Basophils amplify type 2 immune responses, but do not serve a protective role, during chronic infection of mice with the filarial nematode Litomosoides sigmodontis

Chronic helminth infections induce a type 2 immune response characterized by eosinophilia, high levels of IgE, and increased T cell production of type 2 cytokines. Because basophils have been shown to be substantial contributors of IL-4 in helminth infections, and because basophils are capable of inducing Th2 differentiation of CD4(+) T cells and IgE isotype switching in B cells, we hypothesized that basophils function to amplify type 2 immune responses in chronic helminth infection. To test this, we evaluated basophil function using the Litomosoides sigmodontis filaria model of chronic helminth infection in BALB/c mice. Time-course studies showed that eosinophilia, parasite Ag-specific CD4(+) T cell production of IL-4 and IL-5 and basophil activation and IL-4 production in response to parasite Ag all peak late (6-8 wk) in the course of L. sigmodontis infection, after parasite-specific IgE has become detectable. Mixed-gender and single-sex worm implantation experiments demonstrated that the relatively late peak of these responses was not dependent on the appearance of circulating microfilariae, but may be due to initial low levels of parasite Ag load and/or habitation of the developing worms in the pleural space. Depletion of basophils throughout the course of L. sigmodontis infection caused significant decreases in total and parasite-specific IgE, eosinophilia, and parasite Ag-driven CD4(+) T cell proliferation and IL-4 production, but did not alter total worm numbers. These results demonstrate that basophils amplify type 2 immune responses, but do not serve a protective role, in chronic infection of mice with the filarial nematode L. sigmodontis.     

Parasite antigen-driven basophils are a major source of IL-4 in human filarial infections

Basophil contribution to the IL-4 pool in filarial infections was assessed using PBMC from 20 patients with active filarial infections and from 9 uninfected subjects. Patient basophils released histamine in response to Brugia malayi Ag (BmAg). They also released IL-4 within 2 h after exposure to BmAg, as assessed by intracellular cytokine flow cytometry. This IL-4 induction was Ag specific, as IL-4 was not detected in BmAg-exposed basophils obtained from uninfected subjects. Although there were, on average, 64 times more CD4(+) T cells than basophils in the peripheral circulation of filaria-infected patients, the absolute numbers of basophils and CD4(+) T cells producing IL-4 per 100000 PBMC were equivalent (geometric mean: 16 IL-4-producing basophils/100000 PBMC vs 22 IL-4-producing CD4(+) T cells/100000 PBMC). Basophils also released IL-4 in response to both low and high concentrations of BmAg, whereas CD4(+) T cells released IL-4 only after incubation with a high concentration of BmAg, raising the possibility that basophils, due to their lower threshold for activation, may actually release IL-4 more frequently than CD4(+) T cells in vivo. Furthermore, IL-4 production in vitro by Ag-stimulated purified basophils or CD4(+) T cells provided evidence that basophils release greater quantities of IL-4 per cell than CD4(+) T cells in response to BmAg. These results suggest that, when Ag-specific IgE is present in a filaria-infected individual, basophils function to amplify the ongoing Th2 response by releasing IL-4 in greater amounts and possibly more frequently than CD4(+) T cells in response to filarial Ag.     

Long term maintenance of IgE-mediated memory in mast cells in the absence of detectable serum IgE

Mast cells and basophils involved in allergic responses do not have clonotypic Ag receptors. However, they can acquire Ag specificity through binding of Ag-specific IgE to FcepsilonRI expressed on their surface. Previous studies demonstrated that IgE binding induced the stabilization and accumulation of FcepsilonRI on the cell surface and resulted in up-regulation of FcepsilonRI. In this study we have further analyzed the maintenance of IgE-mediated memory in mast cells and basophils in vivo by comparing kinetics of serum IgE levels, FcepsilonRI expression, and ability to induce systemic anaphylaxis. A single i.v. injection of trinitrophenyl-specific IgE induced 8-fold up-regulation of FcepsilonRI expression on peritoneal mast cells in B cell-deficient (micro m(-/-)) mice. Serum IgE levels became undetectable by day 6, but the treatment of mice with anti-IgE mAb induced a significant drop in body temperature on days 14, 28, and 42. The administration of trinitrophenyl -BSA, but not BSA, in place of anti-IgE mAb gave similar results, indicating the Ag specificity of the allergic response. This long term maintenance of Ag-specific reactivity in the allergic response was also observed in normal mice passively sensitized with IgE even though the duration was shorter than that in B cell-deficient mice. The appearance of IgE with a different specificity did not interfere with the maintenance of IgE-mediated memory of mast cells and basophils. These results suggest that IgE-mediated stabilization and up-regulation of FcepsilonRI enables mast cells and basophils not only to acquire Ag specificity, but also to maintain memory in vivo for lengthy periods of time.     

Humoral immune functions in IL-4 transgenic mice

We have analyzed mice expressing IL-4 as a transgene, and found that expression of this lymphokine has profound effects on B cell function. B cells from transgenic mice exhibit phenotypic changes, including an increase in size and elevated expression of class II MHC. IL-4 increases the quantity of IgE produced by transgenic-derived B cells in response to LPS stimulation. In vivo, IL-4 markedly affects the serum Ig isotype repertoire. Serum levels of IgG1 and IgE are elevated, and levels of IgG2a, IgG2b, and IgG3 are depressed in IL-4 transgenic mice. Ag-specific antibody responses to immunization with hapten-carrier conjugates are also affected by IL-4. Transgenic mice show increased anti-hapten IgE and IgG1 and reduced anti-hapten IgG2a, IgG2b, and IgG3, compared with wild-type mice. Ag-specific IgE is substantially induced by T cell-dependent Ag, but not T cell-independent Ag, suggesting that cognate T-B interactions in addition to IL-4 are required for generating IgE responses in vivo. In vivo treatment with the anti-IL-4 mAb 11B11 reverses many of the isotype alterations in the transgenic mice, indicating that these changes arise as a direct consequence of IL-4 secretion.     

Dual mechanisms of potentiation of murine antigen-specific IgE production by cyclosporin A in vitro.

Concomitant administration of cyclosporin A (CsA) with Ag has been shown to augment the production of Ag-specific IgE in vivo. We demonstrate that addition of CsA also markedly potentiated Ag-specific IgE in vitro. Low doses of CsA (3 and 10 ng/ml) added at the time of culture initiation selectively enhanced Ag-specific IgE but not IgA or IgG1 production, whereas higher doses (30 ng/ml) suppressed production of all the isotypes. Augmented IgE production was found to correlate with enhanced production of IL-4 and diminished production of IFN-gamma. Delayed addition (after 2 days) of low doses of CsA to Ag-stimulated cultures did not potentiate IgE production, even though CsA differentially affected levels of IL-4 and IFN-gamma. CsA enhanced Ag-mediated cognate T/B interaction was not affected by neutralizing doses of anti-IL-4, suggesting Ag-mediated lymphocytic "synapses" may be inaccessible to anti-IL-4. The effect of CsA on Ag presentation was determined by pulsing peritoneal exudate cells, spleen cells, or primed B cells with Ag and low doses of CsA before incubation with primed splenocytes. Enhanced Ag-specific IgE responses were detected with no effect on IL-4 or IFN-gamma levels. Thus, our study indicates that CsA potentiation of Ag-specific IgE response is due to cumulative action of CsA on two independent pathways: first, CsA differentially modulates IL-4 and IFN-gamma levels during the early phase of cognate Th2/B cell interaction; and second, CsA directly affects APC and IgE isotype-specific amplifying cellular components without apparently affecting the secretory levels of IL-4 and IFN-gamma. Dual mechanisms of CsA-potentiated IgE production are consistent with the hypothesis of two-tiered T cell regulation of Ag-specific IgE responses.     

Restoration of the antibody response to IgE/antigen complexes in CD23-deficient mice by CD23+ spleen or bone marrow cells

Mice immunized with IgE/Ag complexes produce significantly more Ag-specific Abs than mice immunized with Ag alone. The enhancement is mediated via the low-affinity receptor for IgE (FcepsilonRII or CD23), as shown by its complete absence in mice pretreated with mAbs specific for CD23 and in CD23-deficient mice. Because the constitutive expression of murine CD23 is limited to B cells and follicular dendritic cells (FDCs), one of these cell types is likely to be involved. One of the suggested modes of action of IgE/CD23 is to increase the ability of B cells to present Ag to T cells, as demonstrated to take place in vitro. Another possibility is that FDCs capture the IgE/Ag complexes and present these directly to B cells. The purpose of the present study was to determine whether CD23+ B cells or FDCs are responsible for the IgE/CD23-mediated enhancement of specific Ab responses in vivo. We show that the enhancement is completely restored in irradiated CD23-deficient mice reconstituted with CD23+ spleen or bone marrow cells. In these mice, the B cells are CD23+ and the FDCs are presumably CD23- because the FDCs are radiation resistant and are reported not to be replaced by donor cells after this type of cell transfer. In contrast, enhancement was not restored in irradiated wild-type mice reconstituted with CD23- cells. These results indicate that CD23+ B cells, and not FDCs, are the cells that capture IgE/Ag complexes and induce enhancement of Ab responses in vivo.     

Regulation of IgE production requires oligomerization of CD23.

Here we describe the production of a rabbit polyclonal Ab (RAS1) raised against the stalk of murine CD23. RAS1 inhibits release of CD23 from the surface of both M12 and B cells resulting in an increase of CD23 on the cell surface. Despite this increase, these cells are unable to bind IgE as determined by FACS. CD23 has previously been shown to bind IgE with both a high (4-10 x 10(7) M(-1)) and low (4-10 x 10(6) M(-1)) affinity. Closer examination by direct binding of (125)I-IgE revealed that RAS1 blocks high affinity binding while having no effect on low affinity binding. These data support the model proposing that oligomers of CD23 mediate high affinity IgE binding. These experiments suggest that RAS1 binding to cell surface CD23 results in a shift from oligomers to monomers, which, according to the model, only bind IgE with low affinity. These experiments also suggest that high affinity binding of IgE is required for IgE regulation by CD23 and is demonstrated by the fact that treatment of Ag/Alum-immunized mice treated with RAS1 results in a significant increase in IgE production similar to the levels seen in CD23-deficient mice. These mice also had significantly decreased levels of serum soluble CD23 and Ag-specific IgG1. RAS1 had no effect on IgE or Ag-specific IgG1 production in CD23-deficient mice.     

Identification of semaphorin 4B as a negative regulator of basophil-mediated immune responses

Basophils are strong mediators of Th2 responses during helminthic infections. Recently, basophils were shown to function as APCs and promote both Th2 skewing and humoral memory responses. However, the mechanisms that regulate basophils are still unclear. In this article, we show that a class IV semaphorin, Sema4B, negatively regulates basophil functions through T cell-basophil contacts. In a screen to identify semaphorins that function in the immune system, we determined that Sema4B is expressed in T and B cells. Interestingly, Sema4B(-/-) mice had considerably increased serum IgE levels despite normal lymphocyte and dendritic cell functions. Recombinant Sema4B significantly inhibited IL-4 and IL-6 production from basophils in response to various stimuli, including IL-3, papain, and FcεRI cross-linking. In addition, T cell-derived Sema4B, which accumulated at contact sites between basophils and CD4(+) T cells, suppressed basophil-mediated Th2 skewing, suggesting that Sema4B regulates basophil responses through cognate cell-cell contacts. Furthermore, Sema4B(-/-) mice had enhanced basophil-mediated memory IgE production, which was abolished by treating with an anti-FcεRIα Ab. Collectively, these results indicate that Sema4B negatively regulates basophil-mediated Th2 and humoral memory responses.     

CD23-bound IgE augments and dominates recall responses through human naive B cells

Human peripheral blood BCRμ(+) B cells express high levels of CD23 and circulate preloaded with IgE. The Ag specificity of CD23-bound IgE presumably differs from the BCR and likely reflects the Ag-specific mix of free serum IgE. CD23-bound IgE is thought to enhance B cell Ag presentation to T cells raising the question of how a B cell might respond when presented with a broad mix of Ags and CD23-bound IgE specificities. We recently reported that an increase in CD23(+) B cells is associated with the development of resistance to schistosomiasis, highlighting the potential importance of CD23-bound IgE in mediating immunity. We sought to determine the relationship between BCR and CD23-bound IgE-mediated B cell activation in the context of schistosomiasis. We found that crude schistosome Ags downregulate basal B cell activation levels in individuals hyperexposed to infectious worms. Schistosome-specific IgE from resistant, occupationally exposed Kenyans recovered responses of B cells to schistosome Ag. Furthermore, cross-linking of CD23 overrode intracellular signals mediated via the BCR, illustrating its critical and dominating role in B cell activation. These results suggest that CD23-bound IgE augments and dominates recall responses through naive B cells.     

An antigen-specific DTH-initiating cell clone. Functional, phenotypical, and partial molecular characterization

The elicitation of delayed-type hypersensitivity (DTH) reactions in mice is due to the sequential action of two different Ag-specific Thy-1+ cells. An early-acting DTH-initiating cell in the lymphoid organs produces a circulating, Ag-specific factor that is functionally analogous to IgE antibody and initiates DTH by sensitizing the local tissue for release of the vasoactive amine serotonin. In picryl chloride (PC1) or oxazolone (OX) contact sensitivity, this DTH-initiating factor is called PC1-F and OX-F respectively, and is Ag-specific, but MHC-unrestricted. The phenotype of polyclonal DTH-initiating cells was recently shown to be unusual for an Ag-specific cell. The phenotype was: Thy-1+, Lyt-1+ (CD5), triple negative (CD4-, CD8-, and CD3-), B220+ (Ly-5, CD45RA), positive for IL-3 receptors, but not IL-2 receptors, and positive for antibodies that react with a putative constant or framework portion of DTH-initiating factors such as anti-PC1-F antibodies and 14-30 mAb. We report here the generation of an Ag-specific DTH-initiating cell clone from nude mice that were immunized and boosted by contact sensitization with OX. By flow microfluorometry analysis, this clone has a similar unique surface phenotype, and by in vivo assay has the same functional abilities, as polyclonal DTH-initiating cells. The clone produces Ag-specific OX-F that acts in an Ag-specific manner to initiate DTH. Moreover, specific cDNA probes and Northern blot analysis of the clone demonstrated that the Ag-specific DTH-initiating cells are Thy-1+, CD3-, and IL-3R+. Thus, DTH initiation is due to an Ag-specific lymphoid cell, that produces an Ag-specific factor, and that has a unique surface phenotype for Ag-specific cells; namely, Thy-1+, CD5+, sIg-, CD4-, CD8-, CD3-, CD45RA+, IL-2R-, and IL-3R+.     

Long-term CD4+ memory T cells from the spleen lack MEL-14, the lymph node homing receptor.

We have characterized the surface phenotype and function of long-lived, Ag-specific memory CD4+ T cells generated in vivo by immunization with keyhole limpet hemocyanin (KLH). CD4+ T cells from the spleens of mice primed more than 2 mo previously with KLH, produced high levels of IL-2 and IL-3, and low levels of IL-4 and IFN-gamma in response to in vitro restimulation with specific Ag. The KLH-primed T cells mediated carrier-specific helper activity for the antibody production by NIP-primed B cells in secondary in vitro responses to NIP-KLH. Subsets of CD4+ T cells from KLH-primed mice were isolated on the basis of surface CD45RB (23G2) by magnetic separation and were examined for functional capacity in several assays of Ag-specific recall. Virtually all of the secretion of IL-2, IL-3, IL-4, and IFN-gamma in response to restimulation with Ag in vitro was associated with, and considerably enriched in, the CD45RB- subset of CD4+ T cells. Similarly, carrier-specific helper function and Ag-specific proliferation in vitro were also confined to the CD45RB-, CD4+ subset of T cells, confirming the previous association of this surface phenotype with memory Th cell activity. We also examined expression of the lymphocyte homing receptor, MEL-14 (gp90MEL), which is required for lymphocyte extravasation to peripheral lymph nodes and is present in high levels on naive T cells. MEL-14 positive and negative subsets of CD4+ T cells from long term KLH-primed mice were evaluated for Ag-specific memory function in terms of lymphokine production, Ag-induced proliferation, and helper activity. Each of these functions was associated exclusively with the MEL-14- subset of CD4+ T cells, which exhibited responses comparable to the CD45RB- subset. These data indicate that memory Th cell function in the spleen is contained within the MEL-14-, CD45RB- subset of CD4+ T cells and suggest that memory helper cells may have different patterns of recirculation from naive T cells.     

Soluble forms of CD40 inhibit biologic responses of human B cells.

We have expressed the CD40 surface Ag as both a soluble 28-kDa molecule and a 57-kDa Fc fusion protein containing the human IgG1 Fc region. Soluble CD40 and the Fc fusion protein inhibited the proliferative response of anti-IgM-activated human B cells to the CD40 mAb G28-5. Similarly, G28-5- and IL-4-induced IgE secretion from PBMC depleted of T cells was effectively blocked by both forms of soluble CD40. Although the soluble constructs of CD40 had only a minimal inhibitory effect on IL-4-mediated proliferation of anti-IgM-activated B cells, IL-4-induced soluble CD23 shedding from both PBMC and T cells depleted of PBMC, and IgE secretion from PBMC, were significantly reduced in a concentration-dependent manner when soluble CD40 was present in the culture. The data presented demonstrate that both soluble forms of the CD40 molecule are biologically active, and suggest that the ligand for CD40 is inducible in IL-4-stimulated cultures and that it mediates both shedding of sCD23 and IgE secretion.     

Functional responses and costimulator dependence of memory CD4+ T cells

To examine the functional characteristics of memory CD4+ T cells, we used an adoptive transfer system to generate a stable population of Ag-specific memory cells in vivo and compared their responses to Ag with those of a similar population of Ag-specific naive cells. Memory cells localized to the spleen and lymph nodes of mice and exhibited extremely rapid recall responses to Ag in vivo, leaving the spleen within 3-5 days of Ag encounter. Unlike their naive counterparts, memory cells produced effector cytokines (IFN-gamma, IL-4, IL-5) within 12-24 h of Ag exposure and did not require multiple cycles of cell division to do so. Memory cells proliferated at lower Ag concentrations than did naive cells, were less dependent on costimulation by B7 molecules, and independent of costimulation by CD40. Furthermore, effector cytokine production by memory cells also occurred in the absence of either B7 or CD40 costimulation. Lastly, memory cells were resistant to tolerance induction. Together, these findings suggest that the threshold for activation of memory CD4+ cells is lower than that of naive cells. This would permit memory cells to rapidly express their effector functions in vivo earlier in the course of a secondary immune response, when the levels of Ag and the availability of costimulation may be relatively low.     

Cutting edge: IPSE/alpha-1, a glycoprotein from Schistosoma mansoni eggs, induces IgE-dependent, antigen-independent IL-4 production by murine basophils in vivo

During infection with the helminth parasite Schistosoma mansoni, the deposition of eggs coincides with the onset of IL-4 production and Th2 development. Although IL-4 is known as a potent inducer of Th2 differentiation, the mechanism by which schistosome eggs induce IL-4 production is not clear. In this study, we demonstrate that the S. mansoni egg Ag (SmEA) induces IgE-dependent IL-4 production by basophils derived from Heligmosomoides polygyrus-infected or OVA/alum-immunized mice in the absence of pathogen-specific IgE. The effect is mediated by the secretory glycoprotein IPSE/alpha-1, because IPSE/alpha-1-depleted SmEA no longer induces cytokine production. Conversely, recombinant IPSE/alpha-1 is sufficient to induce IL-4 production. Importantly, the injection of SmEA or recombinant IPSE/alpha-1 into H. polygyrus-infected 4get/KN2 IL-4 reporter mice rapidly induces the dose-dependent IL-4 production by basophils in the liver, a major site of egg deposition. Thus, IPSE/alpha-1 induces basophils to produce IL-4 even in the absence of Ag-specific IgE.     

SHIP represses Th2 skewing by inhibiting IL-4 production from basophils

We report that SHIP(-/-) mice, compared to SHIP(+/+) mice, are Th2 skewed with elevated serum IgE and twice as many splenic CD4(+) Th2 cells that, when stimulated with anti-CD3, produce more IL-4 and less IFN-γ. Exploring the reason for this Th2 skewing, we found that freshly isolated SHIP(-/-) splenic and bone marrow basophils are present in elevated numbers and secrete far more IL-4 in response to IL-3 or to FcεRI stimulation than do WT basophils. These SHIP(-/-) basophils markedly skew wild-type macrophage colony stimulating factor-derived macrophages toward an M2 phenotype, stimulate OT-II CD4(+) Th cells to differentiate into Th2 cells, and trigger SHIP(+/+) B cells to become IgE-producing cells. All these effects are completely abrogated with neutralizing anti-IL-4 Ab. Exploring the cell signaling pathways responsible for hyperproduction of IL-4 by SHIP(-/-) basophils, we found that IL-3-induced activation of the PI3K pathway is significantly enhanced and that PI3K inhibitors, especially a p110α inhibitor, dramatically suppresses IL-4 production from these cells. In vivo studies, in which basophils were depleted from mast cell-deficient SHIP(+/+) and SHIP(-/-) mice, confirmed the central role that basophils play in the Th2 skewing of naive SHIP-deficient mice. Taken together, these studies demonstrate that SHIP is a potent negative regulator of IL-4 production from basophils and thus may be a novel therapeutic target for Th1- and Th2-related diseases.     

Vaccination with heat-killed Listeria as adjuvant reverses established allergen-induced airway hyperreactivity and inflammation: role of CD8+ T cells and IL-18

Asthma is a respiratory disorder characterized by airway hyperreactivity (AHR) and inflammation and is associated with high serum IgE and overproduction of IL-4, IL-5, and IL-13 by allergen-specific Th2 cells. Our previous studies demonstrated that heat-killed Listeria monocytogenes (HKL) as an adjuvant in immunotherapy successfully reversed ongoing Ag-specific Th2-dominated responses toward Th1-dominated responses, but it was unclear if such immune modulation could reverse ongoing, established disease in target organs such as the lung. In this paper we show that a single dose of Ag plus HKL as adjuvant significantly reduced AHR in a murine model for asthma and reversed established AHR when given late after allergen sensitization. HKL as adjuvant also dramatically inhibited airway inflammation, eosinophilia, and mucus production, significantly reduced Ag-specific IgE and IL-4 production, and dramatically increased Ag-specific IFN-gamma synthesis. The inhibitory effect of HKL on AHR depended on the presence of IL-12 and CD8+ T cells and was associated with an increase of IL-18 mRNA expression. Thus, our results demonstrate that HKL as an adjuvant for immunotherapy mediates immune deviation from a pathological Th2-dominated response toward a protective immune response in peripheral lymphoid tissues and in the lungs and may be clinically effective in the treatment of patients with established asthma and allergic disease.     

Cutting edge: Helminth infection induces IgE in the absence of mu- or delta-chain expression

Infections with helminth parasites are associated with an IgE isotype switch and high serum IgE concentrations. IgE is rapidly bound by the high affinity IgE receptor (Fc epsilonRI), thereby sensitizing Fc epsilonRI-bearing basophils and mast cells for IgE-inducible effector functions such as IL-4 production. The development of Ab-secreting B cells is dependent on IgM and consequently, muMT mice, which lack surface IgM, are considered devoid of Abs. In this study we report the unexpected finding that C57BL/6 muMT mice generate robust IgE responses upon infection with three distinct helminth parasites, Heligmosomoides polygyrus, Trichuris muris, and Schistosoma mansoni. IgE is produced despite an apparent block in B cell development and licenses basophils for IgE-induced IL-4 production. Our findings reveal the existence of an evolutionarily conserved, IgM-independent pathway for the production of IgE upon infection with helminth parasites.     

Induction of negative signal through CD2 during antigen-specific T cell activation.

We have investigated the role of CD2 molecules in Ag-specific T cell activation by using a mouse model system in which the function of CD2 can be analyzed without the apparent influence of major accessory molecules, such as CD4 or LFA-1. Transfection of the CD2 gene into a CD2- T cell hybridoma confers the enhancement of IL-2 production upon Ag stimulation. Anti-CD2 mAb inhibits the Ag-specific response of the CD2-transfectant, not only to the level of CD2- cells but to the background. B cells, but not MHC class II-transfected L cells, serve as APC to induce the inhibition of Ag response. The complete abrogation of the response is observed only upon the stimulation through TCR with Ag in the presence of APC but not through either TCR-CD3 or other molecules such as Thy-1. Furthermore, the inhibition can also be observed when anti-CD2 mAb is immobilized on culture plates, suggesting that the inhibition of Ag response results from transducing the negative signal through the CD2 molecule. The experiments on cytoplasmic domain-deleted CD2-transfected T cells reveal that the cytoplasmic portion is responsible for the CD2-mediated abrogation of Ag responses. These results imply that CD2 has important roles in T cell responses not only as an activation and adhesion molecule but also as a regulatory molecule of Ag-specific responses through the TCR.     

In situ analysis reveals physical interactions between CD11b+ dendritic cells and antigen-specific CD4 T cells after subcutaneous injection of antigen

In situ staining techniques were used to visualize physical interactions between dendritic cell subsets and naive Ag-specific CD4 T cells in the lymph node. Before injection of Ag, CD8(+) dendritic cells and naive OVA-specific CD4 T cells were uniformly distributed throughout the T cell-rich paracortex, whereas CD11b(+) dendritic cells were located mainly in the outer edges of the paracortex near the B cell-rich follicles. Many OVA-specific CD4 T cells were in contact with CD8(+) dendritic cells in the absence of OVA. Within 24 h after s.c. injection of soluble OVA, the OVA-specific CD4 T cells redistributed to the outer paracortex and interacted with CD11b(+), but not CD8(+) dendritic cells. This behavior correlated with the uptake of OVA and the presence of peptide-MHC complexes on the surface of CD11b(+) dendritic cells, and subsequent IL-2 production by the Ag-specific CD4 T cells. These results are consistent with the possibility that CD11b(+) dendritic cells play a central role in the activation of CD4 T cells in response to s.c. Ag.     

Cutting edge: activation of NK T cells by CD1d and alpha-galactosylceramide directs conventional T cells to the acquisition of a Th2 phenotype.

NK T cells recognize glycolipid Ags such as alpha-galactosylceramide (alpha-GalCer) presented by the MHC class I-like molecule CD1d. In this paper we have studied the in vivo effects of alpha-GalCer on the generation of adaptive immune responses. Treatment of mice with alpha-GalCer resulted in rapid activation of NK T cells and production of the cytokines IL-4 and IFN-gamma. However, after this initial stimulation, NK T cells became polarized for the production of IL-4. Further, as soon as 6 days after alpha-GalCer injection, a marked increase in serum IgE levels was observed. Administration of alpha-GalCer at the time of priming of mice with protein Ag resulted in the generation of Ag-specific Th2 cells and a profound increase in the production of IgE. Collectively, these findings indicate that alpha-GalCer may be useful for modulating immune responses toward a Th2 phenotype during prophylaxis and therapy.