Donata Vercelli

Regulation of immunoglobulin (Ig)E synthesis in the hyper-IgE syndrome.

The hyper-IgE (HIE) syndrome is characterized by high IgE serum levels, chronic dermatitis, and recurrent infections. The mechanisms responsible for hyperproduction of IgE in HIE patients are presently unknown. We investigated whether spontaneous in vitro IgE synthesis by PBMC from seven HIE patients was sensitive to signals (cell adhesion, T/B cell cognate interaction and lymphokines: IL-4, IL-6, and IFN-gamma) known to regulate IgE induction in normals. Our results show that, unlike IL-4 dependent IgE synthesis induced in normals, spontaneous IgE production by PBMC from HIE patients was not blocked by monoclonal antibodies to CD2, CD4, CD3, and MHC class II antigens. Furthermore, antibodies to IL-4 and IL-6 did not significantly suppress IgE production. IFN-gamma had no significant effects on spontaneous in vitro IgE synthesis. To test whether an imbalance in lymphokine production might underlie hyperproduction of IgE in HIE patients, mitogen-induced secretion of IL-4 and IFN-gamma by PBMC was assessed. No significant difference was detected between HIE patients and normal controls. Thus, ongoing IgE synthesis in the HIE syndrome is largely independent of cell-cell interactions and endogenous lymphokines, and is due to a terminally differentiated B cell population, no longer sensitive to regulatory signals.

Regulation of isotype switching

In the past year, our understanding of the mechanisms that regulate isotype switching has significantly progressed. Activation of germ line transcription by isotype-specific cytokines is emerging as a crucial mechanism for increasing the accessibility of a particular switch region, and targeting switch recombination.

Regulation of IgE synthesis in humans

Atopic disease is characterized by a high and sustained synthesis of IgE following stimulation by environmental antigens (allergens). IgE binds to high-affinity Fce receptors (FceR1) 2 on mast cells and basophils (1) and to low-affinity FceR (FceR2) on B cells, monocytes, eosinophils, and platelets (2). The reintroduction of the antigen, which crosslinks FceR-bound IgE, induces the release of a variety of mediators, which trigger a cascade of events resulting in the clinical symptoms of allergy. At present, the pharmacologic management of allergic diseases is aimed mainly at controlling the symptoms resulting from the release of mediators by effector cells. The development of a rational therapeutic approach to the treatment of allergic diseases will require a deeper understanding of the mechanisms responsible for the regulation of IgE responses. In 1986, we discussed the current views about the regulation of human IgE synthesis by T cells (3). Various laboratories had by then reported that,At the present time, in no other system are the signals required for isotype switching better understood than in the IgE system. IgE switching is therefore becoming a general model for directed isotype switching in mice and humans. The study of IgE regulation has proposed a paradigm of general importance for immunology in the nineties, namely, the requirement for at least two signals in order to trigger the final event, in this case DNA switch recombination to the IgE isotype. The first signal is delivered by a cytokine, IL-4 or IL-13, and is responsible for the choice of the isotype. The second signal is typically delivered upon engagement of CD40 on B cells by the CD40 ligand expressed on T cells, and results in switching and production of IgE. We shall herein discuss the two-signal model for IgE switching in detail, stressing in particular the cross-talk between signals, and the mechanisms responsible for IgE amplification.

Induction of interleukin-4-dependent IgE synthesis and interleukin-5-dependent eosinophil differentiation by supernatants of a human helper T-cell clone

The human alloreactive T-cell clone A1 produces interleukin (IL)-4, IL-5, and granulocyte macrophage-colony stimulating factor (GM-CSF), but not IL-2 or interferon-γ (IFN-γ), as assessed by probing for mRNA transcripts, immunoassays, or bioassays. Supernatants from clone A1 induced IgE synthesis by normal human peripheral blood mononuclear cells. IL-4 was shown to be necessary and sufficient for the induction of IgE by clone A1 supernatants. In contrast, IgE induction by clone A1 supernatants and recombinant (r) IL-4 was inhibited by IFN-γ. This suggests that the absence of IFN-γ from the IL-4-containing A1 supernatants was important for their IgE-inducing ability. Supernatants from clone A1 could also specifically induce the growth of cord blood cell progenitors and their differentiation into eosinophils but not into basophils. rIL-5, but not rIL-4 or rGM-CSF, also induced eosinophil maturation. These data suggest that IL-5 secreted by clone A1 was responsible for its ability to induce eosinophil differentiation. The implications of the concomitant production of IL-4 and IL-5 by a single T-cell clone are discussed.

Regulation of IgE synthesis.

The role of IgE in allergic disease is by now well recognized. The study of the cellular basis of IgE regulation has been actively pursued to gain insights into the pathogenesis of a disease that affects a considerable proportion of the population. More recently, the molecular events underlying IgE synthesis have become the focus of intense interest, in an attempt to characterize the signals involved in isotype-specific regulation of immunoglobulin synthesis. Recent evidence from different laboratories indicates that induction of IgE synthesis requires two signals: one is IgE isotype-specific, and is delivered by IL-4, the other is a B cell activating signal that can be delivered through a variety of pathways. We will herein review what is currently known about the mechanisms underlying the synthesis of IgE in humans, as they are revealed through cellular and molecular biology studies.

Role of protein tyrosine kinases and phosphatases in isotype switching: crosslinking CD45 to CD40 inhibits IgE isotype switching in human B cells

Protein tyrosine kinases and protein tyrosine phosphatases play an important role in the transduction of signals via antigen receptors in T and B cells, and in CD40-dependent B-cell activation. To examine the role of tyrosine kinases and phosphatases in B-cell isotype switching, we examined the effects of the engagement of the transmembrane phosphatase CD45 on the synthesis of IgE induced by IL-4 and anti-CD40 monoclonal antibody (mAb). Crosslinking CD45 to CD40 using biotinylated mAbs and avidin strongly inhibited CD40-mediated IgE synthesis in IL-4-treated human B cells. CD40/CD45 crosslinking did not affect epsilon germline transcription in B cells stimulated with IL-4, but strongly inhibited induction of S mu/S epsilon switch recombination as detected by a nested primer polymerase chain reaction assay. The B-cell src-type tyrosine kinase lyn, which is activated following CD40 engagement, is a potential target for the effects of CD45 observed in our experiments, because CD45/CD40 crosslinking resulted in the inhibition of CD40-mediated lyn phosphorylation and activation. These results suggest an important role for protein tyrosine kinases and phosphatases in CD40-mediated induction of isotype switching to IgE.

Regulation of IgE synthesis: from the membrane to the genes

ConclusionsThe data discussed above clearly indicate that the second signal(s) required for IgE production can be delivered to B cells through different pathways. At the present time, no unifying explanatory mechanism can reconcile all different observations. Not much is known about signalling via MHC class II molecules and CD40; the role played by hydrocortisone is not clear, and other molecules — in addition to CD40 and its ligand — might be involved in non-cognate T-B cell interactions. Signalling via CD21 is not sufficient for IgE induction, and EBV-induced proteins essential for IgE induction have not been identified. The observation that engagement of CD40 [32] and full expression of latent EBV proteins [21] share the ability to protect human B cells from programmed cell death (apoptosis) is certainly intriguing, but it is unclear whether this effect is in any way related to IgE induction. It seems plausible that the pathways mentioned above may share the ability to activate the switch recombination machinery in B cells in which the ɛ locus has been transcriptionally activated and made accessible by IL-4.

Regulation of human IgE synthesis.

Allergic diseases result from the interaction with IgE bound to cell surface receptors. Therefore, rational therapeutic approaches to allergic diseases would be aimed at decreasing IgE and/or at blocking the binding of IgE to effector cells such as mast cells and monocytes. Our investigation of the mechanism of IgE synthesis in man shows that IgE synthesis by peripheral blood mononuclear cells (PBMC) absolutely requires the presence of IL-4 and requires endogenous IL-6, because antibody to IL-6 inhibits IgE production completely. IgE synthesis requires T/B cell contact and involves interactions between B cell surface MHC Class II molecules and T cell surface receptors, as antibodies to both of these cell surface molecules inhibit IgE synthesis. Furthermore, alloreactive T cell clones which are unable to engage the B cell MHC Class II molecules fail to induce IgE synthesis in spite of their ability to secrete IL-4. Studies on the immunoglobulin sites that are involved in IgE binding to high affinity receptors on mast cells and basophils have used recombinant fragments of IgE to block mast cell binding. These studies suggest that a stretch of 76 amino acids which straddles the C epsilon 2 and C epsilon 3 domains is essential for this binding. Parallel studies on IgE binding to low affinity receptors on monocytes and B cells suggest that sequences within C epsilon 3 are involved in this binding. Peptides or analogues that inhibit IgE binding to its cellular receptors may be useful in the treatment of allergic diseases.

Interleukin 4 Dependent Induction of IgE Synthesis and CD23 Expression by the Supernatants of a Human Helper T Cell Clone

The supernatants of the human alloreactive helper T cell clone A1, which secretes interleukins (IL) 4 and 5, could induce IgE synthesis by normal peripheral blood mononuclear cell and expression of Fc epsilon R2/CD23 on normal human monocytes. Both effects were IL-4 dependent, since they could be induced by recombinant IL-4 alone and were inhibited by an anti-IL-4 antibody. The activation of such clones may contribute to the high IgE production and the high percentage of circulating CD23+ monocytes observed in allergic disorders.