The Effect of IFN-gamma, Alum and Complete Freund Adjuvant on TNP-KLH Induced Ig.G(1), IgE and IgG(2a) Responses in Mice

Adjuvants are considered to play an important role in directing the isotype and amount of antibodies produced upon immunization by conducting the development of either Th-1 or Th-2 cells upon T-cell stimulation. This is based on the different cytokine production patterns that were observed after in vitro resttmulation of T cells isolated from mice immunized with antigen either adsorbed on alum or emulsified in complete Freund adjuvant (CFA). However, other studies suggest that primarily the type of antigen determines which isotypes are produced and to what extent. In these studies, however, IgE was not determined. Therefore, this study examined whether alum and CFA influenced the amount and/or ratio of IgG(1), IgE and IgG(2a) produced after TNP-KLH immunization. Similar levels of IgG(1), IgE and IgG(2a) antibodies were found upon immunization with TNP-KLH either adsorbed on alum or emulsified in CFA. Moreover, administration of IFN-gamma in combination with TNP-KLH adsorbed on alum did not increase the amount of IgG(2a) produced. IFN-gamma treatment resulted in an increased IL-6 and decreased IFN-gamma production by spleen cells upon Con A stimulation, whereas it did not change the IL-4 production in similar conditions. The presented results suggest that upon immunization with TNP-KLH high IL-4 levels are produced, resulting in an antibody response that is dominated by IgG(1), independent of the adjuvant employed. The IL-4 inducing property of TNP-KLH is substantiated by the finding that repeated immunization of mice with TNP-KI, without adjuvant, increases the serum total IgE level. The presented data suggest that the carrier part of TNP-KLH preferentially results in Th-2 cell activity after which the adjuvant merely enhances the antibody responses generated.     

The polyclonal and antigen-specific IgE and IgG subclass response of mice injected with ovalbumin in alum or complete Freund's adjuvant

BALB/c mice were injected ip with 1 microgram ovalbumin (OVA) in alum or complete Freund's adjuvant (cFA) and the changes of the IgE and IgG subclass serum levels and isotypes of the anti-OVA specific antibodies determined by radioimmunoassays. By Day 10, OVA in alum had induced a 5- to 10-fold increase of the IgE serum level and an initial decrease of the IgG subclass levels which subsequently increased to two to threefold over the preinjection level. OVA in cFA induced a gradual twofold increase of the IgE serum level, a rapid fourfold increase of the IgG2a level occurring by Day 7, and a gradual two to threefold increase of the other IgG subclasses. Over 90% of the anti-OVA antibodies were of the IgGl isotype with both adjuvants; OVA in alum induced slightly more IgGl anti-OVA antibodies than cFA. In contrast, the OVA in alum injected mice formed significantly more (5- to 10-fold) IgE anti-OVA antibodies than the cFA-injected mice. OVA in alum also induced a large nonspecific increase of the IgE serum level because only approximately 40% of the increase observed on Day 14 was absorbable with OVA, whereas approximately 90% the IgE increase in cFA injected mice was absorbable with OVA. The data demonstrate that mice form mainly IgGl and IgE antibodies to OVA irrespective of the adjuvant. The low specific and lack of nonspecific IgE formation by mice injected with OVA in cFA may be the result of cFA-induced interferon-gamma (IFN-gamma) production because IFN-gamma has been shown to stimulate IgG2a and inhibit IgE secretion in vitro.     

Relationships between cell-mediated immunity and the IgE antibody response: II. Delayed hypersensitivity and antibody production to DNP-Ascaris conjugates

Rats of the W/F strain were immunized with DNP-Ascaris conjugates using complete Freund's adjuvant (CFA), Al(OH)₃ gel (alum), or B. pertussis vaccine as adjuvants. Cell-mediated immunity was assessed by lymphotoxin in vitro and by delayed hypersensitivity in vivo. IgE and IgG antibody determinations were made on serum pools obtained at various times during the primary and secondary responses. Although delayed hypersensitivity appeared earlier than lymphotoxin, these two parameters correlated during the primary but not during the secondary response. The discrepancies suggested that different cells may be responsible for these two phenomena. Antibody production was influenced by the adjuvant used. CFA led to IgG antibody responses to both hapten and carrier but not to IgE antibody production. The use of B. pertussis resulted in both IgE and IgG antibody production. In the case of alum, anti-hapten antibodies appeared during the primary response while anti-carrier antibodies of both IgE and IgG classes were detected after booster. The results indicated that cell-mediated immunity, IgE, and IgG antibodies appeared independently in an ordered, temporal sequence, and that these responses were not mutually exclusive but were under strong modulatory influences of the various adjuvants used.     

IgE immune complexes stimulate an increase in lung mast cell progenitors in a mouse model of allergic airway inflammation

Mast cell numbers and allergen specific IgE are increased in the lungs of patients with allergic asthma and this can be reproduced in mouse models. The increased number of mast cells is likely due to recruitment of mast cell progenitors that mature in situ. We hypothesized that formation of IgE immune complexes in the lungs of sensitized mice increase the migration of mast cell progenitors to this organ. To study this, a model of allergic airway inflammation where mice were immunized with ovalbumin (OVA) in alum twice followed by three daily intranasal challenges of either OVA coupled to trinitrophenyl (TNP) alone or as immune complexes with IgE-anti-TNP, was used. Mast cell progenitors were quantified by a limiting dilution assay. IgE immune complex challenge of sensitized mice elicited three times more mast cell progenitors per lung than challenge with the same dose of antigen alone. This dose of antigen challenge alone did not increase the levels of mast cell progenitors compared to unchallenged mice. IgE immune complex challenge of sensitized mice also enhanced the frequency of mast cell progenitors per 10(6) mononuclear cells by 2.1-fold. The enhancement of lung mast cell progenitors by IgE immune complex challenge was lost in FcRγ deficient mice but not in CD23 deficient mice. Our data show that IgE immune complex challenge enhances the number of mast cell progenitors in the lung through activation of an Fc receptor associated with the FcRγ chain. This most likely takes place via activation of FcεRI, although activation via FcγRIV or a combination of the two receptors cannot be excluded. IgE immune complex-mediated enhancement of lung MCp numbers is a new reason to target IgE in therapies against allergic asthma.     

Enhanced protective efficacy of a chimeric form of the schistosomiasis vaccine antigen Sm-TSP-2

The large extracellular loop of the Schistosoma mansoni tetraspanin, Sm-TSP-2, when fused to a thioredoxin partner and formulated with Freund's adjuvants, has been shown to be an efficacious vaccine against murine schistosomiasis. Moreover, Sm-TSP-2 is uniquely recognised by IgG(1) and IgG(3) from putatively resistant individuals resident in S. mansoni endemic areas in Brazil. In the present study, we expressed Sm-TSP-2 at high yield and in soluble form in E. coli without the need for a solubility enhancing fusion partner. We also expressed in E. coli a chimera called Sm-TSP-2/5B, which consisted of Sm-TSP-2 fused to the immunogenic 5B region of the hookworm aspartic protease and vaccine antigen, Na-APR-1. Sm-TSP-2 formulated with alum/CpG showed significant reductions in adult worm and liver egg burdens in two separate murine schistosomiasis challenge studies. Sm-TSP-2/5B afforded significantly greater protection than Sm-TSP-2 alone when both antigens were formulated with alum/CpG. The enhanced protection obtained with the chimeric fusion protein was associated with increased production of anti-Sm-TSP-2 antibodies and IL-4, IL-10 and IFN-γ from spleen cells of vaccinated animals. Sera from 666 individuals from Brazil who were infected with S. mansoni were screened for potentially deleterious IgE responses to Sm-TSP-2. Anti-Sm-TSP-2 IgE to this protein was not detected (also shown previously for Na-APR-1), suggesting that the chimeric antigen Sm-TSP-2/5B could be used to safely and effectively vaccinate people in areas where schistosomes and hookworms are endemic.     

Antigen concentration determines helper T cell subset participation in IgE antibody responses.

A recently developed in vitro system for antigen-stimulated primary and secondary murine IgE antibody responses has been used to define (a) the relative participation of the Th1 and Th2 cell-derived lymphokines IFN-gamma and IL-4, respectively, in such responses, and (b) the role of antigen concentration in determining functional helper T cell activity. These studies confirm that IL-4 and IFN-gamma exert regulatory effects on IgE synthesis, but the nature and extent of their respective effects on primary and secondary IgE responses differ. Thus, primary IgE responses are considerably more sensitive to and dependent on IL-4 than are secondary IgE responses since (1) anti-IL-4 monoclonal antibody totally inhibited primary IgE responses, but only partially affected secondary responses; and (2) exogenously added IL-4 could stimulate primary IgE responses to optimal antigen concentrations, but had no effect on secondary IgE production. Likewise, antigen-stimulated primary IgE responses are about eightfold more sensitive than are secondary responses to the inhibitory effects of IFN-gamma. Studying the effect of antigen dose on the quantity of IgE antibody produced revealed that although IFN-gamma could be detected by ELISA in cultures exhibiting high-dose antigen-dependent diminution of IgE production, anti-IFN-gamma monoclonal antibody could not reverse this phenomenon. Thus, IFN-gamma is not solely responsible for decreased IgE synthesis associated with high-dose antigen exposure. IL-4 activity was detected in the fluid from cultures stimulated with low, but not high, levels of antigen. Moreover, addition of exogenous IL-4 restored IgE production to normal levels in cultures exposed to high antigen concentrations. Therefore, it appears that high levels of antigen result in selective stimulation of Th1 cells which produce IFN-gamma, and diminished activation of IL-4-producing Th2 cells. These results help explain observations regarding the influence of antigen dose on the generation of experimental and clinical IgE antibody responses in vivo.     

IgE antibody against hepatitis B surface antigen in mice

Hepatitis B surface antigen (HBs antigen) was examined for elicitation of IgE production by injection into mice. The Prausnitz-Küstner (PK)-type skin test in the rat was employed for detection of IgE antibody to HBs antigen, because no sufficient purified HBs antigen was available as the challenging antigen for the passive cutaneous anaphylaxis (PCA) test in rats. The positive PK test was considered to be due to IgE antibody, since the active principle was inactivated by heating the sera at 56 C for 30 min, did not bind to protein A and was eliminated by anti-mouse IgE antisera. These data indicate that the PK-type test in rats can be used for detection of mouse IgE antibody when the amount of a test sample is not sufficient for the PCA test in rats.     

Establishment of an allergic rhinitis model in mice for the evaluation of nasal symptoms

The purpose of our study was to establish a new model of allergic rhinitis in mice, eliciting symptoms such as sneezing, infiltration of eosinophils into the nasal mucosa, and antigen-specific IgE production. One of the major human T-cell epitopes in Cry j 1, an allergen of Japanese cedar pollen, is also a major murine T-cell epitope in B10.S mice. Thus we tried to establish an allergic rhinitis model in B10.S mice with Cry j 1 as the antigen. We sensitized B10.S mice subcutaneously with Cry j 1/alum three times at intervals of 1 week. Five weeks after the final sensitization, we challenged the mice by instilling Cry j 1 intranasally from the day after intranasal histamine pretreatment. Soon after, we counted the number of sneezes. We then evaluated the infiltration of eosinophils into the nasal tissues and also measured the serum levels of antigen-specific IgE antibody. In addition, we confirmed the effects of ketotifen fumarate and dexamethasone hydrochloride on these animals. In Cry j 1-sensitized B10.S mice, sneezes, eosinophil peroxidase (EPO) activity in nasal tissues, and Cry j 1-specific IgE clearly increased after intranasal histamine pretreatment and 5 days of continuous intranasal Cry j 1 challenge. Both ketotifen and dexamethasone inhibited the increase in sneezing, and dexamethasone also inhibited EPO activity and Cry j 1-specific IgE. Thus we succeeded in establishing a new model of allergic rhinitis in Cry j 1-sensitized B10.S mice, which exhibited sneezing, eosinophil infiltration into the nasal mucosa, and Cry j 1-specific IgE production.     

Critical role of protein kinase C betaII in activation of mast cells by monomeric IgE

Accumulating evidence suggests that IgE-mediated activation of mast cells occurs even in the absence of antigen, which is referred to as "monomeric IgE" responses. Although monomeric IgE was found to induce a wide variety of responses, such as up-regulation of the FcepsilonRI, survival, cytokine production, histamine synthesis, and adhesion to fibronectin, it remains to be clarified how mast cells are activated in the absence of antigen. It has been controversial whether monomeric IgE responses are mediated by a similar signaling mechanism to antigen stimulation, although recent studies suggest that IgE can induce the FcepsilonRI aggregation even in the absence of antigen. In this study, we focused on the role of conventional protein kinase C (cPKC), since this response is suppressed by a specific inhibitor for cPKC. Monomeric IgE-induced Ca(2+) influx was not observed in a mouse mastocytoma cell line, which lacks the expression of PKCbetaII, although Ca(2+) influx induced by cross-linking of the FcepsilonRI was intact. Transfection of PKCbetaII cDNA was found to restore the Ca(2+) influx induced by monomeric IgE in this cell line. Furthermore, the dominant negative form of PKCbetaII (PKCbetaII/T500V) significantly suppressed the Ca(2+) influx, histamine synthesis, and interleukin-6 production in another mouse mast cell line, which is highly sensitive to monomeric IgE. Expression of PKCbetaII/T500V was found not to affect the antigen-induced responses. These results suggest that PKCbetaII plays a critical role in monomeric IgE responses, but not in antigen responses.     

Carrier-specific unresponsiveness in reaginic antibody formation. II. Suppression of hapten and carrier responsiveness in presensitized rats.

By inducing carrier-specific tolerance to sheep γ-globulin (SGG) in rats challenged with TNP-SGG in alum, it has been possible to study the effect of helper T-cell Unresponsiveness on IgE anti-TNP antibody formation. Rats primed to either the carrier (SGG) or the hapten (TNP as TNP-KLH) were treated with a single high dose (10 mg) of soluble SGG resulting in a suppression of both IgE anti-TNP and anti-SGG antibody which was maintained following a normally immunogenic secondary challenge with TNP-SGG in alum. This suppression was relatively long lasting, with no detectable IgE responsiveness to hapten or carrier observed for up to 8 weeks after tolerance induction. Suppressed animals were able to respond to the hapten when challenged with TNP-KLH, indicating that the induced effect did not directly involve the IgE antibody producing cells, but rather the carrier-specific helper cells. These results parallel our previous findings for IgM and IgG responses in a similar system. Such relatively long lasting and easily induced suppression in IgE antibody formation to specific protein antigens in primed animals may eventually provide a clinically useful means of allergic desensitization to large protein allergens.     

Control of IgE and IgG1 antibody production in mice.

The production of IgE and IgG1 was studied in untreated, thymectomized. splenectomized, anti-thymocyte serum-treated, or sublethally X-irradiated mice. Dinitrophenyl Ascaris and ovalbumin were used as antigens, and aluminum hydroxide was used as adjuvant. A suppression of IgE production was observed in adult thymectomized mice, although the kinetic pattern of the antibody response was the same as in control animals. IgG1 antibody production was not affected by thymectomy. Splenectomy did not change either IgE or IgG1 production. A single dose of rabbit anti-thymocyte serum (ATS) given 8 days after immunization inhibited IgE antibody production. The effect of ATS was dose dependent and also varied with the amount of antigen used, the immune response to high doses being more susceptible to the effect of ATS. No alteration in IgG1 production was caused by ATS even when IgE antibody formation was completely inhibited. When preceding immunization, sublethal irradiation enhanced IgE antibody formation and partially suppressed IgG1 production; applied after immunization, irradiation caused an enhancement of IgE production which was inversely proportional to the interval elapsed between the two procedures. On the other hand, the IgG1 antibody production was fairly resistant to the same treatment. The results suggest a clearcut separation between the mechanisms regulating IgE and IgG1 production in mice.     

IgE peptide-specific CTL inhibit IgE production: a transient IgE suppression model in wild-type and HLA-A2.1 transgenic mice

Effect of IgE peptide-specific CTL on IgE antibody production was studied in mouse models. CTL elicited in B6.A2Kb tg mice against a human IgE peptide nonamer, pWV, lysed human IgE-secreting U266 myeloma cells and inhibit IgE production by these cells. U266 transfected with mouse A2Kb transgene (U266-A2Kb) were optimally lysed by these CTL, because the α3 domain of A2Kb interacts well with the CD8 co-receptors. The CTL generated were more effective in inhibiting IgE production by U266-A2Kb cells than lysing these cells. IgE production by and progression of U266 myeloma were suppressed in B6.A2Kb tg mice rendered tolerant to these cells and vaccinated with pWV along with CpG. We also studied the CTL response elicited in wild-type mice by a mouse nonameric IgE peptide, PI-1, along with CpG. This treatment caused a transient suppression of the IgE response in mice previously sensitized to an antigen. In mice treated with this regimen repeatedly, the IgE response was fully recovered 20 days after each treatment. Notably, while IgE peptide/CpG-treated mice remained unresponsive to antigen challenge in vivo, antigen-specific IgE production can be elicited by antigen in cultured splenocytes from these mice. Moreover, IgE peptide/CpG also inhibited an on-going IgE response, including IgE production by bone marrow cells. Taken together, these observations indicate that a CTL-based IgE peptide vaccine targeting IgE-secreting B/plasma cells may be safely employed as a therapeutic approach for suppressing IgE production.     

IgE isotype switch and IgE production are enhanced in IL-21-deficient but not IFN-gamma-deficient mice in a Th2-biased response

IgE plays a critical role in the pathogenesis of allergy and asthma. Therefore, suppression of IgE production would provide therapeutic benefits to patients suffering from these diseases. We have reported that the production of IgE is regulated differently in the spleen vs. the draining lymph nodes (LN). IgE isotype switch and IgE producing B cell expansion occur in the draining LN after antigen (Ag) immunization, but do not happen in the spleen. In addition, a population of pre-existing IgE+ cells is observed in the spleen of normal or sham immunized mice, but is not present in the draining LN. To further understand the regulation of IgE production in different lymphoid organs, and the potential inhibitory factors of IgE isotype switch in the spleen, the involvement of IL-21 and IFN-gamma in regulating IgE production was investigated by using the IL-21 and the IFN-gamma deficient mice. We found that in the absence of IL-21 IgE isotype switch and IgE+ cell clonal expansion were dramatically enhanced in the spleen and IgE isotype switch was partially increased in the draining LN. In addition, IgE production of the pre-existing CD19-CD5+B220(low) IgE+ cells in the spleen was also increased in the absence of IL-21 under physiological conditions. In contrast, using the IFN-gamma deficient mice, we did not observe a negative impact of IFN-gamma on either IgE isotype switch or IgE production. Our data suggest that IL-21 appears to be a critical cytokine to keep low IgE levels under physiological and pathological conditions.     

Elicitation of antigen-induced primary and secondary murine IgE antibody responses in vitro

Described herein are methods for eliciting and quantitating primary and secondary murine IgE antibody responses in vitro, and the important role of antigen concentration in determining the level of IgE produced during an immune response. The methods for quantitating IgE antibody levels in culture supernatant fluids and in serum by ELISA are presented in detail. The specificity of such methods was confirmed in that (1) no other isotype of antibody registered in the IgE-ELISA, and (2) parallel determinations of IgE antibody concentrations could be obtained by independent analysis using Fc epsilon RI-dependent basophil degranulation. We examined various parameters of cell donor immunization and lymphoid cell culture which allow for optimal in vitro primary and secondary IgE responses. High relative antigen doses result in diminished IgE antibody responses in experimental animals, a finding confirmed herein. High antigen concentrations in vitro also result in relative suppression of IgE antibody synthesis. This was also true for in vitro production of IgG1 and IgA antibodies. Conversely, IgM and IgG2a responses were elicited at both low and high antigen concentrations; IgG2b and IgG3 were not produced under the conditions of priming and culture used herein. Finally, production of IgE in vitro depended on the presence of carrier-primed CD4+ T cells and hapten-specific B cells. Generation of maximal IgE antibody secretion, and hence elicitation of an allergic reaction, is thus dependent on the amount of antigen acting as stimulant for the immune response.     

The action of interleukin-4 on antigen-specific IgG1 and IgE production by interaction of in vivo primed B cells and carrier-specific cloned Th2 cells

The production of anti-trinitrophenyl (TNP) antibodies of different isotypes from in vivo primed B cells was studied using the plaque-forming cell method. It was shown that these B cells secrete anti-trinitrophenyl antibodies of different isotypes only in the presence of Th2 cells specific for keyhole limpet hemocyanin (KLH) and the hapten-carrier conjugate TNP-KLH. Lipopolysaccharide-stimulated primed B cells without cells from the Th2 clone did not produce anti-TNP-specific IgG1 or IgE antibodies even in the presence of the hapten-carrier antigen TNP-KLH. Supernatants from these Th2 clones cultured with antigen-presenting cells and the complete antigen were unable to activate primed B cells for antibody secretion. Cognate interaction between primed B cells and carrier-specific Th2 cells is a prerequisite for hapten-specific IgG1 or IgE production. Anti-IL-4 antibody inhibited secretion of anti-hapten IgE antibody. Therefore, for production of anti-hapten antibody of the IgE isotype IL-4 is also necessary.     

Stimulation by adjuvants of cell-mediated immune response in Plasmodium berghei infected mice

Immunological adjuvants (alum, liposomes and saponin) were utilized to stimulate cell-mediated immune response in Plasmodium berghei infected Balb/c mice. It was shown that malaria antigen mixed with adjuvant induced appreciably delayed type hypersensitivity and production of migration inhibition factor compared to antigen alone.     

人参多糖对新甲型H1N1流感病毒灭活疫苗的免疫增强作用

在流感灭活疫苗中添加佐剂可以提高疫苗的免疫原性,节约抗原用量。一些天然中草药多糖具有潜在的佐剂效应。本文探讨了人参多糖（ginseng polysaccharide,GPS）在新甲型H1N1流感病毒裂解型灭活疫苗中的佐剂效应。将不同剂量GPS与新甲型H1N1流感病毒灭活疫苗混合,共同免疫小鼠一次,通过检测免疫后在小鼠体内诱导产生的疫苗特异性IgM、IgG、IgG1和IgG2a抗体情况来评价GPS作为流感病毒灭活疫苗佐剂的免疫增强效果,并与不添加佐剂的疫苗和加有铝佐剂的疫苗的免疫效果作比较。结果显示,GPS与铝佐剂一样能显著提高和维持疫苗特异性IgG抗体滴度,同时提高IgM抗体水平,其中800μgGPS的佐剂效果最好。因此我们认为GPS可以作为流感病毒灭活疫苗的一种候选佐剂。     

Effects of dietary lactosucrose (4G-beta-D-galactosylsucrose) on the IgE response in mice

In this study, we examined the effects of dietary lactosucrose (LS, a non-digestible oligosaccharide) on the IgE response in mice immunized with ovalbumin (OVA)/alum. In addition to IgG1 and IgG2a responses, the anti-OVA IgE response in mice fed LS diets was dose-dependently suppressed, as compared with the control mice, while the serum total IgG levels were comparable. Moreover, dietary LS feeding inhibited antigen-specific IgE and IgG1 productions even after a second immunization. Regarding with cytokine production, when stimulated in vitro with OVA, splenocytes obtained from LS-fed mice produced a similar level of IFN-gamma, and lower levels of IL-4 and IL-5, as compared with the control mice. But IL-10 production by OVA-stimulated splenocytes was augmented in LS-fed mice, suggesting that IL-10 producing cells are responsible for the immunoregulatory effect of LS. Our findings indicate the further possibility that dietary LS supplementation can be used to prevent IgE-mediated allergic diseases.     

Aluminium hydroxide adjuvant initiates strong antigen-specific Th2 responses in the absence of IL-4- or IL-13-mediated signaling

Previous studies demonstrate that aluminium hydroxide adjuvant (alum) produces increased Th1 responses in IL-4-deficient mice compared with wild-type animals, although the continued production of IL-5 by spleen cells from these mice also indicates that Th2 responses are induced. In the present study, we demonstrate that alum can induce Th2-associated IL-4 and IL-5 production in the absence of IL-4 signaling in mice deficient in either IL-4Ralpha or Stat6. The Th2 responses observed could not be due to IL-13 as IL-13 responses are also impaired in IL-4Ralpha- and Stat6-deficient mice. We also detected higher levels of IL-4 in IL-4Ralpha gene-deficient, though not Stat6-deficient, mice compared with their wild-type counterparts. The increased levels of IL-4 could be explained by the IL-4R being unavailable to neutralize this cytokine in IL-4Ralpha-deficient mice. While levels of IL-5 production in IL-4Ralpha- or Stat6-deficient mice were similar to IL-4-deficient and wild-type mice, other type 2-associated responses, which are largely or wholly IL-4 dependent, such as the production of IgG1 or IgE Abs, were either reduced or absent. We conclude that alum adjuvants can induce IL-4 production and Th2 responses independently of IL-4 or IL-13, negating the requirement for an early source of IL-4 in the Th2 response induced by this adjuvant.     

Different T helper cell subsets elicited in mice utilizing two different adjuvant vehicles: the role of endogenous interleukin 1 in proliferative responses

Mice were immunized with either poly(Glu, Arg, Ala) or poly(Glu, Lys, Phe) contained in two different adjuvant preparations, alum (A1K(SO4)2) or complete Freund's adjuvant (CFA), and in vitro antigen-driven proliferative responses of lymph node cells were assayed 4-16 days later. After immunization with antigens on alum, endogenous interleukin 1 (IL-1) as well as interleukin 4 (IL-4) production was required for the proliferation of the elicited T cells as inclusion of either polyclonal goat anti-mouse IL-1 alpha or monoclonal anti-mouse IL-4 (11B11) in the cultures inhibited proliferative responses to antigen. In contrast, proliferative responses of cells elicited by antigen in CFA were not inhibited by either anti-IL-1 or anti-IL-4. Monoclonal antimouse CD4 (GK 1.5) inhibited proliferative responses regardless of which adjuvant was used to elicit antigen-reactive cells. These data indicated that phenotypically different subpopulations of CD4+ cells were elicited by the same antigen administered in different adjuvant preparations, Th2-like cells after immunization with polymers on alum and Th1-like cells after immunization with antigens in CFA. An examination of the isotypes of polymer-specific antibodies present in the sera of immunized mice also supported this conclusion.