Short-chain fatty acids inhibit the activation of T lymphocytes and myeloid cells and induce innate immune tolerance.

The intestinal microbiota contributes to gut immune homeostasis, where short-chain fatty acids (SCFAs) function as the major mediators. We aimed to elucidate the immunomodulatory effects of acetate, propionate, and butyrate. With that in mind, we sought to characterise the expression of SCFA receptors and transporters as well as SCFAs' impact on the activation of different immune cells. Whereas all three SCFAs decreased tumour necrosis factor (TNF)-α production in activated T cells, only butyrate and propionate inhibited interferon (IFN)-γ, interleukin (IL)-17, IL-13, and IL-10 production. Butyrate and propionate inhibited the expression of the chemokine receptors CCR9 and CCR10 in activated T- and B-cells, respectively. Similarly, butyrate and propionate were effective inhibitors of IL-1ß, IL-6, TNF-α, and IL-10 production in myeloid cells upon lipopolysaccharide and R848 stimulation. Acetate was less efficient at inhibiting cytokine production except for IFN-α. Moreover, SCFAs inhibited the production of IL-6 and TNF-α in monocytes, myeloid dendritic cells (mDC), and plasmacytoid dendritic cells (pDC), whereas acetate effects were relatively more prominent in pDCs. In monocytes and mDCs, acetate was a less efficient inhibitor, but it was equally effective in inhibiting pDCs activation. We also studied the ability of SCFAs to induce trained immunity or tolerance. Butyrate and propionate - but not acetate - prevented Toll-like receptor-mediated activation in SCFA-trained cells, as demonstrated by a reduced production of IL-6 and TNF-α. Our findings indicate that butyrate and propionate are equally efficient in inhibiting the adaptive and innate immune response and did not induce trained immunity. The findings may be explained by differential SCFA receptor and transporter expression profiles of the immune cells.

Interaction of mouse splenocytes and macrophages with bacterial strains in vitro: the effect of age in the immune response.

Probiotics influence the immune system, both at the local and systemic level. Recent findings suggest the relation between microbiota and the immune system alters with age. Our objective was to address direct effects of six bacterial strains on immune cells from young and aged mice: Lactobacillus plantarum WCFS1, Lactobacillus casei BL23, Lactococcus lactis MG1363, Bifidobacterium breve ATCC15700, Bifidobacterium infantis ATCC15697, and Akkermansia muciniphila ATCC BAA-835. We used splenocytes and naïve or interferon-γ-stimulated bone marrow-derived macrophages (BMDM) as responder populations. All tested bacterial strains induced phenotypic and cytokine responses in splenocytes and BMDM. Based on magnitude of the cellular inflammatory response and cytokine profiles, two subgroups of bacteria were identified, i.e. L. plantarum and L. casei versus B. breve, B. infantis, and A. muciniphila. The latter group of bacteria induced high levels of cytokines produced under inflammatory conditions, including tumour necrosis factor (TNF), interleukin (IL)-6 and IL-10. Responses to L. lactis showed features of both subgroups. In addition, we compared responses by splenocytes and BMDM derived from young mice to those of aged mice, and found that splenocytes and BMDM derived from aged mice had an increased IL-10 production and dysregulated IL-6 and TNF production compared to young immune cells. Overall, our study shows differential inflammatory responses to distinct bacterial strains, and profound age-dependent effects. These findings, moreover, support the view that immune environment importantly influences bacterial immune effects.

A Gly15Arg mutation in the interleukin-10 gene reduces secretion of interleukin-10 in Crohn disease.

BACKGROUND: Genetic susceptibility, probably involving cytokines and their receptors, plays an important role in inflammatory bowel disease (IBD). In this study we examine the potential role of the interleukin-10 (IL-10) gene as a susceptibility gene in IBD. METHODS: We studied 17 sib-pairs with either Crohn disease (CD) or ulcerative colitis. After microsatellite analysis for allele-sharing, the IL-10 gene of sib-pairs who shared alleles was screened for nucleotide alterations in and around exons and the promoter region. The IL-10 promoter polymorphism at position -1082 was also determined. Function was evaluated by measuring IL-10 secretion by peripheral blood mononuclear cells stimulated with lipopolysaccharide or phorbol ester. The activity of recombinant immature wild-type and mutated IL-10 was tested in a proliferation assay with a human monocytic leukaemia cell line (HL60 cells). RESULTS: DNA sequencing revealed a G --> A point mutation in exon 1 at base position 43 in one sib-pair, both affected with CD. It was also found in 2 of their healthy siblings, but not in 75 unrelated healthy controls. This mutation results in a glycine to arginine substitution at amino acid position 15 of the leader sequence (Gly15Arg). The in vitro IL-10 secretion by mononuclear cells of the IL-10 Gly15Arg carriers was about 50% of healthy controls, matched for the -1082 polymorphism in the IL-10 promoter region. Incubation of HL60 cells with recombinant mutated IL-10 showed a markedly reduced cell proliferation compared to wild-type IL-10. CONCLUSION: A Gly15Arg mutation in the leader sequence of IL-10 was found in a multiple CD-affected family. This altered leader sequence decreases IL-10 secretion, thereby reducing the anti-inflammatory effect.

1alpha,25-Dihydroxyvitamin d3 has a direct effect on naive CD4(+) T cells to enhance the development of Th2 cells.

1alpha,25-Dihydroxyvitamin D3 (vitD3) is an immunoregulatory hormone with beneficial effects on Th1 mediated autoimmune diseases. Although the inhibitory effects of vitD3 on macrophages and dendritic cells are well documented, any direct effects of vitD3 on Th cell development are not clearly defined. Using CD4(+)Mel14(+) T cells derived from mice on a BALB/c and a C57BL/6 genetic background we examined the effect of vitD3 on Th cell development. We demonstrated that vitD3 affects Th cell polarization by inhibiting Th1 (IFN-gamma production) and augmenting Th2 cell development (IL-4, IL-5, and IL-10 production). These effects were observed in cultures driven with splenic APC and Ag, as well as with anti-CD3 and anti-CD28 alone, indicating that CD4(+) cells can also be direct targets for vitD3. The enhanced Th2 development by vitD3 was found in both BALB/c and C57BL/6 mice. An increased expression of the Th2-specific transcription factors GATA-3 and c-maf correlated with the increased production of Th2 cytokines after vitD3 treatment. The vitD3-induced effects were largely mediated via IL-4, because neutralization of IL-4 almost completely abrogated the augmented Th2 cell development after vitD3 treatment. These findings suggest that vitD3 acts directly on Th cells and can, in the absence of APC, enhance the development of a Th2 phenotype and augment the expression of the transcription factors c-maf and GATA-3. Our findings suggest that the beneficial effects of vitD3 in autoimmune diseases and transplantation operate through prevention of strong Th1 responses via the action on the APC, while simultaneously directly acting on the T cell to enhance Th2 cell development.

Differential ultraviolet-B-induced immunomodulation in XPA, XPC, and CSB DNA repair-deficient mice.

Ultraviolet B irradiation has serious consequences for cellular immunity and can suppress the rejection of skin tumors and the resistance to infectious diseases. DNA damage plays a crucial role in these immunomodulatory effects of ultraviolet B, as impaired repair of ultraviolet-B-induced DNA damage has been shown to cause suppression of cellular immunity. Ultraviolet-B-induced DNA damage is repaired by the nucleotide excision repair mechanism very efficiently. Nucleotide excision repair comprises two subpathways: transcription-coupled and global genome repair. In this study the immunologic consequences of specific nucleotide excision repair defects in three mouse models, XPA, XPC, and CSB mutant mice, were investigated. XPA mice carry a total nucleotide excision repair defect, whereas XPC and CSB mice only lack global genome and transcription-coupled nucleotide excision repair, respectively. Our data demonstrate that cellular immune parameters in XPA, XPC, and CSB mice are normal compared with their wild-type (control) littermates. This may indicate that the reported altered cellular responses in xeroderma pigmentosum patients are not constitutive but could be due to external factors, such as ultraviolet B. Upon exposure to ultraviolet B, only XPA mice are very sensitive to ultraviolet-B-induced inhibition of Th1-mediated contact hypersensitivity responses and interferon-gamma production in skin draining lymph nodes. Lipopolysaccharide-stimulated tumor necrosis factor alpha and interleukin-10 production are significantly augmented in both XPA and CSB mice after ultraviolet B exposure. Lymph node cell numbers were increased very significantly in XPA, mildly increased in CSB, and not in XPC mice. In general XPC mice do not exhibit any indication of enhanced ultraviolet B susceptibility with regard to the immune parameters analyzed. These data suggest that both global genome repair and transcription-coupled repair are needed to prevent immunomodulation by ultraviolet B, whereas transcription-coupled repair is the major DNA repair subpathway of nucleotide excision repair that prevents the acute ultraviolet-B-induced effects such as erythema.

Selective development of a strong Th2 cytokine profile in high-risk children who develop atopy: risk factors and regulatory role of IFN-gamma, IL-4 and IL-10.

BACKGROUND: The immunological processes in early life and their relation to allergic sensitization leading to a Th2 cytokine profile are still not well understood. OBJECTIVE: To analyse the environmental and genetic risk factors and immunological responses at birth in relation to the development of atopic disease at 12 months of age in a longitudinal study of high-risk children. METHODS: High-risk children were followed from birth till 12 months of age. Mononuclear cells obtained at birth and 6 and 12 months thereafter were analysed for their proliferative and cytokine responses after polyclonal and allergen-specific stimulation. RESULTS: At 12 months of age 25% children had developed an atopic disease. Two atopic parents, parental smoking and atopic dermatitis of at least one of the parents were significant risk factors. In cord blood of newborns who developed atopy, an increased percentage of CD4+CD45RO+ cells and an increased polyclonal-stimulated proliferation were observed. Furthermore, an impaired allergen-induced, but not polyclonal-stimulated IFN-gamma production was found, suggesting a regulatory defect. At 6 and 12 months of age, a strong Th2 profile (characterized by increased levels of IL-4, IL-5, and IL-13) after both polyclonal and, to a lesser extent, allergen-specific stimulation was found in the children developing atopy. Allergen-induced IL-10 production at 12 months of age was only observed in the non-atopic children. CONCLUSION: Our data indicate that the first 6 months of life represent a critical time window for the initiation of immunological changes resulting in the development of atopy. The selective development of a Th2 cytokine profile in high-risk children who develop atopy is due to increased production of Th2 cytokines, possibly caused by impaired allergen-induced IFN-gamma production in the neonatal period. Furthermore, the decreased allergen-induced IL-10 levels observed in the atopic children at 12 months of age may result in a lack of down-regulation of the inflammatory process.

Regulation of helper T cell responses to staphylococcal superantigens.

Staphylococcal superantigens (sAgs) including toxic shock syndrome toxin-1 (TSST-1) and related enterotoxins are exoproteins with unique immunobiological properties. They bind to major histocompatibility complex (MHC) class II molecules of antigen-presenting cells outside the peptide groove, and induce massive proliferation of T cells bearing specific V beta determinants. This tri-molecular interaction leads to uncontrolled release of various proinflammatory cytokines especially interferon-gamma (IFN-gamma) and tumor necrosis factor-a (TNF-alpha), the key cytokines causing sAg-mediated shock. The effector T cells involved in this hyper-immune response are predominantly of the T helper-1 (Th1) phenotype. There is also some evidence that polarization to a Th2 response with the production of classical anti-inflammatory cytokines (such as interleukins IL-4 and IL-6) also occurs. Moreover, the emergence of a novel regulatory T cell (Tr1) subset, producing mainly IL-10 but little or no IL-2 and IL-4, has recently been described following repeated sAg stimulation. In this review, the current knowledge regarding the regulation of T helper cell subsets in response to staphylococcal sAgs is critically evaluated, and the role of various cytokines which directly influence T cell differentiation and polarization is summarized. Particular emphasis is directed towards pro-inflammatory as well as anti-inflammatory and regulatory effector functions during toxic shock. Based on this review, we propose that a delayed production of IL-10 by Tr1 cells may be the most prominent driving force in the down-regulation of the Th1 hyper-immune response, and the critical determinant for the eventual recovery of the host.

UVB irradiation modulates systemic immune responses by affecting cytokine production of antigen-presenting cells.

The immunosuppressive effects of UVB irradiation have been well documented. The production of cytokines by keratinocytes is considered to play a major role in the induction of local as well as systemic immunosuppression. It is thought that partly due to the interaction of locally produced cytokines with antigen-presenting cells (APC) systemic effects, like antigen-specific tolerance, can be induced. In this study we examined the effect of UVB irradiation on cytokine profiles of peripheral APC as well as the functional consequences. Our results indicate that UVB irradiation impairs T(h)1-mediated immune responses in vivo by suppression of the systemic IL-12p70 production. Splenic APC from UVB-exposed mice showed an enhanced production of prostaglandin E(2), IL-1, IL-6 and tumor necrosis factor-alpha after in vitro stimulation. Also, spleen cells from UVB irradiated IL-4(-/-) mice showed increased IL-6 levels. These APC were less efficient in inducing IFN-gamma production by CD4(+) T cells and suppressed IgM production by B cells. We conclude that the altered cytokine profile of peripheral APC can be responsible for the systemic effects of UVB irradiation on the T(h)1/T(h)2 balance as well as on B cell responses.

Critical illness polyneuropathy and myopathy (CIPNM): evidence for local immune activation by cytokine-expression in the muscle tissue.

In a longitudinal prospective study a muscle biopsy was taken from 30/32 (33%) of the 98 patients who developed critical illness polyneuropathy and myopathy (CIPNM). Neuropathic changes were found in 37%, myopathic in 40%, and a combination in 23% of the biopsies. The immunohistopathology showed macrophages and Th-cells in 40% and 60% of the muscle biopsies respectively. Small mainly perivascular infiltrates contained macrophages and Th-cells. ICAM-1, VCAM and MAC were found on the vascular endothelium in 58%, 53% and 79% respectively. In all biopsies there was an upregulation of both HLA-I and HLA-DR. Proinflammatory cytokines and TNFalphaR75 were also produced locally (IL-1beta in 71%, IFN-gamma in 40%, IL-12 in 73%, TNFalphaR75 in 90%). The anti-inflammatory cytokine IL-10 was simultaneously expressed in 96% of the biopsies. HLA-DR, TNFalphaR75 and IL-10 differed significantly when compared with control muscle biopsies. Our data provide evidence that small numbers of activated leukocytes producing both pro- and anti-inflammatory cytokines infiltrate skeletal muscle of CIPNM patients. We propose that the local balance of leukocyte activities is of importance in the pathophysiology of muscle weakness in CIPNM.

Interleukin 4 receptors on human bronchial epithelial cells. An in vivo and in vitro analysis of expression and function.

Asthma is considered a Th2-like disease, characterized by locally increased levels of interleukin (IL) 4. The bronchial epithelium plays an important role in the initiation and perpetuation of inflammatory reactions within the airways. However, little is known about the presence of IL-4 receptors on human bronchial epithelial cells, or the effects of IL-4 on these cells. In this report, definitive evidence of IL-4 receptor expression on human bronchial epithelial cells using several methods is presented. IL-4 receptor expression on human bronchial epithelial cells in vivo was demonstrated using in situ hybridization and immunohistochemistry. No difference in IL-4 receptor protein expression was observed between bronchial biopsies of healthy subjects compared to allergic asthmatics. Cultured human bronchial epithelial cells also expressed IL-4 receptor mRNA and protein (as determined by RT-PCR analysis and flow cytometry, respectively). IL-4 receptor protein expression by bronchial epithelial cells could be increased by stimulation with PMA+calcium ionophore, whereas IL-1beta and IL-6 decreased IL-4 receptor expression. A cyclic AMP analogue and IL-4 had no effect. Finally, it is shown that the IL-4 receptor is functionally active as IL-4 stimulates the release of IL-8, monocyte chemoattractant protein 1, and particularly IL-1 receptor antagonist by human bronchial epithelial cells. It is concluded that human bronchial epithelial cells express IL-4 receptors both in vivo and in vitro. Stimulation of human bronchial epithelial cells by IL-4 may result in the release of both pro- and anti-inflammatory mediators known to be upregulated in asthmatic airways.

A mouse model to study immunity against pseudorabies virus infection: significance of CD4+ and CD8+ cells in protective immunity.

In this study we firstly established a vaccination/challenge model to study pseudorabies virus infection in mice. The mouse model was used to investigate the significance of CD4+ and CD8+ cells and of IFN gamma production in protective immunity. Functional depletion of CD4+ and CD8+ and IFN gamma was obtained in vivo by intraperitoneal injection of alginate-encapsulated anti-CD4, -CD8 or -IFN gamma producing hybridoma's before and at the moment of vaccination. The observed protective immunity was correlated with underlying immunologic responses such as PRV-specific DTH reactivity, lymphoproliferation and cytotoxicity. The significance of CD4+ and CD8+ cells and of IFN gamma production was also investigated for these immunological responses by the same in vivo depletion technique. The results demonstrated that protective vaccination of mice, that could be induced by immunization with 10(7) plaque forming units of the avirulent PRV mutant NIA3 TK-, was characterized by a typical anti-viral Th1 type immune response. A clear PRV-specific, CD4-dependent DTH reactivity and a classical CD8-dependent, MHC-restricted cytotoxicity was induced after protective immunization and the humoral immune response had a bias towards PRV-specific IgG2a formation. In vivo treatment with anti-CD8 and anti-IFN gamma demonstrated that the cytotoxic response and humoral IgG2a response, respectively, were strongly reduced, whereas protection against lethal challenge was unaffected. On the other hand anti-CD4 treatment reduced the induced protection so that 30% of the mice died after lethal challenge. The results of our study demonstrated that CD4+, DTH like effector cells are a crucial effector mechanism for protective immunity against PRV.

Levels of soluble intercellular adhesion molecule-1, soluble E-selectin, tumor necrosis factor-alpha, and soluble tumor necrosis factor receptor p55 and p75 in atopic children.

During inflammation, membrane expression of adhesion molecules and tumor necrosis factor (TNF)-receptors (TNF-R) are increased, and soluble forms of these molecules are released. This study analyzed plasma levels of sICAM-1 and sE-selectin as well as TNF-alpha, sTNF-R55, and sTNF-R75 in nonallergic (NAA) and allergic asthma patients (AA), atopic dermatitis patients (AD), and healthy children (HC) by ELISA. Plasma levels of sICAM-1, sE-selectin, and sTNF-R, but not TNF-alpha, were detectable, but were not significantly different between the patient groups and healthy children. In the AA group, a significant correlation (rs = 0.78, P = 0.008) was found between sICAM-1 and sE-selectin levels. Furthermore, a significant correlation was found between sTNF-R55 and sTNF-R75 levels in the AA group (rs = 0.70, P = 0.025) and in the AD group (rs = 0.69, P = 0.027). In AD patients, a significant correlation was observed between sE-selectin and the disease severity, as measured by the SCORAD index (rs = 0.73, P = 0.038). Our data demonstrate that plasma levels of sICAM-1, sE-selectin, TNF-alpha, sTNF-R55, and sTNF-R75 were not different between atopic and nonatopic children during a stable phase of the disease. In AD patients, levels of sE-selectin seemed to be related to clinical severity of the disease.

Monoclonal gammopathies in aging mu, kappa-transgenic mice: involvement of the B-1 cell lineage.

Monoclonal gammopathies (MG) are monoclonal proliferative disorders of B cells at the differentiation stage of Ig production. They can be detected in the serum, either as transient or as persistent homogenous immunoglobulin (H-Ig) components. The exact phenotype, localization, and cell lineage origin of the precursor cells of MG are unknown, but may be crucial for both the correct diagnosis and for timely efficient treatment of the malignant forms. We used for the first time transgenic (Tg) mice (Sp6; mu/kappa) to study the origin of MG. In the mu, kappa Tg mice a small proportion of B cells can still produce endogenous IgM. These cells are of B-1 cell origin. The MG in Tg mice showed a later onset and a lower frequency than those in littermate control mice, mainly due to a four times lower frequency of benign monoclonal gammopathy. The 10% of B-1 cells that were able to produce endogenous Ig led to the development of MG in a frequency that was half the number of MG found in normal littermates. None of the MG in Tg mice produced an Ig of the Tg origin and therefore it can be concluded that they originated from B-1 cells.

The role of cytokines in ultraviolet-B induced immunosuppression.

Cytokines play an important role in the mechanisms resulting in ultraviolet B-induced immunosuppression. They play a crucial role in the induction of local as well as systemic immunomodulatory events. Ultraviolet B irradiation directly induces the release of cytokines in the epidermis, resulting in complex and diverse interactions on epidermal cells, which results in migration out of the skin and infiltration into the skin of different cell types. Partly because of the interaction of locally produced cytokines with antigen-presenting cells, systemic effects, such as antigen-specific tolerance, can be observed. This review describes the different mechanisms whereby ultraviolet B exposure affects immune functions with special emphasis on the role of T-cell subsets and cytokines.

Increased adjuvant efficacy in stimulation of antibody responses after macrophage elimination in vivo.

To investigate whether macrophages influence the efficacy of adjuvants, we locally eliminated lymph node macrophages with dichloromethylene diphosphonate containing-liposomes before primary immunization. After macrophage elimination, animals were immunized with a soluble antigen (TNP-KLH; 2,4,6-trinitrophenyl-keyhole limpet haemocyanin) either in phosphate-buffered saline (PBS), in complete Freund's adjuvant (CFA), or in Specol is a water-in-oil emulsion, considered to be less aggressive than CFA, but equally effective. A secondary immunization followed with TNP-KLH. In vivo macrophage elimination before Specol/TNP-KLH immunization resulted in increased adjuvant efficacy as measured by (primary) antibody responses. This suggests that the activity of Specol is not primarily mediated through macrophages but rather through other antigen-presenting cell types (e.g. dendritic cells, B cells, fibroblasts). The overall quality of produced antibodies, in terms of isotype distribution and affinity maturation, remained the same. After primary injection, CFA/TNP-KLH immunization resulted in significantly higher antibody responses in macrophage-depleted animals and antibody responses did not increase significantly after secondary immunization. However, a booster effect was found when macrophages were not eliminated before CFA/TNP-KLH immunization, suggesting that the presence of macrophages during the first weeks of the primary immune response is essential for the induction of an effective secondary response in CFA immunizations. In conclusion, macrophage depletion before immunization with a soluble T-cell-dependent antigen combined with Specol may enhance specific antibody responses without changing the isotype or affinity of the antibodies.

Intrathecal production of interleukin-12 and gamma interferon in patients with bacterial meningitis.

To assess the role of interleukin-12 (IL-12) and gamma interferon (IFN-gamma) in children with bacterial meningitis, bioactive IL-12 (p70) and the inactive subunit p40 and IFN-gamma were measured in serum and cerebrospinal fluid (CSF) from 35 children with bacterial meningitis and 10 control subjects. The production of IFN-gamma is induced by IL-12 with tumor necrosis factor alpha (TNF-alpha) as a costimulator and inhibited by IL-10. CSF concentrations of IL-12 p40 as well as those of IFN-gamma were markedly elevated, whereas IL-12 p70 was hardly detectable. Detectable CSF levels of IFN-gamma correlated positively with IL-12 p40 (r = 0.40, P = 0.02) and TNF-alpha (r = 0.46, P = 0.04) but not with IL-6, IL-8, or IL-10. In contrast to CSF levels of TNF-alpha, IL-12, and IL-10, those of IFN-gamma were significantly higher in patients with pneumococcal meningitis than in children with meningitis caused by Haemophilus influenzae and Neisseria meningitidis, presumably because of a high CSF TNF-alpha/IL-10 ratio in the former. We suggest that IL-12- and TNF-alpha-induced IFN-gamma production may contribute to the natural immunity against microorganisms in the CSF compartment during the acute phase of bacterial meningitis.

Development of airway hyperresponsiveness is dependent on interferon-gamma and independent of eosinophil infiltration.

In this study the role of interleukin (IL)4, IL5, interferon (IFN) gamma, and tumor necrosis factor (TNF) alpha in the development of airway hyperresponsiveness and inflammatory cell infiltration was investigated using a murine model for allergic asthma. Mice were sensitized with ovalbumin and subsequently challenged repeatedly with ovalbumin aerosols. During the challenge period, mice were treated with monoclonal antibodies directed against IL4, IL5, IFN gamma, or TNF alpha. Control antibody-treated mice showed airway hyperresponsiveness to methacholine and the presence of eosinophils in bronchoalveolar lavage (BAL). Treatment with antibodies to IFN gamma completely abolished development of airway hyperresponsiveness in ovalbumin-challenged animals. After treatment with antibodies to TNF alpha, airway hyperresponsiveness in the ovalbumin-challenged animals was partially but not significantly inhibited. Antibodies to IL4 or IL5 did not inhibit airway hyperresponsiveness. The presence of eosinophils in BAL of ovalbumin-challenged mice was completely inhibited after treatment with antibodies to IL5. Treatment with antibodies to IL4, IFN gamma, or TNF alpha had no effect on eosinophilia. Because IFN gamma and IL5 have either an effect on the induction of airway hyperresponsiveness or on the development of eosinophil infiltration, our results suggest that the two phenomena are differentially regulated.

Impaired T cell functions preceding lymphoproliferative disorders in mice neonatally tolerized to transplantation antigens.

In A/J (H-2a) (A) mice, the neonatal i.v. injection of (B10 x A)F1 spleen cells (SC) induces partial transplantation tolerance (TT) to C57BL/10ScSn (H-2b) (B10) skin allografts, chronic host-versus-graft disease (HVGD) and lethal lymphoproliferative disorders (LPD). They produce anti-T-cell autoantibodies (ATA), and the proliferative responses of their SC to the T cell mitogen Con A are decreased. We found that, similar to ATA, the hyporeactivity of T cells developed earlier (at 1-2 weeks of age) than splenomegaly. The proportions of both CD4+ and CD8+ T cells were not reduced in the spleens of tolerized mice without manifest LPD. The supernatants (SN) of Con A-stimulated tolerized SC contained no, or only small amounts of interleukin-2 (IL-2). Thus, in the tolerized mice, ATA and T cell deficiency preceded the development of LPD. ATA and the decreased amount of the T cell growth factor IL-2 might play a role in the defective T cell activation.

Soluble tumour necrosis factor receptor release after anti-CD3 monoclonal antibody treatment in mice is independent of tumour necrosis factor-alpha release.

The involvement of TNF-alpha in the release of soluble TNF receptors was assessed in mice, treated with anti-CD3 monoclonal antibodies. After treatment with three different anti-CD3 mAb, we simultaneously studied serum levels of TNF-alpha, soluble TNF receptor P55 and P75. All three anti-CD3 mAb triggered the release of both of the soluble TNF receptors, whereas only one of the anti-CD3 mAb triggered TNF-alpha release. These data demonstrate that in our model soluble TNF receptor release is independent of TNF-alpha release.

Role of IL-4 in persistent IgE formation.

Antigen-specific immunoglobulin E (IgE) responses against T-cell-dependent antigens, like allergens, can only be generated by cognate interaction between B-cells and T-helper (Th2) cells. This interaction is a prerequisite, donating the two signals that are essential for IgE production: CD40 ligation with its ligand gp39 and exposure to interleukin (IL)-4. Cytokine-mediated immunotherapy geared at intervention in allergic diseases, therefore aims at inhibiting the production or action of IL-4. In our view, based on two findings, this approach is simplistic. The first is that anti-IL-4 based approach (by complex formation between IL-4 and soluble IL-4 receptors or serum binding proteins) may actually increase the persistence of IL-4 and its effects instead of inhibiting them. Secondly, we have good evidence in mouse model systems that a period of exposure to IL-4 will result in an increased population of gamma 1,epsilon-double positive B-cells allowing an increased serum IgE level to persist for extensive periods of time. These B-cells turn out to be partially independent of IL-4 for their IgE formation. Moreover, these B-cells are partially independent of a cognate interaction with T-cells for their subsequent IgE synthesis. Therefore, we hypothesize that an approach geared solely at inhibiting IL-4 is not sufficient for decreasing persistent IgE levels in allergic patients.