Anti-carbamylated protein antibodies in systemic lupus erythematosus patients with articular involvement.

Objective Several studies have evaluated the prevalence of rheumatoid factor (RF) and anti-citrullinated proteins antibodies (ACPA) in systemic lupus erythematosus (SLE) patients but no data are available on the anti-carbamylated proteins (anti-CarP), a new biomarker for rheumatoid arthritis (RA). We evaluated the anti-CarP prevalence in SLE patients with joint involvement and the associations with different phenotypes. Methods Seventy-eight SLE patients with joint involvement were enrolled (F/M 73/5; mean ± SD age 47.6 ± 11.2 years; mean ± SD disease duration 214.3 ± 115.6 months). As control groups, we evaluated SLE patients without joint manifestations ( N = 15), RA ( N = 78) and healthy individuals (HS, N = 98). Anti-CarP were assessed by home-made ELISA in all patients and controls, RF and ACPA in SLE patients with joint involvement (commercial ELISA kit). Results The prevalence of anti-CarP in SLE patients with joint involvement was similar to RA ( p = NS) and significantly higher compared with SLE without joint involvement and HS ( p < 0.0001, p < 0.0001, respectively). Four patients were positive for all three antibodies: seventy-five percent of these showed Jaccoud arthropathy. Fourty-five percent of ACPA-ve/RF-ve patients were anti-CarP + ve. Conclusions The evaluation of anti-CarP in SLE joint involvement demonstrated a prevalence of almost 50%, similar to RA and significantly higher than SLE without joint involvement and HS.

Specific antibodies to diisocyanate and work-related respiratory symptoms in apprentice car-painters.

BACKGROUND: Isocyanates are the main cause of occupational asthma in most countries. Study of immunological markers of diisocyanate asthma may identify individuals at risk. OBJECTIVES: (1) To study changes in specific antibodies to hexamethylene diisocyanates (HDI); (2) to describe the incidence of work-related respiratory symptoms in relation to changes in specific antibody levels. METHODS: Prospective study in 385 apprentice car-painters during their 18 months of training. Participants were assessed on entering and completing their training using questionnaires, methacholine challenges and measurements of HDI-specific immunoglobulin E (IgE), immunoglobulin G (IgG) and subclass 4 of IgG (IgG4) antibodies. RESULTS: Complete data are available for 298 subjects. 13 subjects (4.4%) reported >or=1 new work-related lower respiratory symptoms and 19 (6.4%), >or=1 new work-related nasal symptoms. Increases in levels of specific IgE and IgG above the 97th and 95th percentiles were significantly associated with duration of exposure. Increase in specific IgG was inversely related to incidence of work-related lower respiratory symptoms (OR = 0.001, 95% CI 0.000 to 0.09) after adjusting for relevant covariates. The rise in specific IgG4 was significantly greater in those who did not develop work-related nasal symptoms (OR = 0.09, 95% CI 0.01 to 0.7). CONCLUSION: In this cohort of apprentice car-painters, a small proportion show increases in HDI-specific IgG and IgE after few months of exposure. Increases in specific IgG and IgG4 appear to have a protective effect on the incidence of work-related lower and upper respiratory symptoms, respectively. Assessment of specific antibodies to isocyanates may help identify subjects at risk of developing symptoms.

Respiratory symptoms, sensitization, and exposure response relationships in spray painters exposed to isocyanates.

RATIONALE: Associations between oligomeric isocyanate exposure, sensitization, and respiratory disease have received little attention, despite the extensive use of isocyanate oligomers. OBJECTIVES: To investigate exposure-response relationships of respiratory symptoms and sensitization in a large population occupationally exposed to isocyanate oligomers during spray painting. METHODS: The prevalence of respiratory symptoms and sensitization was assessed in 581 workers in the spray-painting industry. Personal exposure was estimated by combining personal task-based inhalatory exposure measurements and time activity information. Specific IgE and IgG to hexamethylene diisocyanate (HDI) were assessed in serum by ImmunoCAP assay and enzyme immunoassays using vapor and liquid phase HDI-human serum albumin (HDI-HSA) and HSA conjugates prepared with oligomeric HDI. MEASUREMENTS AND MAIN RESULTS: Respiratory symptoms were more prevalent in exposed workers than among comparison office workers. Log-linear exposure-response associations were found for asthmalike symptoms, chronic obstructive pulmonary disease-like symptoms, and work-related chest tightness (prevalence ratios for an interquartile range increase in exposure of 1.2, 1.3 and 2.0, respectively; P

Diisocyanate conjugate and immunoassay characteristics influence detection of specific antibodies in HDI-exposed workers.

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.

Evaluation of antibody binding to diisocyanate protein conjugates in a general population.

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 > or = 0.1 and > or = 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.

Isocyanate vapor-induced antigenicity of human albumin.

BACKGROUND: The bioreactivity of isocyanate, a leading cause of occupational asthma, has led to uncertainty regarding the chemical's antigenicity and mechanisms that elicit immunopathology. OBJECTIVE: To understand better the biologically relevant antigenic forms of hexamethylene diisocyanate (HDI), commonly used in the auto body industry. METHODS: Human albumin was exposed to HDI vapors through a novel approach designed to model the air-liquid interface of the human airway. Vapor HDI-exposed albumin was characterized by electrophoresis, chemical substitution analysis, mass spectrometry, and serology studies on auto body shop workers (N=203) and HDI asthmatics (N=11). RESULTS: HDI vapors caused significant changes in the shape and/or charge of human albumin, which differed from albumin exposed to liquid phase HDI, with lower isocyanate substitution ratios and distinct electrophoretic mobility. Specific sites of vapor HDI conjugation to albumin were identified at His(247) and Lys(414). Vapor HDI-exposed albumin was specifically recognized by the humoral arm of the human immune system, with a strong dependence on albumin as the carrier. Vapor HDI-exposed albumin-specific IgG titers were significantly associated with HDI exposure (P=.001), and specific IgE was detectable in 55% (6/11) of isocyanate asthmatics versus 1.5% (3/203) of exposed healthy workers. Parallel studies using HDI-exposed albumin conjugates produced by previously published methods showed less significant associations of HDI-specific IgG and IgE with exposure and disease, respectively. CONCLUSION: HDI-albumin conjugates produced by novel vapor phase exposure methods may be more physiologically relevant than those produced by previously published methods and of greater utility in characterizing the immune responses associated with HDI exposure and asthma.

Human gamma/delta T-cell proliferation and IFN-gamma production induced by hexamethylene diisocyanate.

BACKGROUND: The human immune response to isocyanate, a leading cause of occupational asthma, remains incompletely characterized, including the cell types involved and the form of the chemical that acts as an antigen. OBJECTIVE: The purpose of this investigation was to characterize human T cells that respond to hexamethylene diisocyanate (HDI), an aliphatic isocyanate routinely used in the automobile body industry. METHODS: Human T-cell lines were generated and characterized from peripheral blood of HDI-exposed and HDI-unexposed subjects, using two different HDI antigens, HDI-conjugated albumin and HDI-exposed human airway epithelial cells (NCI-H292). Flow cytometry was used to characterize the phenotype of HDI-responsive T cells. ELISA and intracellular staining techniques were used to evaluate HDI-induced cytokine production. DNA sequence analysis of T-cell receptors was used to further define clonal populations of HDI-responsive T cells. RESULTS: HDI antigen preparations but not "mock exposed" control antigens lead to increased proliferation of specific cell types, CD3+CD4-CD8(dim) and/or CD3+CD4-CD8- cells, from HDI-exposed but not from HDI-unexposed subjects. These HDI-responsive T cells expressed unique oligoclonal gamma/delta rather than alpha/beta T-cell receptors, with characteristics suggestive of antigen-mediated selection and specificity. The HDI-stimulated gamma/delta T cells were associated with T(H)1-like cytokines and produce IFN-gamma but not IL-5 or IL-13. CONCLUSIONS: These data are the first to demonstrate that HDI can selectively stimulate gamma/delta T cells with the potential to modulate the human immune response to exposure.

Differential roles for CD4 and CD8 T cells after diisocyanate sensitization: genetic control of TH2-induced lung inflammation.

BACKGROUND: Exposure to diisocyanates is a major cause of occupational asthma. We previously developed a novel mouse model of diisocyanate-induced asthma involving epicutaneous sensitization to hexamethylene diisocyanate (HDI) that demonstrates many features of the human disease, including airway eosinophilia and mucus hypersecretion. OBJECTIVE: To determine what factors are critical for the development of HDI-induced airway inflammation, we investigated the strain distribution of this response and the roles of CD4(+) and CD8(+) T cells. METHODS: Mice were epicutaneously exposed to HDI and then challenged with HDI, either by means of inhalation to induce airway inflammation or on the ear to induce contact hypersensitivity (CHS). Lymph node cytokine production and serum antibodies were also measured. RESULTS: Induction of airway eosinophilia was highly dependent on the mouse strain used, with C57BL/6, A/J, CBA, C3H, and C57BL/10 mice all having significantly fewer eosinophils than BALB/c mice. HDI-specific antibodies and lymph node IL-5 and IL-13 production were also diminished in non-BALB/c strains. In contrast, CHS to HDI developed in all strains tested. Studies in mice deficient in either CD4(+) or CD8(+) T cells revealed that CD4(+) T cells were critical for HDI-induced airway eosinophilia, whereas CD8(+) T cells were the major effector cells in CHS. CONCLUSION: The data suggest that, in contrast to CHS, induction of T(H)2 responses after epicutaneous exposure to diisocyanates is strongly genetically influenced. Furthermore, the lung inflammatory response to inhaled HDI appears to depend primarily on effective generation of these CD4(+) T(H)2 responses, as is the case in atopic asthma.

Diisocyanate-exposed auto body shop workers: a one-year follow-up.

BACKGROUND: Diisocyanates currently are the most commonly identified cause of occupational asthma in industrialized countries. Auto body shops, a common hexamethylene diisocyanate (HDI) exposure setting, are difficult to study due to their small size and episodic exposures. OBJECTIVES: A 1-year follow-up was undertaken as an adjunct to the cross-sectional SPRAY study (Survey of Painters & Repairers of Auto bodies by Yale) to investigate the effects of HDI on auto body shop workers over time and whether or not the healthy worker effect may exist in this industry. METHODS AND RESULTS: Forty-eight workers from seven shops were re-contacted. Thirty-four subjects who stayed at the same shop and 11 who left their original shop participated. No statistically significant changes in physiology, symptoms, and immunologic responses from baseline to follow-up were noted. However, significant differences between those who left the shops and those who stayed were noted. Those who left were younger, less experienced in the industry, and more likely to have a history of asthma (23 vs. 3%; P < 0.05), bronchial hyper-responsiveness (23 vs. 9%), HDI-specific IgG (64 vs. 29%; P < 0.05), and HDI-specific proliferation (S.I. 2.0 vs. 1.3; P < 0.05). CONCLUSIONS: The differences in workers who stayed at their shop compared to those who left, combined with the low asthma prevalence and high job turnover rate, all suggest that a healthy worker effect may exist in the auto body industry, and may in part account for the low prevalence of asthma noted in SPRAY and other cross-sectional studies of diisocyante workers.

A novel mouse model of diisocyanate-induced asthma showing allergic-type inflammation in the lung after inhaled antigen challenge.

BACKGROUND: Exposure to diisocyanates, a group of highly reactive, low-molecular-weight compounds, is a major cause of occupational asthma. In contrast to mouse models of atopic asthma, previous mouse models of diisocyanate-induced asthma have failed to show lung inflammation with characteristics of human disease. OBJECTIVE: Our goal was to establish a novel mouse model of diisocyanate-induced asthma in which lung inflammation reminiscent of that seen in human asthma is generated after inhaled antigen challenge. METHODS: BALB/c mice were epicutaneously sensitized to hexamethylene diisocyanate (HDI) and then challenged with an HDI-protein conjugate administered by means of an intranasal droplet. RESULTS: HDI sensitization resulted in development of contact hypersensitivity and HDI-specific antibody production. Most importantly, however, vigorous inflammatory responses with characteristics of human asthma were generated in the lung after inhaled HDI challenge. Challenge of sensitized, but not unsensitized, mice resulted in airway eosinophilia, mucus hypersecretion, and production of T(H)1-type (IFN-gamma) and T(H)2-type (IL-4, IL-5, and IL-13) cytokines by lung inflammatory cells. Despite the mixed T(H)1/T(H)2 response induced by HDI sensitization, use of cytokine-deficient mice revealed that airway eosinophilia was mediated by T(H)2 cytokines and not by IFN-gamma. CONCLUSION: We report a novel mouse model of diisocyanate-induced asthma that, in contrast to previous models, demonstrates antigen-induced lung inflammation with characteristics of human disease. This model will allow investigation of the immunopathogenesis of diisocyanate-induced asthma and should provide insight into this common form of occupational disease.

Respiratory hypersensitivity in guinea pigs sensitized to 1,6-hexamethylene diisocyanate (HDI): comparison of results obtained with the monomer and homopolymers of HDI.

This study used guinea pigs that were sensitized to the biuret or isocyanurate type homopolymers of 1,6 hexamethylene diisocyanate (HDI). Induction was either by intradermal injection or repeated inhalation exposures. For comparison, groups of guinea pigs were sensitized to monomeric HDI. Naive animals served as negative controls. Two and three weeks following induction, animals were challenged by inhalation with the hapten and homologous protein conjugate of the hapten, respectively. Assessments were based on changes in respiratory rate, serum IgG(1)-antibody titer, and influx of eosinophilic granulocytes in airways. Guinea pigs induced and challenged with the HDI-monomer did not display appreciable changes in respiratory rate, whilst the re-challenge with the HDI-protein conjugate caused unequivocal changes in respiratory patterns, including a marked bronchial influx of eosinophilic granulocytes. In contrast, animals induced and challenged with either the free or conjugated aerosols of HDI-homopolymers failed to elicit specific physiological or morphological pulmonary responses. IgG(1) antibodies were observed in all groups receiving monomeric HDI or HDI-homopolymers. Based on the comparative assessment of antibody titers following intradermal injections, it appeared that monomeric HDI was more potent to induce specific IgG(1) antibodies than the homopolymers of HDI. In summary, with respect to induction of allergy and asthma, the data presented here suggest that the homopolymeric forms of HDI appear to be less potent asthmagens, if any, than monomeric HDI.

Subclinical immunologic and physiologic responses in hexamethylene diisocyanate-exposed auto body shop workers.

BACKGROUND: Diisocyanates are potent sensitizing agents and currently the most commonly identified cause of occupational asthma in industrialized countries. However, diisocyanate asthma is difficult to diagnose and exposure and host risk factors are unclear. Auto body shops, one of the most common hexamethylene diisocyanate (HDI) exposure settings, are particularly difficult to study due to their small size and episodic exposures. Surveillance studies of such workers are limited. OBJECTIVES: We have initiated a cross-sectional field epidemiologic study, Survey of Painters and Repairers of Auto bodies by Yale (SPRAY), to characterize the effects of diisocyanate exposures on actively employed auto body shop workers. Methods and Results We present here questionnaire, physiologic, immunologic, and exposure data on 75 subjects enrolled in the study. No overt cases of clinically apparent diisocyanate asthma were identified based on spirometry, methacholine challenge, peak flows, and symptoms. HDI-specific lymphocyte proliferation was present in 30% of HDI-exposed workers and HDI-specific IgG in 34% of HDI-exposed workers, but they were not associated. HDI-specific IgE was detected in two workers. HDI-specific lymphocyte proliferation, increased methacholine responsiveness, and symptoms of chest tightness and shortness of breath were more common in the most heavily HDI-exposed workers, the painters. More long-term follow-up of this cohort should clarify the significance of these HDI-specific immunologic responses, physiologic changes, and symptoms. CONCLUSIONS: These findings demonstrate the presence of HDI-specific immune responses in a large proportion of healthy HDI-exposed workers.

Recent developments in diisocyanate asthma.

Despite recent advances in our understanding of diisocyanate-induced asthma, this disease remains a perplexing phenomenon. Studies reported in the past year have focused on: (i) diisocyanate antigens; (ii) the role of airway and skin epithelium; (iii) human immune responses to exposure; (iv) neurogenic pathways; and (v) genetic factors that may confer susceptibility. These studies support the hypothesis that diisocyanate asthma results from the host immune response to these chemicals, and may represent a mixed T helper type 1/2 response. A better understanding of the pathogenesis of diisocyanate asthma should facilitate the development of better diagnostic tests and strategies for disease surveillance and intervention.

The foreign body reaction to a biodegradable biomaterial differs between rats and mice.

Before a biomaterial can be applied in the clinic, biocompatibility must be tested in in vivo models, by monitoring the foreign body reaction. In this study, we compared the foreign body reaction (FBR) to the biodegradable biomaterial hexamethylenediisocyanate crosslinked dermal sheep collagen (HDSC) between several strains of rats and mice. HDSC disks were implanted subcutaneously on the backs of AO, BN, F344, LEW, and PVG rats and on the backs of 129 SVEV, BALB/c, and C57BL/6 mice. Materials were explanted after 7, 14, 21, and 28 days and processed for (immuno) light and transmission electron microscopic evaluation. In all rat strains, giant cell formation and phagocytosis of HDSC bundles were comparable. In addition, in the PVG rat, many plasma cells infiltrated the HDSC disks. Only a few T cells were present in AO and PVG rats, whereas, in F344 and LEW rats, the presence of T cells was more pronounced. BN rats showed an intermediate T-cell infiltration. In mice, the FBR to HDSC was comparable between the different strains. Compared with rats, giant cell formation was limited, whereas stroma formation was more abundant. Phagocytosis of HDSC bundles rarely occurred in mice, whereas calcification was observed more often. It is concluded that the FBR to HDSC clearly differs between rats and mice. This has consequences for assessment studies on biocompatibility and also on fundamental biomaterial research.

Isocyanate-conjugated human lung epithelial cell proteins: A link between exposure and asthma?

BACKGROUND: Isocyanates are a group of highly reactive cross-linking chemicals that cause airway inflammation and asthma in exposed individuals. Isocyanates have been detected along the airway epithelia of exposed workers and animals, prompting the hypothesis that isocyanates can directly bind to epithelial cell proteins. OBJECTIVE: We tested the hypothesis that hexamethylene diisocyanate (HDI) binds directly to lung epithelial cell proteins and initiated studies to evaluate the immunostimulatory potential of HDI-conjugated lung epithelial cell proteins. METHODS: Human lung epithelial cell lines were exposed to vapor- and liquid-phase HDI, and the cellular proteins were analyzed for HDI conjugation by Western blotting and tested for the ability to induce lymphocyte proliferation in vitro. RESULTS: A number of epithelial cell polypeptides, ranging from 25 to 110 kd in apparent molecular weight, were conjugated with HDI after exposure of the human lung epithelial cell lines (A549 and NCI-H292) to HDI concentrations greater than 0.005% (vol/vol) in the liquid phase. Vapor-phase HDI exposure resulted in a more restricted HDI conjugation pattern, with major HDI-conjugated polypeptides migrating at 47, 71, and 91 kd. HDI-conjugated epithelial cell proteins specifically stimulated proliferation of PBMCs from subjects with isocyanate-induced asthma but not HDI-exposed nonasthmatic individuals or atopic subjects with nonisocyanate-related asthma. CONCLUSIONS: The data demonstrate that epithelial cell proteins readily react with HDI and that HDI-conjugated epithelial cell proteins can stimulate lymphocyte proliferation. Further characterization and evaluation of HDI-conjugated epithelial cell proteins will elucidate their potential role in the pathogenesis of isocyanate-induced asthma.

Diisocyanate antigen-enhanced production of monocyte chemoattractant protein-1, IL-8, and tumor necrosis factor-alpha by peripheral mononuclear cells of workers with occupational asthma.

BACKGROUND: Previous studies have shown a significant association between confirmed diisocyanate-induced asthma (DOA) and in vitro production of diisocyanate antigen-stimulated histamine-releasing factors by PBMCs. Chemokines found in PBMC supernatants are known to express histamine-releasing factor activity. OBJECTIVE: PBMCs of diisocyanate-exposed workers were tested in vitro for diisocyanate antigen-specific enhancement of monocyte chemoattractant protein-1 (MCP-1), monocyte chemoattractant protein-3 (MCP-3), macrophage inflammatory protein-1alpha, RANTES, IL-8, and T-cell cytokines that could play a regulatory role in chemokine synthesis (IL-4, IL-5, IFN-gamma, and TNF-alpha. METHODS: Secretion of chemokines and cytokines was determined by quantitative immunochemical assays of PBMC supernatants. Synthesis of mRNA for beta-chemokines was determined by reverse transcription-polymerase chain reaction. RESULTS: PBMCs of workers with DOA showed significantly enhanced secretion for MCP-1 compared with diisocyanate-exposed asymptomatic workers (P < .05). In vitro induction of antigen-stimulated MCP-1 mRNA synthesis in cultured PBMCs was demonstrated by reverse-transcription polymerase chain reaction. Quantitation of cytokines in supernatants showed increased mean production of IL-8 and TNF-alpha. IFN-gamma, IL-4, and IL-5 were not enhanced in subjects with DOA. CONCLUSION: Antigen stimulation of MCP-1 and TNF-alpha suggest that diisocyanate-specific cellular immune reactions result in activation of macrophages, which may be important in the pathogenesis of DOA.

The kinetics of cytokine production by draining lymph node cells following primary exposure of mice to chemical allergens.

Skin sensitization with chemical allergens is associated with the activation and proliferation of lymphocytes in lymph nodes draining the site of exposure. As lymphocyte activation is regulated by the action of cytokines, we have investigated the nature and kinetics of cytokine production by draining lymph node cells (LNC) from mice, following their primary exposure to chemical allergens. Both interleukin-1 (IL-1) and IL-6 were induced in a biphasic manner following primary exposure of mice to oxazolone or to dicyclohexylmethane-4,4'-diisocyanate (HMDI). The initial phase of production occurred when LNC were prepared from mice 8-20 hr following exposure, while the second peak was coincident with the maximal proliferative response at 72 hr. Increased IL-4 production was observed only when LNC were prepared 96 hr following sensitization. Despite vigorous lymphocyte proliferation there was no evidence for IL-2 production by draining LNC. The ordered and transient pattern of cytokine production that occurs during the afferent phase of contact sensitization suggests that sequential cytokine signals may be involved in regulating the characteristics of the response generated within the draining lymph node.