Specific suppressor T cells in rats active in the afferent phase of contact hypersensitivity

The optimal conditions for the induction of contact hypersensitivity in rats and the characteristics of its suppression were studied using the sensitizing haptens dinitrofluorobenzene (DNFB) and trinitrochlorobenzene (TNCB). The hypersensitivity was shown to be hapten specific in so far as TNCB did not sensitize for DNFB responses but sensitization with DNFB did allow a marginal response in rats challenged with TNCB. Suppression of the sensitization to DNFB and TNCB could be generated by intravenous injection of dinitrobenzenesulphonic acid (DNBS) or trinitrobenzenesulphonic acid (TNBS), respectively, up to 3 weeks before sensitization. This suppression was hapten specific and could be transferred with splenic T cells enriched for lymphocytes carrying the OX8 (Tc/s) cell marker. Only the induction phase of sensitization, however, could be suppressed in that way. No suppression acting upon the effector phase could be detected except for a nonspecific local suppression at the site of a previous challenge with an antigen to which the rat was specifically suppressed. This study shows that suppression of contact hypersensitivity in rats is mediated by specific suppressor T cells of which the activation pathway apparently differs from that postulated for mice.     

Induction of in vivo hyporesponsiveness to contact allergens by hapten-modified Ia+ keratinocytes.

Because our previous in vitro studies of hapten-modified Ia+ keratinocytes (KC) indicated that these cells induced anergy in Ag-specific Th1 cells, we assayed such cells for their ability to induce unresponsiveness in an in vivo animal model system of delayed type hypersensitivity (allergic contact dermatitis). Naive animals that were treated with i.p. injections of FITC-modified Ia+ cultured Langerhans cells (cLC) developed allergic contact dermatitis to subsequent hapten challenge; whereas, animals treated with similar doses of FITC-Ia+ KC failed to become sensitized to epicutaneous application of FITC, as evidenced by absent ear swelling responses to a FITC challenge. Those animals that were first treated with intraperitoneal injections of hapten modified Ia+ KC could not be sensitized when they were subsequently exposed to sensitizing doses of FITC; whereas a similar first exposure to FITC-cultured Langerhans cells did not interfere with epicutaneous sensitization. This hyporesponsiveness to sensitization was hapten specific, as FITC-Ia+ KC-treated animals were hyporesponsive only to FITC but not to the irrelevant hapten, TNCB. Additionally, Ia- KC failed to induce unresponsiveness. Additional studies indicate that the hyporesponsiveness was not passively transferrable with splenocytes and was not related to the I-J MHC locus. In contrast to our in vitro studies, the unresponsiveness induced by hapten-modified Ia+ KC in vivo was transient in nature. These data indicate that hapten-modified Ia+ KC function in vivo as nonstimulatory accessory cells, by generating down-regulatory signals that can interfere with the induction of contact hypersensitivity.     

Orally induced tolerance generates an efferently acting suppressor T cell and an acceptor T cell that together down-regulate contact sensitivity

Intragastric administration of the hapten trinitrochlorobenzene (TNCB) suppresses development of contact sensitivity (CS) to attempted epicutaneous sensitization with TNCB. Suppression induced by feeding TNCB is hapten specific and can be transferred to normal animals with lymphoid cells from fed mice. The lymphoid cells in hapten-fed mice that cause suppression of CS have been identified as Thy-1.2-positive cells in spleen and mesenteric nodes. The suppression with Peyer's patch cells from hapten-fed mice appears to be attributable to cells bearing Thy-1.2 antigen (T cell) and to cells with surface Ig (B cell). Feeding TNCB induces an efferent-acting suppressor T cell (Ts eff), as well as an intermediary acceptor T cell (T acc) with which it interacts to block adoptive transfer of CS with immune cells. Ts eff emanating from hapten-fed mice was identified by its specificity for the hapten, insensitivity to pretreatment with cyclophosphamide (CY), ability to produce soluble suppressor factor (SSF), and requirement for T acc to be functional. The presence of T acc in hapten-fed mice, on the other hand, was confirmed by its sensitivity to treatment with CY, interaction with Ts eff or SSF, and the ability to produce nonspecific inhibitor of TDTH cells. Thus, the suppressor T cells that are induced by administering the hapten intragastrically appear to function much like the cells of the suppressor T cell cascade that are induced by giving hapten via parenteral routes.     

Phenotypic and functional characterization of ultraviolet radiation-induced regulatory T cells

Sensitization through UV-exposed skin induces regulatory T cells (Treg). In contrast to the classical CD4+CD25+ Treg that act contact dependent, UV-induced Treg (UV-Treg) suppress via IL-10, indicating a distinct subtype that requires further characterization. Depletion studies revealed that UV-Treg express the glucocorticoid-induced TNF family-related receptor (GITR) and the surface molecule neuropilin-1. The injection of T cells from UV-tolerized mice after depletion of UV-Treg into naive recipients enabled a contact hypersensitivity response, indicating that tolerization also induces T effector cells. Adoptive transfer experiments using IL-10-deficient mice indicated that the IL-10 required for suppression is derived from UV-Treg and not from host-derived cells. Activation of UV-Treg is Ag specific, however, once activated suppression is nonspecific (bystander suppression). Hence, speculations exist about the therapeutic potential of Treg generated in response to Ag that are not necessarily the precise Ag driving the pathogenic process. Thus, we studied the consequences of multiple injections of 2,4-dintrofluorobenzene (DNFB)-specific Treg into ears of naive mice followed by multiple DNFB challenges. DNFB-specific Treg were injected once weekly into the left ears of naive mice and DNFB challenge was performed always 24 h later. After three injections, a challenging dose of DNFB was applied on the right ear. This resulted in pronounced ear swelling, indicating that the subsequent boosting of DNFB-specific Treg had caused sensitization of the naive mice against DNFB. These data demonstrate that UV-Treg express GITR and neuropilin-1 and act via bystander suppression. However, constant boosting of Treg with Ag doses in the challenging range results in final sensitization that might limit their therapeutic potential.     

Suppression of delayed-type hypersensitivity to third party "bystander" alloantigens by antigen-specific suppressor T cells

Delayed-type hypersensitivity (DTH) against alloantigens can be induced by sc immunization with allogeneic cells. The induction of DTH can be suppressed by iv preimmunization of the mice with similar allogeneic spleen cells, provided the cells are irradiated before injection. This suppression is mediated by T cells. The suppressor activity can be induced not only by H-2-and non-H-2-coded antigens, but also by H-2 subregion-coded antigens. Suppression induced by K, I, or D subregion-coded antigens is specific for that particular subregion as well as for its haplotype. I-J-coded alloantigens were found to not be necessary for the induction of antigen-specific suppressor T cells. After restimulation of suppressor T cells by the "specific" alloantigens, the DTH to simultaneously administered third-party alloantigens becomes suppressed as well. This nonspecific suppression of DTH to third party "bystander" alloantigens also occurs when the specific and the third-party antigens are presented on separate cells, provided that both cell types are administered together at the same site. The simultaneous presentation of both sets of alloantigens during the induction phase of DTH only is sufficient to prevent the normal development of DTH to the third-party antigens.     

Immune suppression with supraoptimal doses of antigen in contact sensitivity. I. Demonstration of suppressor cells and their sensitivity to cyclophosphamide.

Immunologic suppression was induced in a mouse model of contact sensitization to DNFB by using supraoptimal doses of antigen. In these studies, in vivo measurement of ear swelling as an indication of immunologic responsiveness correlated well with measurement of in vitro antigen-induced cell proliferation. This unresponsiveness was specific, since supraoptimal doses of DNFB did not interfere with the development of contact sensitivity to another contactant, oxazolone. The decrease in responsiveness is a form of active suppression, as lymphoid cells from supraoptimally sensitized donors transferred suppression to normal recipients. Furthermore, pretreatment with cyclophosphamide (Cy) reversed the suppression seen in supraoptimally sensitized animals but had no effect on the optimal sensitization regimen. These results indicate that supraoptimal doses of contactants can activate suppressor cells and that precursors of these cells are sensitive to Cy. Such suppressors regenerate within 7 to 14 days after Cy treatment. The ability of Cy pretreatment to affect supraoptimal sensitization without affecting optimal sensitization confirms other reports indicating that the observed results of Cy treatment depend critically upon the dose of antigen used.     

Tolerance and contact sensitivity to DNFB in mice. VI. Inhibition of afferent sensitivity by suppressor T cells in adoptive tolerance.

Tolerance in contact sensitivity to DNFB can be adoptively transferred to normal mice with lymph node cells from tolerant donors. This tolerance is antigen specific and is mediated by T cells, i.e., "suppressor" T cells. Experiments were carried out to investigate the mechanism(s) by which the suppressor T cells induce tolerance to DNFB contact sensitivity. The suppressor cells were effective only if they were present during the early stages of the afferent limb of sensitization. As measured by DNA synthesis, cell proliferation in the draining lymph nodes of recipients of suppressor cells was found to be significantly less than in control animals indicating that the suppressor cells acted, at least in part, by limiting or inhibiting DNFB-induced cell proliferation. This inhibition was shown to be antigen specific since the DNFB suppressor cells did not inhibit cell proliferation induced by oxazolone, an unrelated contact sensitizer. The ability to DNFB tolerant cells to block afferent sensitization pathways differs from the mechanism of tolerance to picryl chloride, reported by others, where efferent pathways are blocked.     

Sensitization through carcinogen-induced Langerhans cell-deficient skin activates specific long-lived suppressor cells for both cellular and humoral immunity

Application of 2,4-dinitrofluorobenzene (DNFB) to BALB/c mouse skin depleted of epidermal Langerhans cells (LC) by the chemical carcinogen 7,12-dimethylbenz[a]anthracene (DMBA) activated cells which suppress both contact sensitivity and antibody production when transferred into naive host mice. Tolerance was induced by a concentration of DNFB optimal for inducing contact sensitivity in solvent-treated control mice. The cellular and humoral responses of hosts to a second antigen, 2,4,6-trinitrochlorobenzene (TNCB), were unaffected by these suppressor cells, demonstrating specificity for DNFB. Suppressor cells for cellular and humoral immunity could still be demonstrated 6 months following activation, by which time some mice had died, presumably of old age. The dose responses to sensitizer for generation of cells which suppressed contact sensitivity and antibody production differed, indicating that separate populations of suppressor cells probably inhibit these responses. Hence, during cutaneous chemical carcinogenesis, depletion of LC may allow activation of specific long-lived suppressor cells capable of inhibiting cellular or humoral antitumor immune responses.     

The induction of tolerance to DNFB contact sensitivity by using hapten-modified lymphoid cells. III. Effects of hapten concentration on the ability of MLS-disparate cells to induce rapid unresponsiveness

We investigated genetic restrictions in the induction of immediate tolerance to DNFB contact sensitivity in mice. Using spleen cells from various donor strains haptenated at 500 micro M DNFB, we were unable to detect any restrictions in tolerance induction in recipients that were either syngeneic or allogeneic to the donor strain. However, if the concentration of hapten used in the in vitro labeling was decreased (from 500 micro M to 2.5 to 5 micro M DNFB), differences in tolerogenesis between the various donor strain haplotypes were found. Haptenated spleen cells labeled with 5 micro M DNFB produced a profound level of unresponsiveness in allogeneic recipients but produced minimal tolerance in syngeneic animals. This tolerant state was shown to be antigen-specific and was not produced by unmodified allogeneic cells alone. Further genetic analysis demonstrated that an efficient tolerant state was produced when the donor of the tolerogen and recipient differed at the MLS locus rather than at either the MHC or minor regions. This phenomenon required viable, Thy 1-bearing cells in the haptenated donor population for efficient tolerogenesis to DNFB contact sensitivity.     

Prevention by the MER tubercle bacillus fraction of immunosuppression induced by cancer chemotherapeutic agents

Summary Contact hypersensitivity (CH) to 2,4 dinitro-1-fluorobenzene (DNFB) was induced in BALB/c mice by DNFB skin application. Development of skin CH was suppressed by exposure of the animals after sensitization to the cancer chemotherapeutic drugs cyclophosphamide (CY), sodium methotrexate (MTX), and 5-fluorouracil (5FU). Unresponsiveness to DNFB was also induced in parallel experiments by a single intravenous injection of dinitrobenzenesulfonate (DNBS), either before or concomitant with sensitization. Potentiation of CH skin reactivity was achieved by administration of CY prior to sensitization.Pretreatment by two injections of the methanol extraction residue (MER) tubercle bacillus fraction restored significantly the ability of animals exposed to CY, MTX, or 5FU to respond to DNFB sensitization. The agent did not impair the potentiation of CH skin reactivity that could be effected by administration of CY prior to sensitization.MER treatment was not effective in reversing hapten-induced (DNBS) tolerance in mice.These findings favor the assumption that MER, under the conditions tested, stimulates the function of positively reacting T cells and exerts no enhancing or protective action on suppressor T cells.     

Neutrophil expression of fas ligand and perforin directs effector CD8 T cell infiltration into antigen-challenged skin

Contact hypersensitivity (CHS) is a T cell response to hapten skin challenge of sensitized individuals proposed to be mediated by hapten-primed CD8 cytolytic T cells. Effector CD8 T cell recruitment into hapten challenge sites to elicit CHS requires prior CXCL1- and CXCL2-mediated neutrophil infiltration into the site. We investigated whether neutrophil activities directing hapten-primed CD8 T cell skin infiltration in response to 2,4-dinitro-1-fluorobenzene (DNFB) required Fas ligand (FasL) and perforin expression. Although DNFB sensitization of gld/perforin(-/-) mice induced hapten-specific CD8 T cells producing IFN-γ and IL-17, these T cells did not infiltrate the DNFB challenge site to elicit CHS but did infiltrate the challenge site and elicit CHS when transferred to hapten-challenged naive wild-type recipients. Hapten-primed wild-type CD8 T cells, however, did not elicit CHS when transferred to naive gld/perforin(-/-) recipients. Wild-type bone marrow neutrophils expressed FasL and perforin, and when transferred to sensitized gld/perforin(-/-) mice, they restored hapten-primed CD8 T cell infiltration into the challenge site and CHS. The FasL/perforin-mediated activity of wild-type neutrophils induced the expression of T cell chemoattractants, CCL1, CCL2, and CCL5, within the hapten-challenged skin. These results indicate FasL/perforin-independent functions of hapten-primed CD8 T cells in CHS and identify new functions for neutrophils in regulating effector CD8 T cell recruitment and immune responses in the skin.     

Tolerance and contact sensitivity to DNFB in mice. V. Induction of tolerance with DNP compounds and with free and membrane-associated DNFB.

Immunologic unresponsiveness (tolerance) was induced in a mouse model of contact sensitization to DNFB. The ability to induce tolerance varied with the chemical reactivity of the tolerogen; DNFB was highly tolerogenic, DNBSO3 was moderately tolerogenic, and DNP-lysine was not tolerogenic. Although DNFB is considered a highly reactive compound, tracer studies of injected DNFB showed that it was rapidly excreted. Further studies were therefore done with DNFB attached to mouse erythrocytes. Tolerance to DNFB-RBC was highly specific in vivo; mice tolerant to DNFB showed normal reactivity to TNCB (picryl chloride.) Cells of mice tolerant to DNFB-RBC were also unresponsive to DNBSO3 in vitro. Tolerance to DNFB, DNBSO3, and DNFB-RBC all required time to develop, suggesting that an active process was involved.     

Immunomodulating activities of staphylococcal enterotoxins. I. Effects on in vivo antibody responses and contact sensitivity reaction

The immunomodulating effects of staphylococcal enterotoxins on in vivo immune responses in C57BL/6 mice were examined. Of the five serological types A (SEA), B, C, D, and E (SEE), only SEA and SEE markedly suppressed the antibody response to sheep red blood cells (SRBC) when injected 1 day before or on the day of immunization with SRBC. Further study of SEA revealed that it did not affect the antibody response to a thymus-independent antigen, salmonella flagella, but did affect the T-cell-mediated immune response. Contact sensitivity to dinitrofluorobenzene (DNFB) was suppressed when SEA was injected before sensitization or before challenge with DNFB, indicating that SEA affected both the afferent and efferent phases of DNFB contact sensitivity. As the suppression of DNFB contact sensitivity could be transferred by anti-Thy-1.2 antibody-sensitive spleen cells of SEA injected donors into normal or DNFB-sensitized recipients, the suppression was thought to be an active one. However, SEA could augment the DNFB contact sensitivity when injected on the third day after sensitization with DNFB. These results indicate that the immunomodulating effects of SEA can be mediated by the T-cell function.     

IL-1 receptor signaling is required at multiple stages of sensitization and elicitation of the contact hypersensitivity response

Contact hypersensitivity (CHS) is a CD8 T cell-mediated response to hapten skin sensitization and challenge. The points at which IL-1R signaling is required during this complex, multistep immune response have not been clearly delineated. The role of IL-1R signaling during 2, 4 dinitro-1-fluorobenezene (DNFB) sensitization to induce hapten-specific CD8 effector T cells and in the trafficking of the effector T cells to the DNFB challenge site to elicit the response were investigated using IL-1R deficient mice. DNFB-sensitized IL-1R(-/-) mice had low CHS responses to hapten challenge that were caused in part by marked decreases in hapten-specific CD8 T cell development to IL-17- and IFN-γ-producing cells during sensitization. Hapten-primed wild type CD8 T cell transfer to naive IL-1R(-/-) mice did not result in T cell activation in response to hapten challenge, indicating a need for IL-1R signaling for the localization or activation, or both, of the CD8 T cells at the challenge site. Decreased CD8 T cell priming in sensitized IL-1R(-/-) mice was associated with marked decreases in hapten-presenting dendritic cell migration from the sensitized skin to draining lymph nodes. Transfer of hapten-presenting dendritic cells from wild type donors to naive IL-1R(-/-) mice resulted in decreased numbers of the dendritic cells in the draining lymph nodes and decreased priming of hapten-specific CD8 T cells compared with dendritic cell transfer to naive wild type recipients. These results indicate that IL-1R signaling is required at multiple steps during the course of sensitization and challenge to elicit CHS.     

Distinct in vivo and in vitro cytokine profiles of draining lymph node cells in acute and chronic phases of contact hypersensitivity: importance of a type 2 cytokine-rich cutaneous milieu for the development of an early-type response in the chronic phase.

Although regional lymph nodes (LN) have been extensively studied as rich sources of effector T cells in contact hypersensitivity (CH), it remains unknown whether T cell responses in the LN reflect those in effector skin sites. We previously showed that repeated elicitation of CH results in a shift in the time course of Ag-specific CH from a delayed-type hypersensitivity response to an early-type response, a reflection of a shift in cutaneous cytokine expression from a type 1 to a type 2 profile. To investigate whether repeated elicitation of CH could also drive T cell development to the type 2 phenotype in the regional draining LN, sequential cytokine gene expression after hapten application was assessed during both the acute and the chronic phase of CH. In the draining LN the shift to type 2 cytokine production was also observed, but more mixed patterns of responses were induced than in the corresponding skin sites. The chronic LN cells (LNC), when stimulated in vitro, produced markedly lower levels of type 1 cytokines and higher levels of type 2 cytokines than the acute LNC. A successful passive transfer of an early-type response by the LNC was only induced in the recipient mice when the skin sites chronically treated with hapten were elicited. These results indicate that an early-type response by regional LNC would take place only in a milieu with sufficient levels of type 2 cytokines.     

Contact hypersensitivity reactions to dinitrofluorobenzene mediated by monoclonal IgE anti-DNP antibodies

Several studies have suggested a possible role for IgE antibodies in the pathogenesis of cutaneous hypersensitivity reactions that reach maximum intensity 24 to 48 hr after antigen challenge. The recent availability of murine monoclonal IgE anti-hapten antibodies has made possible the direct examination of the range of cutaneous inflammatory reactions that can be mediated by such antibodies. We have examined the effects of passively sensitizing BALB/c mice with monoclonal IgE anti-dinitrophenyl (DNP) antibody 48 hr before antigen challenge. Inflammatory responses were assessed by measuring ear swelling in mice challenged on the ears with the reactive hapten 2,4-dinitrofluorobenzene (DNFB). Compared with unsensitized controls, the ears of mice passively sensitized with IgE anti-DNP displayed a biphasic pattern of ear swelling after DNFB challenge. An early, transient response (present within 15 to 30 min of challenge and returning to control levels within 4 to 9 hr) was followed by a second, more persistent increase in ear swelling that peaked 24 to 48 hr after challenge. This biphasic pattern of ear swelling seen in IgE-sensitized mice was temporally indistinguishable from that observed in mice conventionally sensitized for allergic contact dermatitis reactions by epicutaneous application of DNFB 5 days before DNFB ear challenge. Antigen specificity of the IgE-mediated contact hypersensitivity reactions was demonstrated by the failure of mice passively sensitized with IgE anti-DNP to display early or delayed ear swelling greater than unsensitized controls when challenged with either of two noncross-reacting haptens, fluorescein isothiocyanate or oxazolone. Mice passively sensitized with a monoclonal IgA anti-DNP antibody (MOPC 315) 48 hr before DNFB challenge failed to display early or delayed ear swelling greater than unsensitized controls. Heat inactivation of the IgE anti-DNP ascitic fluid at 56 degrees C for 30 min completely abolished its capacity to passively sensitize mice for contact hypersensitivity reactions after DNFB challenge. These results document the existence of an antigen-specific, IgE-mediated, delayed-in-time cutaneous hypersensitivity response that can be elicited by epicutaneous challenge (contract) with a reactive hapten.     

Trans-vivo Delayed Type Hypersensitivity Assay for Antigen Specific Regulation

Delayed-type hypersensitivity response (DTH) is a rapid in vivo manifestation of T cell-dependent immune response to a foreign antigen (Ag) that the host immune system has experienced in the recent past. DTH reactions are often divided into a sensitization phase, referring to the initial antigen experience, and a challenge phase, which usually follows several days after sensitization. The lack of a delayed-type hypersensitivity response to a recall Ag demonstrated by skin testing is often regarded as an evidence of anergy. The traditional DTH assay has been effectively used in diagnosing many microbial infections.Despite sharing similar immune features such as lymphocyte infiltration, edema, and tissue necrosis, the direct DTH is not a feasible diagnostic technique in transplant patients because of the possibility of direct injection resulting in sensitization to donor antigens and graft loss. To avoid this problem, the human-to-mouse "trans-vivo" DTH assay was developed ^1,2. This test is essentially a transfer DTH assay, in which human peripheral blood mononuclear cells (PBMCs) and specific antigens were injected subcutaneously into the pinnae or footpad of a naïve mouse and DTH-like swelling is measured after 18-24 hr ³. The antigen presentation by human antigen presenting cells such as macrophages or DCs to T cells in highly vascular mouse tissue triggers the inflammatory cascade and attracts mouse immune cells resulting in swelling responses. The response is antigen-specific and requires prior antigen sensitization. A positive donor-reactive DTH response in the Tv-DTH assay reflects that the transplant patient has developed a pro-inflammatory immune disposition toward graft alloantigens.The most important feature of this assay is that it can also be used to detect regulatory T cells, which cause bystander suppression. Bystander suppression of a DTH recall response in the presence of donor antigen is characteristic of transplant recipients with accepted allografts ^2,4-14. The monitoring of transplant recipients for alloreactivity and regulation by Tv-DTH may identify a subset of patients who could benefit from reduction of immunosuppression without elevated risk of rejection or deteriorating renal function.A promising area is the application of the Tv-DTH assay in monitoring of autoimmunity^15,16 and also in tumor immunology ¹⁷.     

Allergic contact dermatitis induced by intratracheal administration of hapten

It is known that the skin is the natural route for induction of allergic contact dermatitis (ACD). Because the lung is another potential portal of entry for sensitizing chemicals, studies were performed to evaluate immune responses to haptens deposited in the lung. As a result, a new method for inducing ACD was developed. Intratracheal (IT) inoculation of 2,4,6-trinitrobenzene sulfonic acid (TNBS) led to maximal ear swelling 24 hr after challenge on the ear with 2,4,6-trinitrochlorobenzene (TNCB) in carrier. This response was specific for immunizing hapten. Furthermore, it was equally possible to induce ACD by intratracheal inoculation of trinitrophenyl- (TNP) modified bronchoalveolar cells (BAC) or haptenated spleen cells. Adoptive transfer studies demonstrated that the hypersensitivity that resulted from IT inoculation of TNBS or TNP BAC could be transferred with cells. With a monoclonal anti-hamster Ia reagent, it was demonstrated that, like Langerhans cells, 80 to 90% of cells in the bronchial lavage fluid expressed Ia determinants on their membrane. These cells were morphologically indistinguishable from macrophages. Because an individual is capable of being sensitized when hapten is introduced by the pulmonary or epicutaneous routes, the possibility is raised that the alveolar macrophages in the lung possess a similar capability for antigen presentation of hapten in the induction of ACD as does the Langerhans cell.     

Key role for mast cells in nonatopic asthma

The mechanisms involved in nonatopic asthma are poorly defined. In particular, the importance of mast cells in the development of nonatopic asthma is not clear. In the mouse, pulmonary hypersensitivity reactions induced by skin sensitization with the low-m.w. compound dinitrofluorobenzene (DNFB) followed by an intra-airway application of the hapten have been featured as a model for nonatopic asthma. In present study, we used this model to examine the role of mast cells in the pathogenesis of nonatopic asthma. First, the effect of DNFB sensitization and intra-airway challenge with dinitrobenzene sulfonic acid (DNS) on mast cell activation was monitored during the early phase of the response in BALB/c mice. Second, mast cell-deficient W/W(v) and Sl/Sl(d) mice and their respective normal (+/+) littermate mice and mast cell-reconstituted W/W(v) mice (bone marrow-derived mast cells-->W/W(v)) were used. Early phase mast cell activation was found, which was maximal 30 min after DNS challenge in DNFB-sensitized BALB/c, +/+ mice but not in mast cell-deficient mice. An acute bronchoconstriction and increase in vascular permeability accompanied the early phase mast cell activation. BALB/c, +/+ and bone marrow-derived mast cell-->W/W(v) mice sensitized with DNFB and DNS-challenged exhibited tracheal hyperreactivity 24 and 48 h after the challenge when compared with vehicle-treated mice. Mucosal exudation and infiltration of neutrophils in bronchoalveolar lavage fluid associated the late phase response. Both mast cell-deficient strains failed to show any features of this hypersensitivity response. Our findings show that mast cells play a key role in the regulation of pulmonary hypersensitivity responses in this murine model for nonatopic asthma.     

IL-1beta is essential for langerhans cell activation and antigen delivery to the lymph nodes during contact sensitization: evidence for a dermal source of IL-1beta

IL-1beta(-/-) mice manifest an impaired contact hypersensitivity response to the hapten trinitrochlorobenzene, with the principle defect expressed during the sensitization phase of this response. Following application of hapten to the skin, epidermal Langerhans cells of IL-1beta(-/-) mice failed to demonstrate the classical phenotype of activation. In addition, the delivery of epicutaneously applied fluorescein isothiocyanate to draining lymph nodes was decreased in IL-1beta(-/-) mice. Hapten delivery to draining lymph nodes could be restored by intradermal injection of recombinant IL-1beta. Reconstitution of lethally irradiated IL-1beta(-/-) mice by transfer of wild-type bone marrow restored hapten-stimulated IL-1beta mRNA expression, demonstrating that IL-1beta production was dependent on bone marrow-derived cells. In wild-type skin, IL-1beta expression was upregulated in a time- and dose-dependent fashion following hapten application. Interestingly, prominent IL-1beta expressing cells were found in the dermis, suggesting that dermal cells may contribute significantly to the contact hypersensitivity response.