhaled TDI and MDI in mice by examining micronucleated polychromatic erythrocytes (PCEs) in bone marrow and peripheral blood and TDI- and MDI-derived adducts in hemoglobin. Male C57Bl/6J mice (8 per group) were exposed head-only to TDI vapour (mean concentrations 1.1, 1.5, and 2.4mg/m(3); the mixture of isomers contained, on the average, 63% 2,4-TDI and 37% 2,6-TDI) or MDI aerosol (mean concentrations 10.7, 20.9 and 23.3mg/m(3)), during 1h/day for 5 consecutive days. Bone marrow and peripheral blood were collected 24h after the last exposure. Inhalation of TDI caused sensory irritation (SI) in the upper respiratory tract, and cumulative effects were observed at the highest exposure level. Inhalation of MDI produced SI and airflow limitation, and influx of inflammatory cells into the lungs. Hemoglobin adducts detected in the exposed mice resulted from direct binding to globin of 2,4- and 2,6-TDI and MDI, and dose-dependent increases were observed especially for 2,4-TDI-derived adducts. Adducts originating from the diamines of TDI (toluene diamine) or MDI (methylene dianiline) were not observed. No significant increase in the frequency of micronucleated PCEs was detected in the bone marrow or peripheral blood of the mice exposed to TDI or MDI. The ratio of PCEs and normochromatic erythrocytes (NCEs) was reduced at the highest concentration of MDI, and a slight reduction of the PCE/NCE ratio, dependent on cumulative inhaled dose, was also seen with TDI. Our results indicate that inhalation of TDI or MDI (1h/day for 5 days), at levels that induce toxic effects and formation of TDI- or MDI-specific adducts in hemoglobin, does not have detectable genotoxic effects in mice, as studied with the micronucleus assay.

Rats repeatedly exposed to toluene diisocyanate exhibit immune reactivity against methyl isocyanate-protein conjugates.

BACKGROUND: Volatile monoisocyanates are formed through thermal degradation when products containing polyurethane are heated. Repeated exposure to diisocyanates, such as toluene diisocyanate (TDI) are a well-known cause of occupational asthma. However, although monoisocyanates are abundant in occupational settings, there are few data concerning their ability to provoke immune reactions and asthma. We compared immune reactivity and respiratory disease following single or repeated inhalation exposures to the monoisocyanates methyl isocyanate (MIC) and isocyanic acid (ICA) with the effects of TDI. METHODS: Isocyanates were administrated either as single vapor exposures or as repeated intranasal instillations in rats. Adverse health effects were monitored by analyzing airway inflammation, respiratory function and weight gain. Immune reactivity caused by repeated exposures was studied by analysis of isocyanate-specific antibodies and airway infiltration of immune competent cells. RESULTS: Repeated exposures to TDI induced airway infiltration of neutrophils and lymphocytes, while neither MIC nor ICA provoked a detectable inflammatory response. Antibodies against isocyanate-albumin conjugates were detected in serum after both exposures to TDI and MIC, but not to ICA. TDI-exposed rats also displayed IgG antibodies against MIC-albumin conjugates. Even though MIC did not induce airway inflammation, single exposure provoked an increase in airway resistance and repeated exposures caused weight loss similar to that of TDI. CONCLUSIONS: Airway exposure to TDI produces an antibody response not only against TDI but also against MIC-protein conjugates. This indicates that immune reactivity against abundant monoisocyanates in occupational environments can occur in individuals pre-sensitized with low abundance but highly sensitizing diisocyanates.

Oropharyngeal aspiration: an alternative route for challenging in a mouse model of chemical-induced asthma.

BACKGROUND: To assess the importance of the route of challenge in an existing mouse model of chemical-induced asthma, we replaced intranasal instillation by oropharyngeal aspiration. To our knowledge, oropharyngeal aspiration as a challenge route has not yet been investigated in a mouse model of chemical-induced asthma. METHODS: On days 1 and 8, mice were dermally sensitized with toluene diisocyanate (TDI) (0.3%) [or vehicle (acetone/olive oil)] and on day 15 they received a single challenge, via oropharyngeal aspiration, with TDI (0.01%) or vehicle. One day after challenge, airway reactivity to methacholine was measured by a forced oscillation technique (FlexiVent) and total and differential cell counts, as well as levels of KC, IL-5, IL-17 and TNF-alpha, were assessed in the bronchoalveolar lavage (BAL) fluid. Lymphocytes from the auricular and mediastinal lymph nodes were cultured to determine the concanavaline A-induced secretion of IL-2, IL-4, IL-10, IL-13, IL-17 and IFN-gamma. Total serum IgE was measured. RESULTS: In TDI-sensitized mice, a significant increase in airway reactivity was found after a single oropharyngeal challenge with TDI. BAL neutrophils and eosinophils were increased 7- and 5-fold, respectively. An upregulation of Th1 (IFN-gamma), Th2 (IL-4, IL-10, IL-13) and Th17 (IL-17) cytokines was found in the auricular lymph nodes, in the mediastinal lymph nodes only IL-4 was upregulated. The total serum IgE level in TDI-sensitized mice was significantly increased when compared to control mice. CONCLUSION: We conclude that challenging mice via oropharyngeal aspiration mimics the characteristics of human asthma well, without the possible drawbacks of other techniques.

How long do the systemic and ventilatory responses to toluene diisocyanate persist in dermally sensitized mice?

BACKGROUND: Years after removal from exposure, workers with occupational asthma still show respiratory symptoms and airway hyperresponsiveness on re-exposure to the offending agent. OBJECTIVE: We investigated the persistence of the respiratory responsiveness to toluene diisocyanate (TDI) in a mouse model. METHODS: BALB/C mice received dermal applications of TDI on days 1 and 8, and a single intranasal instillation of TDI on day 10, 15, 20, 25, 30, 40, 50, 60, or 90. After instillation, early (1 hour) changes in ventilatory function and methacholine responsiveness (22 hours) were assessed. Cell counts and macrophage inflammatory protein 2 were measured in bronchoalveolar lavage. Total serum IgE, IgG(1), and IgG(2a) were quantified. Lymphocyte subpopulations were assessed in auricular and cervical lymph nodes, and release of IL-4 and IFN-gamma by these lymph node cells was measured. RESULTS: Toluene diisocyanate-treated mice showed immediate ventilatory changes, increased methacholine reactivity, and an influx of neutrophils and macrophage inflammatory protein 2 in bronchoalveolar lavage as long as 50 days after initial treatment. These mice also showed a relative increase in CD19(+) cells and a decrease in CD4(+) and CD8(+) cells in auricular lymph nodes. Increased release of IL-4 and IFN-gamma in auricular lymph node cells was observed only until 20 days after sensitization. Total serum IgE, IgG(1), and IgG(2a) remained significantly elevated in TDI-sensitized mice until 90 days after dermal sensitization. CONCLUSION: Ventilatory and lung inflammatory responses decrease with increasing delay between sensitization and challenge, despite persistent humoral signs of sensitization.

Dermal exposure determines the outcome of repeated airway exposure in a long-term chemical-induced asthma-like mouse model.

BACKGROUND: Exposure to diisocyanates is an important cause of occupational asthma (OA) in the industrialized world. Since OA occurs after long-term exposure to diisocyanates, we developed a chronic mouse model of chemical-induced asthma where toluene diisocyanate (TDI) was administered at two different exposure sites. OBJECTIVES: Evaluating the effect of long-term respiratory isocyanate exposure - with or without prior dermal exposure- on sensitization, inflammatory responses and airway hyperreactivity (AHR). METHODS: On days 1 and 8, BALB/c mice were dermally treated (20 µl/ear) with 0.5% 2,4-toluene diisocyanate TDI or the vehicle acetone olive oil (AOO) (3:2). Starting from day 15, mice received intranasal instillations with 0.1% TDI of vehicle five times in a week, for five successive weeks. One day after the last instillation airway hyperreactivity (AHR) to methacholine was assessed, followed by an evaluation of pulmonary inflammation and structural lung changes. Immune-related parameters were assessed in the lungs (BAL and tissue), blood, cervical- and auricular lymph nodes. RESULTS: Mice repeatedly intranasally exposed to TDI showed systemic sensitization and a mixed Th1/Th2 type immune response, without the presence of AHR. However, when mice are first dermally sensitized with TDI, followed by repeated intranasal TDI challenges, this results in a pronounced Th2 response and AHR. CONCLUSION: Dermal exposure to TDI determines airway hyperreactivity after repeated airway exposure to TDI.

Contact sensitization to 4,4'-diaminodiphenylmethane and to isocyanates among general dermatology patients.

BACKGROUND: Diisocyanates and 4,4'-diaminodiphenylmethane (MDA) are industrial sensitizers. Occupational asthma is a risk among workers exposed to diisocyanates. Exposure may also lead to contact sensitization and allergic contact dermatitis. OBJECTIVE: The aim of this study was to determine the occurrence of contact sensitization to MDA and to diisocyanates among general dermatology patients. PATIENTS AND METHODS: Patch testing with MDA was carried out in 1595 patients. Diphenylmethane-4,4'-diisocyanate (MDI) and toluene-2,4-diisocyanate (TDI) were tested in 1023 patients and isophorone diisocyanate (IPDI) and 1,6-hexamethylene diisocyanate (HDI) in 433 patients. The clinical data and sources of exposure are analysed. RESULTS: MDA reactions were seen in 17 (1.1%) patients and MDI reactions in 4 patients. Six MDA-positive patients reacted to p-phenylenediamine and two to epoxy chemicals. 5/10 of the TDI reactions were seen concurrently with reactions to MDI, MDA, HDI, or to IPDI. IPDI reactions were seen in eight patients and HDI reactions in two patients. Possible sources of exposure were traced in most patients, although the association with the current dermatitis was not apparent in all cases. CONCLUSION: (Di)isocyanates may induce contact sensitization with or without allergic contact dermatitis.

Comparative studies of lymph node cell subpopulations and cytokine expression in murine model for testing the potentials of chemicals to induce respiratory sensitization.

OBJECTIVES: To investigate immunological changes in lymph nodes based on expression of cell-specific receptors and cytokine expression profile and accompanying inflammatory reactions in lungs of mice treated with chemicals of known potentials to induce respiratory sensitization and those in which activity in this regard is unclear. MATERIALS AND METHODS: On day 1 and 7, Balb/c mice received toluene-2,4-diisocyanate (TDI), trimellitic anhydride (TMA), 1-chloro-2,4-dinitrobenzene (DNCB), glutaraldehyde (GA), formaldehyde (FA), benzalkonium chloride (ChB) or vehicle. On day 14, they received a single intranasal instillation with the same chemical or vehicle. On day 15, auricular lymph nodes (LN) were excised and used for analyzes of T-, B-cells, expression of CD44 and for the estimation of IL-4 and IFN-gamma production after in vitro stimulation with concanavalin A (ConA) and also for IL-4 and IFN-gamma mRNA expression analyses using Real-Time PCR. Inflammatory changes in lungs were observed by estimation of TNF-alpha and MIP-2 concentrations and cell numbers and their type in BAL. RESULTS: There were no significant changes in cell subpopulations of T helper cells in LN. The percent of B cells was significantly increased after treatment with DNCB, TDI, and GA. Increased expression of CD44 on T cells was also observed. Both IL-4 and IFN-gamma were found increased in TDI- and FA-treated mice, while only IL-4 was increased in TMA-treated mice. Real-Time PCR analyses, however, showed increased IL-4 mRNA expression for TDI- and TMA-, and IFN-gamma mRNA expression for DNCB-treated mice. We haven't observed significant changes in inflammatory reactions in the lungs of exposed animals. CONCLUSIONS: Studying immunological changes with first determining the activation status of T cells followed by analyzes of expression of mRNA for Th1 and Th2 cytokines in murine model could be a useful method for assessment of the potentials of chemicals to induce respiratory sensitization but is not sufficient. Addition of ventilatory measurements, but not necessarily inflammatory reactions, could complete the model.

Maternal allergic contact dermatitis causes increased asthma risk in offspring.

BACKGROUND: Offspring of asthmatic mothers have increased risk of developing asthma, based on human epidemiologic data and experimental animal models. The objective of this study was to determine whether maternal allergy at non-pulmonary sites can increase asthma risk in offspring. METHODS: BALB/c female mice received 2 topical applications of vehicle, dinitrochlorobenzene, or toluene diisocyanate before mating with untreated males. Dinitrochlorobenzene is a skin-sensitizer only and known to induce a Th1 response, while toluene diisocyanate is both a skin and respiratory sensitizer that causes a Th2 response. Both cause allergic contact dermatitis. Offspring underwent an intentionally suboptimal protocol of allergen sensitization and aerosol challenge, followed by evaluation of airway hyperresponsiveness, allergic airway inflammation, and cytokine production. Mothers were tested for allergic airway disease, evidence of dermatitis, cellularity of the draining lymph nodes, and systemic cytokine levels. The role of interleukin-4 was also explored using interleukin-4 deficient mice. RESULTS: Offspring of toluene diisocyanate but not dinitrochlorobenzene-treated mothers developed an asthmatic phenotype following allergen sensitization and challenge, seen as increased Penh values, airway inflammation, bronchoalveolar lavage total cell counts and eosinophilia, and Th2 cytokine imbalance in the lung. Toluene diisocyanate treated interleukin-4 deficient mothers were able to transfer asthma risk to offspring. Mothers in both experimental groups developed allergic contact dermatitis, but not allergic airway disease. CONCLUSION: Maternal non-respiratory allergy (Th2-skewed dermatitis caused by toluene diisocyanate) can result in the maternal transmission of asthma risk in mice.

Correlation of bacterial mutagenicity and hamster cell transformation with tumorigenicity induced by 2,4-toluenediamine.

In the presence of rat liver microsome enzymes, 2,4-toluenediamine (TDA) was mutagenic for several tester strains of Salmonella typhimurium. TDA induced morphological transformation in an in vitro carcinogenesis system using secondary culture target cells prepared from cryopreserved primary Syrian hamster embryo cells. These results now correlate bacterial mutagenicity and in vitro morphological transformation with the reported tumorigenicity of this compound.

Respiratory response to toluene diisocyanate depends on prior frequency and concentration of dermal sensitization in mice.

Occupational asthma is the principal cause of work-related respiratory disease in the industrial world. In the absence of satisfactory models for predicting the potential of low molecular weight chemicals to cause asthma, we verified that dermal sensitization prior to intranasal challenge influences the respiratory response using toluene diisocyanate (TDI), a known respiratory sensitizer. BALB/c mice received TDI or vehicle (acetone/olive oil) on each ear on three consecutive days (days 1, 2, and 3; 0.3 or 3% TDI) or only once (day 1, 1% TDI). On day 7, the mice received similar dermal applications of vehicle or the same concentration of TDI as before ("boost"). On day 10, they received an intranasal dose of TDI (0.1%) or vehicle. Ventilatory function was monitored by whole body plethysmography for 40 min after intranasal application, and reactivity to inhaled methacholine was assessed 24 h later. Pulmonary inflammation was assessed by bronchoalveolar lavage and histology. Mice that received an intranasal dose of TDI without having received a prior dermal application of TDI did not exhibit any ventilatory response or inflammatory changes compared to vehicle controls. In contrast, mice that had received prior application(s) of TDI, even if only on day 7, exhibited the following: ventilatory responses, compatible with bronchoconstriction, immediately after intranasal application with TDI; enhanced methacholine responsiveness 24 h later; and pulmonary inflammation characterized by neutrophils. This was, however, not the case in mice that received the highest dermal amount of TDI (3% on days 1, 2, and 3). These findings suggest that respiratory response to TDI depends on prior frequency and concentration of dermal sensitization in mice.

Developmental toxicity evaluation of inhaled toluene diisocyanate vapor in CD rats.

Mated female CD (Sprague-Dawley) rats, 25/group, were exposed to toluene diisocyanate (TDI) vapor, for six h/day on gestational days (gd) 6 through 15, at 0.00, 0.02, 0.10, or 0.50 p.p.m.. Maternal clinical signs, body weights, and feed and water consumption were recorded throughout gestation. At termination (gd 21), maternal body, gravid uterine, and liver weights were recorded. Corpora lutea were counted, and implantation sites were identified: resorptions and dead and live fetuses. All live fetuses were examined for external alterations. One-half of the live fetuses/litter were examined for visceral (including craniofacial) alterations. The remaining intact fetuses/litter were stained with alizarin red S and examined for ossified skeletal alterations. Maternal toxicity at 0.50 ppm consisted of reduced body weights, body weight gains, feed consumption, and clinical signs of toxicity. Water consumption was unaffected. Gestational parameters exhibited no significant treatment-related changes, including pre- and postimplantation loss, sex ratio/litter, or fetal body weights/litter. Incidences of individual malformations, malformations by category (external, visceral, and skeletal), total malformations, individual external and visceral variations, variations by category, and total variations were unaffected. Of 111 skeletal variants observed, only 1, incidence of poorly ossified cervical centrum 5, was increased at 0.50 ppm, indicating possible minimal fetotoxicity, although it occurred in the absence of any other indications of developmental toxicity. Therefore, exposure to TDI vapor by inhalation, during major organogenesis in CD rats, resulted in maternal toxicity and minimal fetotoxicity at 0.50 ppm no observed adverse effect level (NOAEL) for maternal and developmental toxicity was 0.10 ppm. No treatment-related embryotoxicity or teratogenicity was observed.

Validity of methods to predict the respiratory sensitizing potential of chemicals: A study with a piperidinyl chlorotriazine derivative that caused an outbreak of occupational asthma.

A piperidinyl chlorotriazine (PCT) derivative, used as a plastic UV-stabilizer, caused an outbreak of occupational asthma. We verified, in BALB/c mice, the sensitizing potential of PCT in comparison to a known respiratory sensitizer (toluene diisocyanate [TDI]) and a known dermal sensitizer (oxazolone), using three different methods in order to evaluate the validity of current models of sensitization. These included the local lymph node assay (LLNA) and the mouse IgE test. In addition, respiratory hyper-reactivity was assessed following a novel protocol involving dermal sensitization (20 microl of a 3% solution on each ear for three days) and intranasal challenge (0.1% or 1%, 10 microl per nostril on day 10), followed, after 24 h, by a methacholine challenge (using whole-body plethysmography), bronchoalveolar lavage, and histology. PCT was also used for structure-activity relationship (SAR) models for (respiratory) sensitization. High concentrations of PCT (10 and 20%) resulted in significant responses in the local lymph node assay (LLNA; stimulation indices (SI) of 2.7 +/- 0.9 and 3.2 +/- 0.6, respectively). The mouse IgE test was positive with 20% PCT only. Methacholine responsiveness was increased only in previously sensitized mice receiving a challenge with TDI or PCT. However, there was no evidence for pulmonary inflammation. The SAR studies indicated that PCT could be a respiratory sensitizer. Based on an approved test protocol such as the LLNA and the mouse IgE test, PCT proved to be a weak sensitizer when compared to TDI and oxazolone. However, in a protocol involving an intranasal challenge, PCT appeared to be a respiratory sensitizer of similar potency to TDI.

Two-generation reproductive toxicity study of inhaled toluene diisocyanate vapor in CD rats.

Twenty-eight 42-day-old pups/sex/group (F0) were exposed to toluene diisocyanate vapor (TDI; 80% 2,4-TDI, 20% 2,6-TDI) by inhalation at 0.0, 0.02, 0.08, or 0.3 ppm, 6 h/day, 5 days/week, for 10 weeks, then mated within groups for 3 weeks, with exposure 7 days/week during mating, gestation, and lactation. F0 maternal animals were not exposed from gestational day (gd) 20 through postnatal day (pnd) 4; maternal exposures resumed on pnd 5. Twenty-eight weanlings/sex/group continued exposure for 12 weeks (starting on pnd 28) and were bred as described above. F0 and F1 parents and ten F1 and F2 weanlings/sex/group were necropsied, and adult reproductive organs, pituitary, liver, kidneys, and upper respiratory tract (target organs) were evaluated histologically in ten/sex/group. Adult toxicity was observed in both sexes and generations at 0.08 and 0.3 ppm, including occasional reductions in body weights and weight gain, clinical signs of toxicity at 0.08 and 0.3 ppm, and histologic changes in the nasal cavities at 0.02, 0.08, and 0.3 ppm (including rhinitis, a nonspecific response to an irritating vapor, at all concentrations). There was no reproductive toxicity, reproductive organ pathology, or effect on gestation or lactation at any exposure concentration. Postnatal toxicity and reduced body weights and weight gains during lactation occurred only in F2 litters at 0.08 and 0.3 ppm. Therefore, under the conditions of this study, a no observed adverse effect level (NOAEL) was not determined for adult toxicity; the NOAEL for reproductive toxicity was at least 0.3 ppm, and the NOAEL for postnatal toxicity was 0.02 ppm.

Pulmonary alterations associated with inhalation of occupational and environmental irritants.

Many gases, vapors, or particles found in occupational and/or environmental settings can act as irritants. In the present study, sensory irritants are characterized by the stimulation of neuropeptide release from sensory nerves in the nasal mucosa, while pulmonary irritants are characterized by recruitment of PMN into bronchoalveolar airspaces, elevation of breathing frequency, and neuropeptide release from sensory fibers innervating the epithelium of the conducting airways. A review of data from our laboratory as well as results from others indicate that asphalt fume is a sensory irritant; toluene diisocyanate (TDI), methyl isocyanate, and machining fluid act as both sensory and pulmonary irritants; while cotton dust, agricultural dusts, microbial products, leather conditioner, and ozone exhibit responses characteristic of pulmonary irritants.

Bronchial responsiveness and inflammation in guinea-pigs exposed to toluene diisocyanate: a study on single and repeated exposure.

The question of whether or not toluene diisocyanate (TDI)-induced airway hyperresponsiveness in the guinea-pig is accompanied by neutrophil influx into bronchoalveolar lavage fluid (BALF) was addressed. Two modes of exposure were studied; (1) acute exposures where animals were exposed to 3 ppm TDI for 1 h and experiments were carried out 30 min, 4 h, 24 h, 48 h and 1 week after the TDI exposures; (2) subacute exposures where animals were exposed to 0.080 and 0.046 ppm TDI for 48 h 1 week, respectively, and experiments were carried out 24 h after the TDI exposures. The changes in airway responsiveness to increasing doses of intravenous acetylcholine (ACh) in anaesthetized and tracheotomized spontaneously breathing guinea-pigs were examined. In order to elucidate the possible relationships of airway responsiveness to cellular infiltration, bronchoalveolar lavage was performed in additional group of guinea-pigs exposed to the same conditions. After acute exposure to 3 ppm TDI, increased bronchial responsiveness was evident within 30 min, lasted 48 h, but had vanished 1 week after the exposure. An influx of neutrophils occurred into the BALF within 1 h after exposure. The influx of neutrophil into BALF lasted 48 h and vanished 1 week after the end of exposure. After 48 h of exposure to TDI at 0.080 ppm, or 0.046 ppm for 1 week, increased bronchial responsiveness was evident 24 h after the end of the both modes of exposure, but no influx of neutrophils was observed into the BALF. It was concluded that even though the neutrophil influx and hyperresponsiveness evolve in the same way after acute exposure to a high concentration of TDI (3 ppm), this is not the case after subchronic exposure to low concentrations of TDI, where a bronchial hyperresponsiveness is observed without detectable neutrophil influx.

A two-centre study for the evaluation and validation of an animal model for the assessment of the potential of small molecular weight chemicals to cause respiratory allergy.

This study evaluated a single intradermal injection model in the guinea pig with subsequent inhalation challenge and serological analysis as a method to predict the potential of chemicals to induce respiratory allergy. Four known respiratory allergens (trimellitic anhydride, diphenyl methane diisocyanate, phthalic anhydride and toluene diisocyanate (TDI)) were screened by two industrial research laboratories using this protocol. Dinitrochlorobenzene, a potent contact allergen, was included as a negative control material. In both laboratories, the respiratory allergens, but not the contact allergen, induced high titre antigen-specific antibodies in treated animals. The inhalation challenge results were similar in both laboratories but were less conclusive in that exposure to free TDI failed to induce pulmonary responses, probably because it fails to penetrate to the deep lung in sufficient concentration. Although the assay shows promise as a means of identifying chemical respiratory sensitisers, its use as a routine screen for the prediction of the ability of materials to induce respiratory allergy in man is probably questionable.

Effect of amino acid mixtures on nasal allergic responses induced by toluene diisocyanate in mice.

We studied the effect of various amino acid mixtures on nasal allergy induced by the intranasal application of toluene diisocyanate (TDI) in mice. In Experiment 1 (Exp. 1), mice were fed a 25% casein, soy protein isolate (SPI), egg white protein (EW) or gluten diet. In Experiment 2 (Exp. 2), mice were fed a 25% amino acid mixture diets patterned after casein (AA-casein), SPI (AA-SPI), EW protein (AA-EW) or gluten (AA-gluten). In Experiment 3 3 (Exp. 3) we modified the glutamine/glutamic acid (Gln/Glu) concentrations in the amino acid mixtures. Mice were fed a 25% AA-SPI, low Gln/Glu AA-SPI (LG-AA-SPI), AA-EW or high Gln/Glu AA-EW (HG-AA-EW) diet. At the 5th week, mice were divided into sensitized (sen-) and non-sensitized (ns-) groups. The mice in sensitized groups were treated with two courses of intranasal application of toluene diisocyanate (TDI) in ethyl acetate for 5 consecutive days, separated by 9 days rest. The non-sensitized groups of mice were treated with a vehicle. Nine days after the second sensitization, all mice were provoked by TDI. Nasal responses and serum IgE concentration were studied. The findings of Exp. 1 showed that the sen-EW group exhibited a lower body weight gain, higher nasal symptom score and higher IgE concentration than the other sensitized groups. The findings dings of Exp. 2 showed that the sen-EW group had a lower body weight gain, higher nasal symptom score and higher IgE concentration than the other sensitized groups. In Exp. 3, the AA-EW group showed a higher total nasal score and IgE concentration than the HG-AA-EW group, however, the findings of LG-AA-SPI and AA-SPI were similar. These findings demonstrated that amino acid mixtures affect nasal allergy induced by the intranasal application of TDI in mice.