Predictive methods for food allergenicity: perspectives and current status.
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Animals↗
Biomedical subjects
R J Dearman
Publications and source records attributed to R J Dearman.
Phenotypic variation in mast cell responsiveness to the inhibitory action of nitric oxide.
OBJECTIVE AND DESIGN: The aim of this study was to investigate the possible phenotypic variations between mast cells in terms of their responsiveness to the inhibitory actions of nitric oxide. MATERIALS: Unfractionated mouse peritoneal cells, purified rat peritoneal mast cells, mouse bone marrow-derived mast cells of the C1.MC/C57.1 line (cultured mouse mast cells, CMMC) and rat basophilic leukemia cells of the RBL-2H3 line were used. METHODS: Mast cells were cultured with interferon-gamma (IFN-gamma)-stimulated mouse peritoneal cells as a source of nitric oxide, or with the nitric oxide donor S-nitrosoglutathione (SNOG). After 24 h culture, the mast cells were challenged with anti-IgE, antigen, or calcium ionophore A23187, and degranulation measured as release of [3H]serotonin. RESULTS: Addition of IFN-gamma to mouse peritoneal cells led to nitric oxide synthesis and this was associated with decreased IgE-mediated mast cell degranulation. IFN-gamma did not induce nitric oxide production by CMMC and degranulation of CMMC was not inhibited by nitric oxide generated by co-cultured IFN-gamma-activated peritoneal cells. The nitric oxide donor SNOG inhibited degranulation of purified rat peritoneal mast cells, but not RBL-2H3 cells, stimulated by either IgE cross-linking or calcium ionophore. CONCLUSIONS: The inhibitory effects of nitric oxide on mast cell degranulation are variable and selective for phenotype. Such phenotypic differences may reflect important variations in regulation of mast cell function.
Animals↗
Divergent antibody isotype responses induced in mice by systemic exposure to proteins: a comparison of ovalbumin with bovine serum albumin.
Whereas many foreign proteins are immunogenic, only a proportion is associated commonly with allergy, having the potential to induce the quality of immune response necessary for IgE antibody production and the development of immediate type hypersensitivity reactions in the gastrointestinal and/or respiratory tracts. In the context of toxicological evaluations there is a need to identify those properties that confer on proteins the ability to provoke allergic reactions. The characteristics of antibody responses induced in BALB/c strain mice following administration of ovalbumin (OVA), a significant human allergen, have been compared with those provoked by bovine serum albumin (BSA), a protein considered to have more limited allergenic potential. Intranasal or intraperitoneal (ip) administration of BSA or OVA elicited vigorous IgG and IgG1 antibody responses. Differential IgE antibody production was observed, however, with OVA stimulating relatively high IgE antibody titres at all doses tested whereas no or low titre IgE antibody was detected following exposure to BSA. Furthermore, a differential capacity for IgG2a antibody responses was observed, with only BSA provoking high titres of this IgG subclass. The relative quality of induced responses was equivalent following administration of these proteins via mucosal (in) tissue or via a non-mucosal (ip) route of exposure. IgG2a antibody production is promoted by the type 1 cytokine interferon gamma (IFN-gamma), whereas IFN-gamma and the type 2 cell product interleukin 4 exert reciprocal antagonistic effects on IgE antibody responses. Although cytokine expression patterns were not analysed in this series of experiments, the differential IgE and IgG subclass antibody responses induced by BSA and OVA are consistent with the preferential activation of T helper (Th) 1- and Th2-type cells, respectively. These data indicate that proteins can provoke in mice characteristic antibody (IgE and IgG) isotype profiles suggestive of discrete T lymphocyte responses and that such differences may be associated with variable allergenic activity.
Administration, Intranasal↗
Differential effects of UVA1 and UVB radiation on Langerhans cell migration in mice.
The UVB (280-315 nm)- and UVA1 (340-400 nm)-induced migration of Langerhans cells (LC) from the epidermis and accumulation of dendritic cells (DC) in the lymph nodes draining the exposed skin site of C3H/HeN mice have been investigated. One minimum erythemal dose (MED) of UVB (1.5 kJ/m2) and of UVA1 (500 kJ/m2) were chosen, which have been shown previously to suppress delayed hypersensitivity (DTH). UVB irradiation resulted in a reduction in epidermal LC numbers, local to the site of the exposure, which was most apparent 12 h after exposure, but, in contrast, UVA1 had no significant effect even at 72 h after exposure. UVA1 did not exert any protection against the UVB-mediated depletion in LC numbers. The reduction in local LC following UVB exposure was prevented by systemic (intraperitoneal) treatment of mice with neutralising antibodies to either tumor necrosis factor (TNF)-alpha or interleukin (IL)-beta 2 h prior to the irradiation. It has been reported previously that UVB exposure caused an increase in the number of dendritic cells (DC) in the lymph nodes draining the irradiated skin site. In the present study we have shown that UVA1 had a similar effect. Pretreatment of the mice with neutralising antibodies to IL-1beta (by intraperitoneal injection) substantially inhibited DC accumulation induced by both UV regimens. However, anti-TNF-alpha antibodies affected only the UVB-induced increase, and did not alter the elevation in DC numbers observed following UVA1 exposure. These results indicate that UVB causes the migration of LC from the epidermis and an accumulation of DC in the draining lymph nodes by a mechanism that requires both TNF-alpha and IL-1beta. In contrast, UVAI does not cause LC migration from the epidermis and the accumulation of DC in the draining lymph nodes observed following UVA1 exposure requires IL-1beta, but not TNF-alpha. It is likely therefore that UVA1 acts through a different mechanism from UVB and may target a cutaneous antigen presenting cell other than LC, such as the dermal DC.
Animals↗
Use of the local lymph node assay for the estimation of relative contact allergenic potency.
The effective toxicological evaluation of skin sensitization demands that potential contact allergens are identified and that the likely risks of sensitization among exposed populations assessed. By definition, chemicals which possess the toxicological property of skin sensitization potentially are capable of causing allergic contact dermatitis (ACD) in humans. However, this hazard is not an all-or-none phenomenon; clear dose-response relationships can be discerned and thresholds identified for both the induction of sensitization and the elicitation of contact dermatitis. Commonly, these parameters are grouped under the heading of potency, determination of which is vital for risk assessment. In the present investigation, the local lymph node assay (LLNA) has been employed to determine the relative potency of a range of 20 chemicals. The parameter used is the estimated concentration required to produce a 3-fold increase in draining lymph-node cell proliferative activity, the EC3 value. These measurements have been compared with an assessment of the human sensitizing potency of the 20 selected chemicals, each being assigned to 1 of 5 classes based on their human sensitizing potency. The EC3 value, derived from LLNA work carried out in acetone/ olive oil vehicle, correlated well with the human classification, with the strongest sensitizers having low EC3 values (<O.1%), weaker sensitizers having EC3 values generally in the 1-10% range, and non-sensitizing chemicals having EC3 values in excess of 100%. In conclusion, the derivation of the EC3 for a chemical provides an objective and quantitative estimate of potency that is of considerable utility for skin sensitization risk assessment.
Allergens↗
Activity of human contact allergens in the murine local lymph node assay.
The murine local lymph node assay (LLNA) is a predictive test for the identification of chemicals that have the potential to cause skin sensitization. Since its original development, the assay has been the subject of national and international evaluation studies and extensive comparisons with guinea pig tests and human data. On the basis of these investigations, the LLNA has recently been endorsed by ICCVAM (Interagency Coordinating Committee on the Validation of Alternative Methods) as a stand-alone method for skin sensitization hazard identification. At the same time, ICCVAM confirmed that, although the LLNA is not an in vitro method, it does represent a refinement in the way animals are used and can provide a means for reducing the number of animals used in sensitization hazard assessment. The investigations described here were designed to explore further the ability of the LLNA to identify accurately those chemicals that cause allergic contact dermatitis in humans. To that end we have measured, amongst 3 independent laboratories, LLNA responses induced by a total of 18 test chemicals, 11 of which are known to cause skin sensitization and 7 of which are believed not to be associated with any significant evidence of allergic contact dermatitis in humans. The LLNA correctly classified 16 of the 18 materials. The 11 chemicals tested which are associated with allergic contact dermatitis in humans were found to be positive in the LLNA. Of the 7 materials believed to be non-sensitizers, 5 were negative in the LLNA and 2 produced positive results. Collectively, these data provide additional evidence that the LLNA is able to discriminate skin sensitizers from those chemicals which do not possess a significant skin sensitization potential and thus provides a method for hazard identification that offers important animal welfare benefits.
Allergens↗
Cytokines and chemokines in the initiation and regulation of epidermal Langerhans cell mobilization.
Langerhans cells (LC) are members of the wider family of dendritic cells. LC reside in the epidermis where they serve as sentinels of the immune system, their responsibilities being to sample the external environment for changes and challenges and to deliver information (antigen) to responsive T lymphocytes within skin draining lymph nodes. The ability of LC to migrate from the epidermis to regional lymph nodes is therefore of pivotal importance to the induction of cutaneous immune responses. The journey that LC have to make from the skin has a number of requirements. Initially it is necessary that LC disassociate themselves from surrounding keratinocytes and are liberated from other influences that encourage their retention in the epidermis. Subsequently, migrating LC must successfully traverse the basement membrane of the dermal-epidermal junction and make their way, via afferent lymphatics, to draining lymph nodes. Effective entry into lymph nodes is necessary, as is correct positioning of cells within the paracortex. There is increasing evidence that both cytokines and chemokines, and their interaction with appropriate receptors expressed by LC, orchestrate the mobilization and movement of these cells. We here consider the parts played by these molecules, and how collectively they induce and direct LC migration.
Animals↗
Langerhans cell migration.
Epidermal Langerhans cells (LC) play pivotal roles in the induction of cutaneous immune responses. Encounter with antigen in the skin, or other stimuli, cause the mobilization of LC and their migration, via afferent lymphatics, to draining lymph nodes where they localize within the paracortex. During their movement from the skin LC acquire the characteristics of immunostimulatory dendritic cells (DC) such that the antigen-bearing cells which accumulate in lymph nodes are able to provoke specific T-lymphocyte responses. Epidermal cytokines initiate and regulate LC migration (and maturation), of particular importance being interleukin-1beta and tumour necrosis factor-alpha. Collectively, these cytokines, together with relevant chemokine receptor-ligand interactions, effect the liberation of LC from the epidermis and their directed movement to, and localization within, peripheral lymph nodes. Described here are the phenotypic changes induced during the activation of LC and the mechanisms through which their migration is regulated.
Antigen-Presenting Cells↗
Regulation of epidermal Langerhans cell migration by lactoferrin.
Lactoferrin (LF) is a member of the transferrin family of iron-binding glycoproteins to which several anti-inflammatory functions have been ascribed. LF has been shown to down-regulate expression of the pro-inflammatory cytokine tumour necrosis factor-alpha (TNF-alpha), although the possibility has been raised that the activity of LF in this regard was indirect and secondary to its ability to bind to and inactivate the bacterial lipopolysaccharide (LPS) used to induce cytokine production. However, the identification of putative membrane receptors for LF raises the possibility that the interaction of LF with its receptor may be one important route through which this protein exerts anti-inflammatory activity. In the present investigations the biological properties of LF have been examined in a model of cutaneous immune function where the allergen-induced migration of epidermal Langerhans cells (LC) from the skin and their subsequent accumulation as dendritic cells (DC) in skin-draining lymph nodes are known to be dependent upon the de novo synthesis of TNF-alpha, but independent of exogenous LPS. Consistent with the protein having direct anti-inflammatory properties, it was found that the intradermal injection of recombinant murine LF (either iron-saturated or iron-depleted LF) inhibited significantly allergen (oxazolone) -induced LC migration and DC accumulation. That these inhibitory effects were secondary to the inhibition of local TNF-alpha synthesis was suggested by the findings that first, LF was unable to inhibit LC migration induced by intradermal injection of TNF-alpha itself, and second, that migration stimulated by local administration of another epidermal cytokine, interleukin 1beta, which is also dependent upon TNF-alpha production, was impaired significantly by prior treatment with LF. Finally, immunohistochemical analyses demonstrated the presence of LF in skin, associated primarily with keratinocytes. Collectively these data support the possession by LF of direct immunomodulatory and/or anti-inflammatory activity, probably associated in this case with inhibition of cytokine production. Furthermore, the results suggest that as a constituent of normal skin, LF may play a role in homeostatic regulation of cutaneous immune function.
Allergens↗
Role of CD4(+) T helper 2-type cells in cutaneous inflammatory responses induced by fluorescein isothiocyanate.
Owing to its skin-sensitizing and fluorochromatic properties, fluorescein isothiocyanate (FITC) is employed frequently as an experimental hapten in mechanistic studies of contact allergy, particularly in the context of the role of migration and activation of Langerhans' cells. In this study we demonstrated that topical exposure of mice to FITC results in the selective development of activated lymph node cells (LNC) expressing a preferential type 2 cytokine-secretion profile, with high levels of interleukin (IL)-4 and IL-10, but low levels of interferon-gamma (IFN-gamma). Negative selection (complement depletion) identified CD4(+) T helper (Th)2-type cells as the primary source in activated LNC of the type 2 cytokines IL-4 and IL-10, whereas the low levels of IFN-gamma produced were derived exclusively from CD8(+) T cytotoxic (Tc) 1-type cells. A biphasic pattern of cutaneous inflammatory reactions was elicited by exposure to FITC, the early phase of which could be transferred passively with serum (presumably immunoglobulin E [IgE] antibody), whereas adoptive transfer experiments demonstrated that Th2-type CD4(+) cells were responsible for the delayed-type component of the dermal hypersensitivity reaction. In contrast with contact allergic reactions induced by other sensitizing haptens, which are considered to be largely Th1/Tc1-mediated immune processes regulated by Th2-type cells, these results suggest therefore that the skin lesions provoked in mice by FITC are primarily a result of the activation of Th2-type cells.
Adoptive Transfer↗
Local lymph node assay: validation assessment for regulatory purposes.
For the prediction of skin sensitization potential of substances, the murine local lymph node assay (LLNA) is an alternative to the widely used guinea pig tests. For more than 10 years, this method has undergone extensive development, evaluation, and validation. In this review, the validation status of the LLNA is considered, specifically with regard to its use for regulatory identification of skin sensitization hazards. The LLNA is a method for the predictive identification of chemicals that have a potential to cause skin sensitization. Activity is measured as a function of lymph node cell proliferative responses stimulated by topical application of test chemicals. The LLNA has successfully passed all reasonable validation stages. It provides a reliable and relevant source of predictive skin sensitization data, which unlike results from guinea pig tests, are reproducible from laboratory to laboratory. In summary, the LLNA is now ready for acceptance as a viable and complete alternative to traditional methods, offering a substantial reduction in animal numbers and refinement opportunities without compromising the standards for the identification of important skin sensitizers.
Allergens↗
Antigen-specific and nonspecific determinants of cytokine production during topical sensitization of mice to chemical allergens.
BACKGROUND: Topical exposure to chemical allergens such as trimellitic anhydride or 1-chloro-2,4-dinitrochlorobenzene results in the accumulation of dendritic cells (DCs) and subsequent rapid up-regulation of CD4 T-cell proliferation and cytokine secretion within draining lymph nodes. OBJECTIVE: We investigated the contribution of antigen-specific and CD40 ligand (CD40L)-mediated signals to chemical allergen-induced CD4 T-cell growth and cytokine production. METHODS: DCs enriched from lymph nodes of allergen-challenged animals by metrizamide centrifugation were used to stimulate cytokine and proliferative responses by magnetic immunobead-sorted CD4 T cells primed in vivo with the same or unrelated allergen. Cultures of DCs and T cells were supplemented with antibodies that block IL-12 and CD40L activity. RESULTS: Proliferation of CD4 T cells was stimulated by DCs primed with the same but not unrelated antigen, whereas IFN-gamma, IL-12, and IL-10 secretion were provoked equally well by DCs primed with either hapten. Blockade of CD40L engagement abrogated production of IFN-gamma (80%) and IL-12 (95%) under antigen-nonspecific stimulatory conditions. In contrast, IL-10 secretion was enhanced after CD40L blockade under both antigen-specific and nonspecific conditions. Primary CD4 T cells activated by mitogen were also influenced by DCs in the same way. CONCLUSION: These results show that during the development of chemical sensitization emerging CD4 T-cell growth and cytokine production are regulated by independent mechanisms requiring antigen presentation and CD40 signaling, respectively.
Allergens↗
Why are some proteins allergens?
The ability of certain proteins to induce an allergic response in susceptible individuals is well established. Symptoms can range from mild erythema or rhinitis, to acute, and possibly fatal, anaphylactic shock. Because such allergic responses require complex interactions between the protein and the immune system, they are notoriously difficult to predict. Nevertheless, it is clear that some proteins are intrinsically more allergenic than others. The challenge for toxicologists is to identify those characteristics that confer on proteins the potential to induce allergic sensitization and allergic disease. Here, we first consider the potential contribution that individual epitopes may make to the allergenicity of a protein. These are the minimal peptide units within proteins that can be recognized by the immune system and are a fundamental requirement for all immune responses, including those resulting in allergic sensitization. It appears that allergens must necessarily contain B-cell epitopes to which immunoglobulin E (IgE) can bind, and T-cell epitopes capable of inducing a type 2 T-lymphocyte response. Nevertheless, it appears doubtful that the presence of appropriate epitopes alone is sufficient to endow a protein with allergenic potential. We therefore consider also the contribution that other features and characteristics of proteins may make to their overall allergenicity. In particular, we consider the effects that resistance to proteolysis, post-translational glycosylation, and enzymatic activity may have. It appears that relative stability in simulated gastric fluid (SGF) sometimes correlates with allergenic activity. However, this is not universally true, and it is known that there are protein allergens, such as some of those associated with oral allergy syndrome, that are unstable. Nevertheless, if stability in SGF is associated with the intrinsic allergenicity of many proteins irrespective of the route of exposure, then this may reflect some more fundamental property of proteins, and possibly their stability in other biologic matrices and/or to intracellular proteases. Post-translational modification appears generally to enhance allergenicity, perhaps by increasing uptake and detection of the protein by the immune system. Some enzymatic activities also enhance allergenicity through what appear to be several different mechanisms, including nonspecific activation of cells participating in the immunologic response. Overall, it appears likely that many factors can contribute to the overall allergenicity of any given protein. Some, such as the presence of epitopes with allergenic potential, may be essential. Others, such as the glycosylation status, resistance to proteolysis, and enzymatic activity, may play a subsidiary but nevertheless critically important role. By better defining the limits within which these factors operate, we can hope to gain a better understanding of the fundamental origins of protein allergenicity, and therefore be in a position to identify and characterize the hazards and risks of allergic disease associated with novel proteins.
Allergens↗
Failure of vehicle to influence local lymph node assay response to benzocaine.
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Animals↗
Intracellular phosphotyrosine induction by major histocompatibility complex class II requires co-aggregation with membrane rafts.
Cross-linking MHC class II molecules human leukocyte antigen (HLA-DR) on the surface of THP-1 cells was found to induce their entry into the glycolipid-enriched membrane fraction of the plasma membrane. At the cellular level, this resulted in the synergistic co-aggregation of class II with cholera toxin, a marker of membrane rafts. The accompanying induction of intracellular protein tyrosine phosphorylation could be inhibited by treating cells with methyl-beta-cyclodextrin, a drug that chelates membrane cholesterol and thereby disperses membrane rafts. Signaling could also be inhibited by treating cells with the Src-family kinase inhibitor PP1. Together, these results show that the induced association of class II molecules with membrane rafts can contribute to their aggregation on the cell surface and mediate an association with intracellular protein-tyrosine kinases.
Biomarkers↗
Selective down-regulation of T cell- and non-T cell-derived tumour necrosis factor alpha by thalidomide: comparisons with dexamethasone.
Both thalidomide and dexamethasone have been shown to inhibit the production of tumour necrosis factor alpha (TNF-alpha), but little is known of their cellular selectivity. Inhibition of monocyte TNF-alpha expression has been implicated in the clinical efficacy of thalidomide, and it has been suggested that the drug modulates only monocyte-derived cytokines. Given the importance of T lymphocyte responses in immunological disorders in which treatment with thalidomide has been successful, it is pertinent to study the effects of this drug on T cell-derived TNF-alpha. In the present investigations we have examined the influence of both thalidomide and dexamethasone on mitogen-induced elaboration of TNF-alpha by CD3+ peripheral blood mononuclear cells (PBMC) and the T cell line MOLT-4. PBMC from healthy human volunteers were stimulated optimally with phytohaemagglutinin (PHA) in the presence of varying concentrations of thalidomide or dexamethasone, and supernatants assayed for TNF-alpha and interleukin 2 (IL-2). Concurrently, PHA-stimulated PBMC were treated with 1 x 10(-1) mM thalidomide or dexamethasone and the cells fixed, permeabilised, stained with anti-CD3 and anti-TNF-alpha fluorescently labelled antibodies and analysed by flow cytometry. MOLT-4 cells were cultured in the presence or absence of the drugs following activation with phorbol myristate acetate (PMA)/ionophore, and supernatants analysed by enzyme-linked immunosorbent assay (ELISA) for cytokine expression. Thalidomide was found to inhibit PBMC-derived TNF-alpha, but not IL-2. In contrast, dexamethasone down-regulated both TNF-alpha and IL-2 in a dose-dependent manner. Thalidomide and dexamethasone both suppressed intracellular levels of TNF-alpha in CD3+ PBMC, reducing percentages of double positive staining cells by 28 and 52%, respectively, compared with controls. In addition, TNF-alpha production by CD3- PBMC was inhibited by 31% by thalidomide and by 47% by dexamethasone. In order to determine whether thalidomide was acting directly on T cells, or indirectly through effects on accessory cells, TNF-alpha production in the T cell line MOLT-4 was investigated. TNF-alpha secretion by PMA/ionophore activated MOLT-4 cells was reduced by 80% following thalidomide treatment and close to background levels following dexamethasone treatment. To verify that thalidomide was acting selectively to down-regulate TNF-alpha, IL-2 production by MOLT-4 cells was also measured and found to be unaffected by the drug. In contrast, dexamethasone reduced MOLT-4-derived IL-2 levels by 20%. These observations suggest that thalidomide, in addition to its known inhibitory effect on monocyte-derived TNF-alpha, is capable also of down-regulating T cell-derived TNF-alpha in a direct and selective manner. In addition, the inhibition of intracellular levels of TNF-alpha strengthens the evidence that the inhibitory effect of thalidomide is at the level of transcription and/or translation and does not reduce cellular TNF-alpha secretion. Such effects could explain the efficacy of thalidomide treatment in various immunological disorders where T cell activation plays an important role in the pathogenesis of the disease.
CD3 Complex↗
Immune responses to contact allergens: novel approaches to hazard evaluation.
Progress in our understanding of the immunobiological mechanisms that cause skin sensitization and allergic contact dermatitis has facilitated consideration of alternative approaches to hazard evaluation. One such is the murine local lymph node assay in which, in contrast to more traditional guinea pig tests, sensitizing activity is measured as a function of events associated with the induction, rather than the elicitation, phase of contact hypersensitivity. Activity in the local lymph node assay is dependent upon all of those immunological events that are initiated following first encounter with chemical allergen and which result in the stimulation of T lymphocyte proliferative responses in lymph nodes draining the site of exposure. In this respect the assay embraces in an holistic way the induction of skin sensitization. With the objective of developing in vitro approaches to hazard identification, consideration has been given to discrete immunological responses that characterize the induction of skin sensitization. Most attention has focused upon the changes induced by chemical allergens in the phenotype and function of epidermal Langerhans cells and in cytokine expression. In addition, attempts have been made to identify contact allergens as a function of their ability to provoke in vitro specific responses by unprimed T lymphocytes. These novel approaches to skin sensitization testing and their potential utility in the context of toxicological evaluations are reviewed in this article.
Allergens↗
Analysis of interleukin 12 protein production and mRNA expression in mice exposed topically to chemical allergens.
Interleukin (IL) 12 is a heterodimeric cytokine which stimulates IFN-gamma production and promotes the development of type 1 T helper cells and T cytotoxic cells from their respective precursors. We have described previously that contact allergens such as 2,4-dinitrochlorobenzene (DNCB), and respiratory allergens such as trimellitic anhydride (TMA) induce discrete type 1 and type 2 cytokine secretion patterns, respectively, following repeated topical exposure of BALB/c strain mice. Under such conditions, we have now examined production by draining LNC of the inducible subunit of IL-12 (p40) and p40 and p35 subunit mRNA expression. Cultured LNC prepared from mice treated with DNCB secreted significantly more IL-12 p40 protein than did TMA- or vehicle-activated LNC, such differences becoming more pronounced as the duration of culture increased. Maximal levels of mRNA expression of the IL-12 p40 subunit were observed after 6-24 h of culture in all treatment groups and declined thereafter. Somewhat higher levels were induced following exposure to DNCB, these differences only reached statistical significance at 24 h of culture. Expression of this subunit by LNC from all treatment groups declined with time in culture. Levels of IL-12 p35 mRNA expression were comparable in cultured LNC prepared from allergen and vehicle treated mice and remained constant throughout the entire culture period. These data indicate that the divergent T cell cytokine responses seen in response to contact and respiratory allergens are associated with differential production of IL-12 p40 protein in the absence of substantial changes in the expression of mRNA for either subunit.
Acetone↗