Use of intranasal IL-12 to target predominantly Th1 responses to nasal and Th2 responses to oral vaccines given with cholera toxin

We have investigated the effects of IL-12 and cholera toxin (CT) on the immune response to tetanus toxoid (TT) given by intranasal or oral routes. CT inhibited IL-12-induced IFN-gamma secretion both in vivo and in vitro. Intranasal administration of IL-12 to mice nasally immunized with the combined vaccine of TT and CT resulted in increased TT-specific IgG2a and IgG3 Abs, while IgG1 and IgE Ab responses were markedly reduced. This shift of the CT-induced immune response toward Th1 type was associated with TT-specific CD4+ T cells secreting IFN-gamma and reduced levels of Th2-type cytokines (i.e., IL-4, IL-5, IL-6, and IL-10). In contrast, intranasal IL-12 enhanced the CT-induced serum IgG1 and IgE Ab responses in mice given the combined vaccine orally. IFN-gamma secretion by TT-specific CD4+ T cells was also enhanced; however, Th2-type cytokine responses were predominant. Mucosal secretory IgA responses to oral or nasal vaccines were not affected by intranasal IL-12. Thus, intranasal IL-12 delivery influences Th cell subset development in mucosal inductive sites that are dependent on the route of vaccine delivery.     

Endogenous IL-12 is required for control of Th2 cytokine responses capable of exacerbating leishmaniasis in normally resistant mice

We investigated the mechanisms by which treatment with anti-IL-12 Ab prevents cure of infection with Leishmania major in resistant C57BL/6 mice. Consistent with delayed production of IL-12, anti-IL-12 Abs could be administered as late as 2 wk after infection to exacerbate disease. Starting at 2 wk of infection, the cultured lymph node cells from mice treated with either polyclonal or monoclonal anti-IL-12 Abs persistently generated 3- to 10-fold more IL-4 and IL-10 in response to L. major Ag compared with cells from mice receiving preimmune goat IgG. Reciprocal decreases in Ag-specific IFN-gamma production were observed in mice receiving anti-IL-12 Abs. A similar reversal of IFN-gamma and IL-4 production accompanied progressive disease induced by pretreatment with a single dose of anti-IFN-gamma mAb. Although IFN-gamma production was suppressed for up to 4 wk in mice treated with monoclonal anti-IL-12 or anti-IFN-gamma, coadministration of neutralizing anti-IL-4 IgG reversed progressive illness. These findings demonstrate that IL-12 produced in vivo is necessary for both the emergence of IFN-gamma producing cells and the down-regulation of Th2 cell responses during murine leishmaniasis. Furthermore, the uninhibited production of IL-4 was required to sustain progressive infection initiated by the decreased IFN-gamma synthesis observed in anti-IL-12 and anti-IFN-gamma-treated mice.     

Lack of type 2 T cell-independent B cell responses and defect in isotype switching in TNF-lymphotoxin alpha-deficient mice

TNF is implicated in in vitro Ig production, but its role in vivo is not clearly defined. Our previous studies had shown that TNF-LT alpha double-deficient mice have defective IgM and IgG primary Ab responses to the T cell-dependent (TD) Ag SRBC. We now extend these studies to secondary responses and to T cell-independent (TI) B cell responses. Injections of the TD Ag SRBC did not induce germinal center formation in the spleen of TNF-LT alpha-deficient mice. Associated with the morphologic defect, there was a defective IgG Ab response and a secondary hyper-IgM response to the TD Ag in TNF-LT alpha-deficient mice. The response to the TI Ag type 2 DNP-alanyl-glycyl-glycyl-Ficoll was essentially absent in TNF-LT alpha-deficient mice, while that to the TI Ag type 1 TNP-LPS was significantly reduced only for IgG2b isotype. Transplantation of bone marrow cells from wild-type mice into irradiated TNF-LT alpha-deficient mice restored the formation of splenic germinal centers and corrected the IgM and IgG responses to both TD and TI Ags. These data suggest that TNF and/or LT alpha signaling are critically required for germinal center formation and for the IgM and IgG responses to both TD and TI type 2 Ags.     

IL-12 down-regulates autoantibody production in mercury-induced autoimmunity

In genetically susceptible H-2s mice, subtoxic doses of mercuric chloride (HgCl2) induce a complex autoimmune syndrome characterized by the production of anti-nucleolar IgG Abs, lymphoproliferation, increased serum levels of IgG1 and IgE Abs, and renal Ig deposits. Mercury-induced autoimmunity in H-2s mice provides a useful model for chemically related autoimmunity in humans. The increase in serum IgG1 and IgE, which are under IL-4 control, suggests a role for the Th2 subset in this syndrome. The IL-12 cytokine induces T cell proliferation and IFN-gamma production and is necessary for differentiation of naive T cells into the Th1 subset. To gain an understanding of T cell control in this syndrome and, in particular, Th1/Th2 regulation, we assessed the effect of IL-12 administration in mercury-induced autoimmunity. Groups of A.SW mice (H-2s) received HgCl2 plus IL-12, HgCl2 alone, or IL-12 alone. IL-12 treatment resulted in a dramatic reduction of the anti-nucleolar Ab titers. IL-12 also inhibited the HgCl2-induced serum IgG1 increase, but, in contrast, did not significantly affect IgE induction in this model. This observation may be related to our unexpected finding that IL-12 further potentiated the HgCl2-triggered IL-4 induction in this model. The levels of renal Ig deposits were similar in mice receiving HgCl2 alone or HgCl2 plus IL-12. Our results indicate that IL-12 can down-regulate the autoimmune component of this experimental syndrome and that the various manifestations of mercury-induced autoimmunity are independently regulated.     

Adenoviral gene delivery elicits distinct pulmonary-associated T helper cell responses to the vector and to its transgene

Replication-deficient adenovirus (Ad) vectors are effective to specifically target the respiratory epithelium for either corrective gene therapy such as cystic fibrosis or for mucosal immunization. As a consequence of transducing the lower respiratory tract with an E1/E3 deleted Ad5 vector, host responses have been characterized by the duration of transgene expression and by the induction of CTL responses. However, limited emphasis has been devoted to understanding the contribution of CD4+ T cell responses to the Ad vector. Both CD4+ and CD8+ T cells migrate into the lung following sequential intratracheal Ad5 transgene instillations. Isolated CD3+ T lymphocytes from the lungs were predominantly of the Th2 type, and after cell sorting, the IL-4-producing T cells were largely CD4+, while IFN-gamma expression was associated with both CD4+ and CD8+ T cells. Ab responses to the Ad5 vector and to the expressed transgene beta-galactosidase (beta gal) revealed elevated bronchial and serum IgA and IgG Abs with low neutralization titers. Analysis of serum IgG subclass responses showed IgG1 and IgG2b with lower IgG2a Abs to Ad5 and IgG2a and IgG2b Ab responses to beta gal. Ad5-specifc CD4+ T cells produced both Th1 (IFN-gamma and IL-2)- and Th2 (IL-4, IL-5, IL-6)-type cytokines, while beta gal-specific CD4+ T cells secreted IFN-gamma and IL-6. This study provides direct evidence for the concomitant induction of Th2- with Th1-type responses in both the pulmonary systemic and mucosal immune compartments to the Ad5 vector as well as a Th1-dominant response to the transgene.     

A 70-kilodalton recombinant heat shock protein of Candida albicans is highly immunogenic and enhances systemic murine candidiasis

The 70-kDa recombinant Candida albicans heat shock protein (CaHsp70) and its 21-kDa C-terminal and 28-kDa N-terminal fragments (CaHsp70-Cter and CaHsp70-Nter, respectively) were studied for their immunogenicity, including proinflammatory cytokine induction in vitro and in vivo, and protection in a murine model of hematogenous candidiasis. The whole protein and its two fragments were strong inducers of both antibody (Ab; immunoglobulin G1 [IgG1] and IgG2b were the prevalent isotypes) and cell-mediated immunity (CMI) responses in mice. CaHsp70 preparations were also recognized as CMI targets by peripheral blood mononuclear cells of healthy human subjects. Inoculation of CaHsp70 preparations into immunized mice induced rapid production of interleukin-6 (IL-6) and tumor necrosis factor alpha, peaking at 2 to 5 h and declining within 24 h. CaHsp70 and CaHsp70-Cter also induced gamma interferon (IFN-gamma), IL-12, and IL-10 but not IL-4 production by CD4+ lymphocytes cocultured with splenic accessory cells from nonimmunized mice. In particular, the production of IFN-gamma was equal if not superior to that induced in the same cells by whole, heat-inactivated fungal cells or the mitogenic lectin concanavalin A. In immunized mice, however, IL-4 but not IL-12 was produced in addition to IFN-gamma upon in vitro stimulation of CD4+ cells with CaHsp70 and CaHsp70-Cter. These animals showed a decreased median survival time compared to nonimmunized mice, and their mortality was strictly associated with organ invasion by fungal hyphae. Their enhanced susceptibility was attributable to the immunization state, as it did not occur in congenitally athymic nude mice, which were unable to raise either Ab or CMI responses to CaHsp70 preparations. Together, our data demonstrate the elevated immunogenicity of CaHsp70, with which, however, no protection against but rather some enhancement of Candida infection seemed to occur in the mouse model used.     

Lipid vesicle size determines the Th1 or Th2 response to entrapped antigen

Understanding the factors that control the differential induction of Th1 and Th2 responses is a key immunologic objective with profound implications for vaccination and immunotherapy of infectious and autoimmune diseases. Using Ag formulated in lipid vesicles prepared from nonionic surfactants, we describe a novel mechanism influencing the balance of the Th1 or Th2 response. Our results indicate that inoculation of BALB/c mice with vesicles with a mean diameter > or = 225 nm preferentially induces Th1 responses, as characterized by increased titers of IgG2a in plasma and elevated IFN-gamma production by lymph node cells. However, preparation of the same quantity of Ag in vesicles with mean diameter of < or = 155 nm induces a Th2 response, as identified by IgG1 in the absence of IgG2a production and increased lymph node IL-5 production. Although large (> or = 225 nm) vesicles could induce IL-12 production, smaller vesicles (< or = 155 nm) could not. However, small vesicles did induce higher levels of IL-1beta production by macrophages than larger vesicles. The role of IL-12 in this response was confirmed in IL-12-deficient mice, whose spleen cells failed to produce IFN-gamma following in vivo priming with Ag prepared in large vesicles. Our results therefore indicate that macrophages respond to endocytosis of large or small vesicles by producing different patterns of cytokines that can subsequently direct the immune response toward a Th1 or a Th2 phenotype.     

Role of IL-4 and IFN-gamma in modulation of immunity to Borrelia burgdorferi in mice

Because T cells appear to modulate the severity of murine Borrelia burgdorferi infections, we decided to examine the possible involvement of T cell-associated cytokines in disease outcome. Comparison of in vitro B. burgdorferi Ag-induced cytokine production in disease-susceptible and -resistant strains revealed striking differences; spleen cells from susceptible C3H mice produced significantly higher levels of IL-2 and IFN-gamma and lower levels of IL-4 than spleen cells from resistant BALB/c mice. Lymph node responses were even more divergent, with C3H mice producing high levels of IFN-gamma, and BALB/c mice producing little or none. This apparent Th1/Th2 cytokine imbalance was also reflected in vivo, since serum from C3H had significantly higher levels of B. burgdorferi-specific IgG2a Ab and lower levels of IgG1 Ab than serum from BALB/c mice. In vivo studies confirmed the importance of IL-4 in early control of spirochete growth, since treatment of either strain with neutralizing anti-IL-4 mAb led to increased joint swelling and higher spirochete burdens in joints compared with those in control mAb-treated mice. In contrast, IFN-gamma may hinder early control of spirochete growth in susceptible C3H mice, since treatment of mice with neutralizing anti-IFN-gamma mAb reduced both joint swelling and joint spirochete burdens compared with those in control mAb-treated mice. These studies indicate opposing roles for IL-4 and IFN-gamma in the modulation of spirochete growth and disease development in B. burgdorferi-infected mice and suggest that differential cytokine production early in infection may contribute to strain-related differences in susceptibility.     

Down-regulating effects of IL-4 and IL-10 on the IFN-gamma response in atopic dermatitis

Atopic dermatitis (AD) is a chronic allergic disease associated with toxin (superantigen)-producing Staphylococcus aureus skin infections, impaired delayed hypersensitivity responses, and the expansion of IL-4-secreting Th2 cells, as well as diminished IFN-gamma synthesis. IL-12 is known to induce IFN-gamma synthesis and to augment Th1 responses. In this study, therefore, we examined the potential role of IL-12 in the immunopathogenesis of AD. We show that, after stimulation with staphylococcal toxic shock syndrome toxin-1 (TSST-1) or IL-12, PBMC from patients with AD are deficient in their ability to produce IFN-gamma. PBMC from AD patients, however, produced normal quantities of IL-12 and expressed normal levels of IL-12R. Induction of IFN-gamma by TSST-1 was decreased by neutralizing anti-IL-12 Ab in normal donors, but not in AD patients. The latter observation is consistent with a defective response to IL-12 in AD PBMC. Because AD is associated with increased production of IL-4 and IL-10, we examined the effect of IL-4 on IL-12- or TSST-1-induced IFN-gamma production in normal donors. IL-4 inhibited IL-12-induced IFN-gamma production. Furthermore, Ab neutralization of IL-4 caused increased production of IFN-gamma in AD PBMC. However, neutralization of IL-10 activity caused an even greater augmentation of IFN-gamma production. Our data suggest that despite normal levels of IL-12 production and IL-12R expression, PBMC from AD patients are unable to generate normal IL-12-induced IFN-gamma responses. This defective response may be due to the excess production of IL-4 and IL-10 in this common allergic condition.     

Bacterial superantigen-induced human lymphocyte responses are nitric oxide dependent and mediated by IL-12 and IFN-gamma

Bacterial superantigens cause marked proliferation of T cells and release of lymphokines. Nitric oxide, derived from the conversion of L-arginine to L-citrulline, inhibits this activation in murine cells. We have now investigated the roles of IL-12, IFN-gamma, lymphotoxin-alpha, and nitric oxide during superantigen-induced human lymphocyte activation. Lymphocyte activation was determined by measurement of proliferative responses and lymphokine release. Both toxic shock syndrome toxin-1 from Staphylococcus aureus and recombinant streptococcal pyrogenic exotoxin A induced proliferation and production of IFN-gamma, lymphotoxin-alpha, and IL-12 by human mononuclear cells in a time-dependent fashion. The release of IFN-gamma was abrogated by a neutralizing Ab to IL-12, but lymphocyte proliferative responses were unaffected. A neutralizing Ab to IFN-gamma prevented the release of lymphotoxin-alpha, but did not affect proliferation. The neutralization of lymphotoxin-alpha using two different Abs did not affect IFN-gamma release or proliferation. In contrast to previous findings in mice, the arginine analogue, NG-monomethyl-L-arginine, significantly inhibited both proliferation and lymphokine release by superantigen-stimulated human cells. Thus, the release of lymphotoxin-alpha by lymphocytes following superantigen stimulation is dependent upon the presence of IFN-gamma; the IFN-gamma response is in turn under the control of IL-12. There is no evidence that nitric oxide plays an inhibitory role during superantigen-mediated human lymphocyte activation. Indeed, arginine is a prerequisite for such activation.     

IL-12 is not required for induction of type 1 cytokine responses in viral infections

To investigate the physiological role of IL-12 in viral infections in terms of T cell cytokine responses involved in virus-specific Ig isotype induction and in antiviral protection, immune responses elicited upon infection of IL-12-deficient mice with lymphocytic choriomeningitis virus (LCMV) or vesicular stomatitis virus (VSV) were studied. Infection of IL-12-deficient mice with LCMV induced a virus-specific type 1 cytokine response as determined by in vitro cytokine secretion patterns as well as by in vivo intracellular cytokine staining of LCMV-specific CD4+ TCR transgenic T cells that had clonally expanded in LCMV-infected IL-12-deficient recipient mice. In addition, LCMV- and VSV-specific IgG responses exhibited normal serum IgG2a/IgG1 ratios, demonstrating again virus-specific CD4+ T cell induction of type 1 phenotype in IL-12-deficient mice upon viral infection. LCMV and VSV immune mice were found to be protected against challenge immunization with recombinant vaccinia viruses expressing either the LCMV- or the VSV-derived glycoprotein, respectively. This protection is known to be mediated by T cell-secreted type 1 cytokines IFN-gamma and TNF-alpha. In contrast, IL-12-deficient mice showed impaired abilities to control infection with the facultative intracellular bacterium Listeria monocytogenes at early time points after infection. However, at later time points of infection, IL-12-deficient mice were able to clear infection. These findings may indicate that viruses are able to induce type 1 T cell responses in the absence of IL-12 as opposed to some bacterial or parasitical infections that are crucially dependent on the presence of IL-12 for the induction of type 1 immune responses.     

Streptococcal preparation OK-432 is a potent inducer of IL-12 and a T helper cell 1 dominant state

Streptococcal preparation OK-432 is a bacterial immunopotentiator extensively used in Japan for adjuvant cancer therapy. Using a C57BL/6 mouse model, OK-432 was found to induce multiple cytokines including the Th1 polarizing cytokine IL-12. Expression of IL-12 protein by murine splenocytes was restricted to macrophages and B cells and led to high levels of IFN-gamma production from both CD4+ and CD8+ T cells. Of the Th2 cytokines IL-4 and IL-10, only IL-10 protein was detected and originated primarily from the adherent cell population. Its expression was delayed relative to IL-12. A similar pattern of cytokine induction was observed from human PBMCs. OK-432-driven IFN-gamma production was inhibited by anti-IL-12 Ab, anti-IL-2 Ab, anti-TNF-alpha Ab, and anti-IL-2R alpha Ab, suggesting that IFN-gamma production from Th1 cells is induced by the cooperation action of these cytokines through the IL-2R alpha pathway. When compared with another widely used immunopotentiator bacillus Calmette-Guérin (BCG), OK-432 was a stronger IL-12 and IFN-gamma inducer. Furthermore, the mechanism of IFN-gamma induction by OK-432 differed from BCG in that coincident granulocyte-macrophage CSF and IL-1 expression played little to no role. These results suggest that OK-432 is a potent multicytokine inducer, specifically a strong inducer of IL-12, and that OK-432 may exert its antitumor effect by promoting a Th1-dominant state.     

CD40 ligand/trimer DNA enhances both humoral and cellular immune responses and induces protective immunity to infectious and tumor challenge

CD40/CD40 ligand interactions have a central role in the induction of both humoral and cellular immunity. In this study, we examined whether a plasmid expressing CD40 ligand/trimer (CD40LT) could enhance immune responses in vivo. BALB/c mice were injected with plasmid expressing beta-galactosidase DNA with or without CD40LT DNA or IL-12 DNA, and immune responses were assessed. Mice vaccinated with beta-gal DNA plus CD40LT DNA or IL-12 DNA had a striking increase in Ag-specific production of IFN-gamma, cytolytic T cell activity, and IgG2a Ab. The mechanism by which CD40LT DNA enhanced these responses was further assessed by treating vaccinated mice with anti-IL-12 mAb or CTLA-4 Ig (CTLA4Ig). Production of IFN-gamma and CTL activity was abrogated by these treatments, suggesting that CD40LT DNA was mediating its effects on IFN-gamma and CTL activity through induction of IL-12 and enhancement of B7 expression, respectively. Physiologic relevance for the ability of CD40LT DNA to enhance immune responses by the aforementioned pathways was shown in two in vivo models. First, with regard to CTL activity, mice vaccinated with CD40LT DNA did not develop metastatic tumor following challenge with lethal dose of tumor. Moreover, in a mouse model requiring IL-12-dependent production of IFN-gamma, mice vaccinated with soluble Leishmania Ag and CD40LT DNA were able to control infection with Leishmania major. These data suggest that CD40LT DNA could be a useful vaccine adjuvant for diseases requiring cellular and/or humoral immunity.     

Immunotherapy of experimental metastases by vaccination with interleukin gene-transduced adenocarcinoma cells sharing tumor-associated antigens. Comparison between IL-12 and IL-2 gene-transduced tumor cell vaccines

We have compared the therapeutic activity and characterized the antitumor response induced by IL-12 and IL-2 gene-transduced tumor cell vaccines. Mice bearing lung metastases of the BALB/c colon carcinoma C51 were treated with syngenic, histologically related, and antigenically cross-reacting irradiated IL-12 (C26/IL12) or IL-2 (C26/IL2) gene-transduced C26 tumor cells given s.c. Vaccination with C26/IL12 cells cured 40% of mice, while vaccination with C26/IL2 cells reduced the number of metastatic nodules without affecting survival. Despite this difference, similar antitumor CTL activation was shown in mice treated with C26/IL12 or C26/IL2 cells. The lytic pattern of CTL was shown to be directed to tumor-associated Ags (TAA) shared between the colon carcinomas C51, C26, and CC36 as well as with other syngenic tumors. Both treatments induced anti-TAA Abs, but only sera from mice treated with C26/IL12 contained Ab that lysed tumor cells in a C-dependent cytotoxicity assay. Early infiltration of activated T cells was found in the lungs of mice vaccinated with C26/IL12. CD4+ lymphocytes purified from the lymph nodes draining the vaccination site or from the spleen showed a higher production of IFN-gamma in response to anti-CD3 mAb in C26/IL12 vaccinated mice, while a higher production of IL-4 was shown in mice vaccinated with C26/IL2 cells. These results indicate that the better therapeutic efficacy of vaccination with C26/IL12 is associated with the production of C-binding Ab, an early infiltration of the metastatic lungs by activated T lymphocytes and a predominant systemic activation of Th1 more than Th2 cells.     

IFN-gamma-deficient mice develop experimental autoimmune uveitis in the context of a deviant effector response

Experimental autoimmune uveitis (EAU) is a T cell-mediated disease that targets the neural retina and serves as a model of human uveitis. Uveitogenic effector T cells have a Th1-like phenotype (high IFN-gamma, low IL-4), and genetic susceptibility to EAU is associated with an elevated Th1 response. Here we investigate whether the ability to produce IFN-gamma is necessary for the development of EAU by immunizing IFN-gamma-deficient (GKO) mice with the uveitogenic protein interphotoreceptor retinoid binding protein (IRBP) and characterize the associated immunologic responses. GKO mice developed EAU comparable in severity and incidence to that of their wild-type littermates. However, the cytokine profile in their uveitic eyes as well as the cytokines produced by primed lymph node cells in response to IRBP showed a distinct profile: undiminished TNF-alpha and elevated IL-5, IL-6, IL-10, and lymphotoxin (but not IL-4) responses. The inflammatory infiltrate in GKO eyes contained an excess of granulocytes and IL-5- and IL-6-producing cells, but uveitic GKO mice did not up-regulate inducible nitric oxide synthase. GKOs had enhanced lymphocyte proliferation and delayed-type hypersensitivity responses to IRBP. Histology of the delayed-type hypersensitivity lesion in GKO had superimposed elements of an allergic-like response. Anti-IRBP Ab isotypes of GKO mice showed a reduction of IgG2a, but no enhancement of IgG1. Comparison of responses in +/+ and +/- wild-type mice revealed some limited evidence of a gene-dose effect. We conclude that IFN-gamma is not required for priming of pathogenic T cells or for effecting the retinal damage and photoreceptor loss typical of EAU. However, what appears to be a grossly similar disease is caused in the GKO by a deviant type of effector response.     

Presence of activated antigen-binding B cells during immunization enhances relative levels of IFN-gamma in T cell responses

To examine the influence of Ag presentation by B cells on immune responses, we have used mice transgenic for an Ig heavy chain from a monoclonal anti-azobenzenearsonate (Ars) Ab to deliver Ag to B cells during immunization. A large proportion of transgene-expressing B cells in these mice binds Ars, while transgenic serum Ig shows poor Ars binding. Transgenic B cells present Ars proteins better than their nonhaptenated counterparts. This is associated with an increase in the proliferative responses of transgenic T cells to Ars protein immunization. Although B cell numbers in the transgenic mice are lower, many B cells in them show an activated phenotype, as identified by altered surface levels of peanut agglutinin reactivity, CD23, CD24, CD44, CD62L, and CD86. Even against nonhaptenated immunogens, transgenic responses show significant enhancement in the relative proportions of the Th1 cytokine IFN-gamma over the Th2 cytokines IL-4 and IL-10. Haptenated immunogens further enhance the predilection of transgenic mice to produce relatively more IFN-gamma. Consistent with this, there is an increase in IgG2a/IgG1 ratios in serum Abs in response to haptenated immunogens in transgenic mice. Adoptive transfer of primed hapten-specific secondary B cells into nontransgenic mice also induces an increase in relative levels of IFN-gamma in response to haptenated immunogens. Thus, presentation of immunogen in vivo by activated Ag-binding B cells contributes to enhanced immunogenicity and a Th1 cytokine bias.     

Heat-killed Listeria monocytogenes as an adjuvant converts established murine Th2-dominated immune responses into Th1-dominated responses

We investigated the capacity of heat-killed Listeria monocytogenes (HKL), a potent stimulator of the innate immune system, as a vaccine adjuvant to modify both primary and secondary Ag-specific immune responses. Mice immunized with the Ag keyhole limpet hemocyanin (KLH) mixed with HKL generated a KLH-specific primary response characterized by production of Th1 cytokines and large quantities of KLH-specific IgG2a Ab. Moreover, administration of KLH with HKL as an adjuvant reversed established immune responses dominated by the production of Th2 cytokines and high levels of KLH-specific IgE and induced a Th1-type response with high levels of IFN-gamma and IgG2a and low levels of IgE and IL-4. Neutralization of IL-12 activity at the time of HKL administration blocked the enhancement of IFN-gamma and reduction of IL-4 production, indicating that IL-12, induced by HKL, was responsible for the adjuvant effects on cytokine production. These results suggest that HKL as an adjuvant during immunization can successfully bias the development of Ag-specific cytokine synthesis toward Th1 cytokine production even in the setting of an ongoing Th2-dominated response. Thus, HKL may be clinically effective in vaccine therapies for diseases such as allergy and asthma, which require the conversion of Th2-dominated immune responses into Th1-dominated responses.     

Mice lacking the IFN-gamma receptor have impaired ability to resolve a lung eosinophilic inflammatory response associated with a prolonged capacity of T cells to exhibit a Th2 cytokine profile

To investigate the modulatory role of IFN-gamma on the induction and maintenance of Th2 mucosal immunity in vivo, experiments were performed in mice lacking the IFN-gamma R. Aerosol OVA challenge of immunized wild-type mice resulted in an infiltration of eosinophils into the lung, associated with the ex vivo production of Th2 cytokines (IL-4 and IL-5) from purified lung Thy1.2+ cells stimulated via the CD3/TCR complex. However, while immunized IFN-gamma R-deficient mice exhibited elevated levels of IgE, IgG1, and reduced levels of IgG2a compared with wild-type mice, there was no difference in the recruitment of eosinophils into the lung or the production of IL-4 and IL-5 from lung T cells on day 3. In contrast, up to 2 mo after a single Ag challenge, eosinophils were still present in the lungs of IFN-gamma R-deficient, but not wild-type, mice. Likewise, lung-derived T cells from IFN-gamma R-deficient mice produced higher levels of IL-4 and IL-5, both at 1 and 2 mo after OVA challenge compared with T cells from wild-type mice. We conclude that endogenous IFN-gamma regulates the humoral isotype Ab pattern, but does not modulate the commitment of T cells to a Th2 phenotype in vivo or the acute infiltration of eosinophils to the lung. However, in the absence of IFN-gamma-mediated signaling, there is a transition from a spontaneously resolving to a persisting eosinophilic inflammation of the lungs, associated with a sustained capacity of lung T cells to secrete a Th2 cytokine profile.