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Risk factors have not been identified that determine susceptibility for development of diisocyanate-induced occupational asthma (DA). We hypothesized that diisocyanate (DI) exposure could modify gene promoter regions regulating transcription of cytokine mediators and thereby influence expression of DA. A cross-sectional study was designed to investigate the promoter methylation status of candidate genes in DI-exposed workers. Subjects consisted of 131 workers in three groups: 40 cases with DA confirmed by a positive specific inhalation challenge (SIC) (DA+), 41 exposed workers with lower respiratory symptoms and negative SIC (DA-), and 50 asymptomatic exposed workers (AWs). We studied four candidate genes (GSTM1, DUSP22, IFN-γ, and IL-4) for which altered promoter methylation has been previously investigated for relationships with a variety of other environmental exposures. Methylation status was determined using methylation-specific quantitative PCR performed on genomic DNA extracted from whole blood. Results showed that relative methylation of IFN-γ promoter was significantly increased in DA+ in comparison with both comparator groups (DA- and AW), and it exhibited good sensitivity (77.5%) and specificity (80%) for identifying DA workers in a multivariate predictive model after adjusting for type of DI exposure, smoking status, methacholine PC₂₀, and gender. IL-4 promoter was slightly less methylated only in DA+ compared with AW among nonsmoking workers. Both GSTM1 and DUSP22 promoter methylations were found not associated with DA. Our finding suggests that exposure to occupational chemicals could play a heretofore undefined mechanistic role via epigenetic modification of specific genes in the promoter region.

BACKGROUND The structural characteristics of diisocyanate chemical protein antigens vary depending upon the methods of production, and may influence diisocyanate antigen immunoassays. The impact of different antigen preparation methods on immunoassay sensitivity, specificity, and predictive value for identifying workers with diisocyanate asthma (DA) has not been systematically evaluated. OBJECTIVE Evaluate the influence of preparation methodology of hexamethylene diisocyanate human serum albumin (HDI-HSA) conjugates on the performance of specific antibody assays for identifying workers with confirmed HDI asthma. METHODS Asthmatic reactions to HDI exposure were assessed in 80 autobody shop workers by specific inhalation challenge (SIC). HDI-specific IgE and IgG in serum were measured by RAST and ELISA with seven different HDI-HSA conjugates prepared in liquid phase with monomeric or polymeric HDI, or vapour-phase monomeric HDI. The HDI : HSA substitution ratios were determined by mass spectrometry. RESULTS DA was confirmed by SIC in 23 subjects. The maximal sensitivity for detecting specific IgE among workers with positive SIC results was higher with RAST and with polymeric vs. monomeric HDI-albumin conjugates (21.7% vs. 8.7%) with a generally high specificity (>or=95%). HDI-HSA specific IgG antibody was also detected in 22-43% of HDI asthmatics depending upon the conjugate used. The specificity of specific IgG varied from 88% to 96%, and it was higher for monomeric (vs. polymeric) HDI-albumin conjugates with low (vs. high) substitution ratios. CONCLUSION The test performance of specific IgE and IgG immunoassays for identifying a positive SIC response varied with different HDI-HSA conjugates. Standard test antigens and common immunoassays must be used to minimize inter-laboratory variability.

Background Specific IgG binding to diisocyanate-human serum albumin (HSA) has been proposed as an indicator of diisocyanate exposure. One residential study reported IgG binding to diisocyanate conjugates in 8% of residents living near a factory using toluene diisocyanate (TDI). Because comparable assays were not performed using individuals distant from such facilities, the significance of this finding is uncertain. Objective To determine the prevalence of diisocyanate specific antibodies in sera from individuals “not known to be exposed” to diisocyanates. Methods Serum samples from 139 anonymous donors without known diisocyanate exposure were assayed by means of enzyme-linked immunosorbent assay for IgG or IgE specific for TDI-HSA, diphenylmethane diisocyanate (MOI)-HSA, and hexamethylene diisocyanate (HDI)-HSA. Positive responses (optical density ≥0.1 and ≥3 SDs above the mean of 8 laboratory controls) were run 3 times. Competitive inhibition was performed for sera exhibiting binding of optical density of at least 0.2. Results We detected IgG reactive with HDI-HSA, diphenylmethane diisocyanate-HSA, and TDI-HSA in 18 (13%), 0, and 7 donors (5%), respectively. Inhibition (>50%) was demonstrated in 6 of 9 participants with elevated HDI-HSA levels and in 2 of 7 with elevated TDI-HSA levels. We detected IgE reactive with the same antigens in 3 donors (2%); however, none were confirmed to be positive using the biotin-streptavidin IgE assay. Conclusions Specific and nonspecific IgG binding to HDI-HSA and TDI-HSA were detected in individuals without known exposure to isocyanates. These antibody measurements may not be reliable indicators of diisocyanate exposure in nonoccupational populations and should not be interpreted as surrogates of diisocyanate exposure in the absence of defined referent populations.

OBJECTIVE To investigate whether genetic variants of N-acetyltransferase (NAT) genes are associated with diisocyanate asthma (DA). METHODS The study population consisted of 354 diisocyanate-exposed workers. Genotyping was performed using a 5'-nuclease polymerase chain reaction assay. RESULTS The NAT2 rs2410556 and NAT2 rs4271002 variants were significantly associated with DA in the univariate analysis. In the first logistic regression model comparing DA+ and asymptomatic worker groups, the rs2410556 (P = 0.004) and rs4271002 (P

Background An outbreak of lung disease among workers in a metal-working plant included 16 biopsy-confirmed cases of hypersensitivity pneumonitis and additional patients with asthma, bronchiolitis and emphysema, usual interstitial pneumonitis, and sarcoidosis. Study design Clinical examination of patients; cross-sectional questionnaire survey of the outbreak plant and two control plant areas, one with and one without MWF exposures, in a separate facility; industrial hygiene survey with laboratory characterization of microbial flora; and immunological investigation. Methods Patients with suspected hypersensitivity pneumonitis underwent a clinical examination including detailed lung function, imaging, and tissue studies. A plant walk-through identified metal-working processes, microbial aerosols, and work practices. Microbial characteristics of the three microbial aerosol-producing processes were characterized. Antibodies to those agents were determined in patient sera. A questionnaire survey was conducted in the case plant and in two areas of a control plant, one with and one without metal-working fluids exposure. Results Thirty-nine (79.6%) patients described symptoms consistent with work-related lung disease, eight received other diagnoses, and two did not complete their examinations. Sixteen patients had hypersensitivity pneumonitis confirmed on biopsy. Mean decrements in lung forced expiratory volume in 1 s and force vital capacity from before to after work were similar in the 16 biopsy-confirmed cases of hypersensitivity pneumonitis ( − 6.3%; − 7.2%) and the 19 symptomatic patients without biopsies ( − 11.2%, − 10.1%). Symptoms were more common in the case plant than in a non-MWF control plant area. Three sources of water-based aerosols were identified that grew similar microbial flora. Although machining increased airborne bacterial levels, the increase was not related to the concentration of viable bacteria in the sumps. Antibody testing did not identify a specific single organisms. Endotoxin levels were similar in case and MWF control plant. Conclusions Lung disease in environments with water-based aerosols may be more common than usually recognized. Patients with HP often present with only subtle abnormalities and may be missed if multiple clinical abnormalities are required to document disease. Am. J. Ind. Med. 39:616–628, 2001. © 2001 Wiley-Liss, Inc.

Although health risks to pesticides containing Bacillus thuringiensis (Bt) have been minimal, the potential allergenicity of these organisms has not been evaluated. Therefore, a health survey was conducted in farm workers before and after exposure to Bt pesticides. Farm workers who picked vegetables that required Bt pesticide spraying were evaluated before the initial spraying operation (n = 48) and 1 and 4 months after (n = 32 and 20, respectively). Two groups of low- (n = 44) and medium- (n = 34) exposure workers not directly exposed to Bt spraying were also assessed. The investigation included questionnaires, nasal/mouth lavages, ventilatory function assessment, and skin tests to indigenous aeroallergens and to a variety of Bt spore and vegetative preparations. To authenticate exposure to the organism present in the commercial preparation, isolates from lavage specimens were tested for Bt genes by DNA-DNA hybridization. Humoral immunoglobulin G (IgG) and immunoglobulin E (IgE) antibody responses to spore and vegetative Bt extracts were assayed. There was no evidence of occupationally related respiratory symptoms. Positive skin-prick tests to several spore extracts were seen chiefly in exposed workers. In particular, there was a significant (p < 0.05) increase in the number of positive skin tests to spore extracts 1 and 4 months after exposure to Bt spray. The number of positive skin test responses was also significantly higher in high (p < 0.05) than in low- or medium-exposure workers. The majority of nasal lavage cultures from exposed workers was positive for the commercial Bt organism, as demonstrated by specific molecular genetic probes. Specific IgE antibodies were present in more high-exposure workers (p < 0.05) than in the low and medium groups. Specific IgG antibodies occurred more in the high (p < 0.05) than in the low-exposure group. Specific IgG and IgE antibodies to vegetative organisms were present in all groups of workers. Exposure to Bt sprays may lead to allergic skin sensitization and induction of IgE and IgG antibodies, or both. Images Figure 1 Figure 2 Figure 3

Background Nasal eosinophils are a biomarker for allergic rhinitis (AR) and are associated with increased symptom severity. Objective To identify predictors of allergic eosinophilic rhinitis (AER) in early childhood in children at higher risk for chronic allergic respiratory disorders. Methods In the Cincinnati Childhood Allergy and Air Pollution Study, infants born to aeroallergen-sensitized and symptomatic parents were examined and underwent skin prick testing (SPT) annually to 15 aeroallergens from 1 to 4 years of age. Wheal circumferences were traced and scanned and areas were determined by computer planimetry. At 4 years, AER was defined as (1) at least 1 positive aeroallergen SPT result, (2) presence of sneezing and runny nose without a cold or influenza, and (3) nasal eosinophilia of at least 5%. Wheal areas at 1 to 3 years were analyzed for an association with AER compared with children without AR. Results At 4 years, 487 children completed rhinitis health histories, SPT, and nasal sampling. Ninety-nine children (22.8%) had AR. Thirty-eight children had AER (8.8% of total sample and 38.4% of AR sample, respectively). At 3 years, for every 1-mm 2 increase in Penicillium species (adjusted odds ratio 1.18, 95% confidence interval 1.06–1.32, P = .002) and maple (adjusted odds ratio 1.07, 95% confidence interval 1.01–1.13, P = .02), wheal area significantly increased the risk of AER at 4 years of age. Conclusion Allergic eosinophilic rhinitis was identified in 8.8% of children at 4 years of age. Age 3 years was the earliest that aeroallergen SPT wheal areas were predictive of AER. Skin testing at 3 years identifies children at risk for an AR phenotype with nasal eosinophilia.

Background Although evidence suggests that ambient exposures to endotoxin and other immunostimulants during early life influence allergic risk, efforts to understand this host-environment relationship have been hampered by a paucity of relevant assays. Objectives These investigations determined whether parameters of house dust extract (HDE) bioactivity were predictive of allergen skin prick test (SPT) reactivity for infants at high risk of allergy participating in the Cincinnati Childhood Allergy and Air Pollution Study (CCAAPS). Methods We conducted a nested case-control study, selecting 99 CCAAPS children who had positive SPT results to at least 1 aeroallergen at age 3 years and 101 subjects with negative SPT results. HDEs were prepared from dust samples collected from the subjects' homes at age 1 year. Murine splenocytes and bone marrow–derived dendritic cells were incubated with HDEs, and supernatant cytokine concentrations were determined by means of ELISA. Alternatively, bone marrow–derived dendritic cells were preincubated with HDEs, and then LPS-induced IL-6 responses were assessed. HDE endotoxin levels were determined by using the limulus amebocyte lysate assay. Results HDEs derived from the homes of children with positive (cases) and negative (control subjects) SPT results had similar bioactivities. However, when cases were considered in isolation, HDEs with higher levels of bioactivity were significantly associated with children who had lower numbers of positive SPT results. Analogous statistical analyses did not identify any association between HDE endotoxin levels and the aeroallergen sensitization profiles of children included in this study. Conclusion HDE immunostimulatory activities predicted the aeroallergen sensitization status of CCAAPS subjects better than HDE endotoxin levels. These results provide the first published evidence that HDE bioassays have clinical relevance in predicting atopic risk.

Indoor mold concentrations were measured in the dust of moldy homes (MH) and reference homes (RH) by quantitative PCR (QPCR) assays for 82 species or related groups of species (assay groups). About 70% of the species and groups were never or only rarely detected. The ratios (MH geometric mean ∶ RH geometric mean) for 6 commonly detected species (Aspergillus ochraceus, A. penicillioides, A. unguis, A. versicolor, Eurotium group, and Cladosporium sphaerospermum) were >1 (Group I). Logistic regression analysis of the sum of the logs of the concentrations of Group I species resulted in a 95% probability for separating MH from RH. These results suggest that it may be possible to evaluate whether a home has an abnormal mold condition by quantifying a limited number of mold species in a dust sample. Also, four common species of Aspergillus were quantified by standard culturing procedures and their concentrations compared to QPCR results. Culturing underestimated the concentrations of these four species by 2 to 3 orders of magnitude compared to QPCR.

Summary The numbers of splenic plasma cells containing b4 or b5 allotypic markers on light chains were determined in b4b5 heterozygous rabbits and in heterozygotes “allotype-suppressed” for b5. The cell count values were compared to relative serum levels of the allotypic markers in each rabbit. Similar comparisons were made for plasma cell and serum percentage of a1 and a2 in normal rabbits and one rabbit allotype-suppressed for a2. General agreement was found between relative serum levels and relative numbers of plasma cells staining for allelic allotypes. In the splenic plasma cells of these heterozygotes, complete cellular segregation of allelic allotypic markers was observed. The allotype-suppressed animals had correspondingly fewer cells containing the allotypic marker which was deficient in the circulating serum immunoglobulins.