Rapid IgE desensitization is antigen specific and impairs early and late mast cell responses targeting FcεRI internalization

Rapid IgE desensitization provides temporary tolerization for patients who have presented severe hypersensitivity reactions to food and drugs, protecting them from anaphylaxis, but the underlying mechanisms are still incompletely understood. Thus, here we develop an effective and reproducible in vitro model of rapid IgE desensitization for mouse BM-derived mast cells (BMMCs) under physiologic calcium conditions, and we characterize its antigen specificity and primary events. BMMCs were challenged with DNP-human serum albumin conjugated (DNP-HSA) and/or OVA antigens, delivered either as a single dose (activation) or as increasing sequential doses (desensitization). Compared to activated cells, desensitized BMMCs had impaired degranulation, calcium flux, secretion of arachidonic acid products, early and late TNF-α production, IL-6 production, and phosphorylation of STAT6 and p38 mitogen-activated protein kinase (p38 MAPK). OVA-desensitized cells responded to DNP and DNP-desensitized cells responded to OVA, proving specificity. Internalization of specific antigen, IgE and high-affinity receptor for IgE (FcεRI) were impaired in desensitized BMMCs. Our results demonstrate that rapid IgE desensitization is antigen specific and inhibits early and late mast cell activation responses and internalization of the antigen/IgE/FcεRI complexes.     

Cbl family proteins: balancing FcεRI-mediated mast cell and basophil activation

The binding of IgE to high-affinity IgE receptors (FcεRI) expressed on the surface of mast cells and basophils initiates a cascade of signaling events that results in the release of a wide array of proinflammatory mediators. In order to limit the intensity and duration of cell activation, FcεRI aggregation has been understood to additionally generate negative signals through the coordinated action of adapters, phosphatases, and ubiquitin ligases. Among them, Cbl family proteins negatively regulate FcεRI-mediated signals mainly by promoting ubiquitination of the activated receptor subunits and associated protein tyrosine kinases. Notably, FcεRI ubiquitination has become recognized as an important signal for the internalization and delivery of engaged receptor complexes to lysosomes for degradation. The surface expression of activated FcεRI complexes is further downregulated through a pathway that is functionally separable from Cbl ligase activity and is dependent on the interaction of Cbl proteins with adapters involved in clathrin-dependent endocytosis. In this article, we review recent advances in our understanding of the molecular mechanisms through which Cbl proteins negatively regulate FcεRI-mediated mast cell and basophil functions.     

Transcriptional response of human mast cells stimulated via the FcεRI and identification of mast cells as a source of IL-11

Background  

In asthma and other allergic disorders, the activation of mast cells by IgE and antigen induces the cells to release histamine and other mediators of inflammation, as well as to produce certain cytokines and chemokines. To search for new mast cell products, we used complementary DNA microarrays to analyze gene expression in human umbilical cord blood-derived mast cells stimulated via the high-affinity IgE receptor (FcεRI).     

Technical advance: soluble OX40 molecule mimics regulatory T cell modulatory activity on FcεRI-dependent mast cell degranulation

Tregs play a central role in modulating FcεRI-dependent MC effector functions in the course of the allergic response. Cellular interaction depends on the constitutive expression of OX40 on Tregs and the OX40L counterpart on MCs. Study of OX40L signaling on MCs is hampered by the need of a highly purified molecule, which triggers OX40L specifically. We now report that sOX40 mimics the physiological activity of Treg interaction by binding to activated MCs. When treated with sOX40, activated MCs showed decreased degranulation and Ca(++) influx, whereas PLC-γ2 phosphorylation remained unaffected. Once injected into experimental animals, sOX40 not only located within the endothelium but also in parenchyma, where it could be found in close proximity and apparently bound to MCs. This soluble molecule triggers MC-OX40L without the requirement of Tregs, thus allowing study of OX40L signaling pathways in MCs and in other OX40L-expressing cell populations. Importantly, as sOX40 inhibits MC degranulation, it may provide an in vivo therapeutic tool in allergic disease.     

Human mast cell subsets - distinct functions in inflammation?

That mast cells participate in inflammatory reactions is beyond argument. A major question posed by mast cell biologists is whether specific functions in inflammation are subserved by different subsets of the mast cell population. We have investigated the two major subsets of human mast cells (MC(T) and MC(TC)), in the chronic inflammatory processes associated with rheumatoid arthritis (RA). Whereas normal synovium contains mainly MC(TC) mast cells, the MC(T) subset is selectively expanded in early RA, in numbers that correlate with synoviocyte hyperplasia and T-lymphocyte infiltration. In contrast, in RA of long duration, MC(TC) mast cells predominate in numbers that correlate with clinical indices of rapidity of disease progression. We suggest that MC(T) mast cells participate in active inflammatory events, whereas MC(TC) mast cells may be more relevant in repair or damage to connective tissues.     

Detection of circulating IgG autoantibody to FcεRIα in sera from chronic spontaneous urticaria patients

BackgroundsChronic spontaneous urticaria (CSU) is a common skin disorder characterized by itchy wheals of at least 6 weeks in duration, wherein the autoimmune mechanism is involved to activate IgE receptors (FcεRIα) on mast cells. We aimed to assess levels of IgG autoantibody against FcεRIα in sera from CSU patients using dot-blot immunoassay.MethodsWe performed a hospital-based cross-sectional study of 125 CSU patients (64 ASST-positive, 61 ASST-negative) and 64 age-and sex-matched healthy controls. The cut-off value of IgG FcεRIα autoantibody was determined as the mean intensity plus two standard deviations of values in controls. Positivity for IgG autoantibody to FcεRIα was analyzed according to clinical parameters of disease duration, urticaria activity score (UAS), ASST, response to antihistamine treatment, complement levels, and the presence of other autoantibodies. Nonparametric tests were applied for statistical analyses.ResultsIgG positivity to FcεRIα was noted in 24.8% of CSU patients and was significantly more frequent in ASST-positive patients than in ASST-negative patients (32.8% vs 16.4%, P = 0.040). Only 3.1% of healthy controls had this autoantibody. Complement 3 levels were significantly lower in anti-FcεRIα antibody-positive patients than antibody-negative patients (109.8 ± 19.9 vs 123.1 ± 30.9, P = 0.035). No significant associations were found between IgG positivity to FcεRIα and UAS, serum total IgE levels, atopic status, clinical responses to antihistamines, or the presence of anti-thyroid and anti-nuclear antibodies.ConclusionThese findings suggest that circulating IgG autoantibody to FcεRIα in a subset of patients may be involved in the autoimmune mechanism of CSU. Further studies are needed to clarify its clinical significance.     

An involvement of neurokinin-1 receptor in FcεRΙ-mediated RBL-2H3 mast cell activation

Objective and design

To determine whether the neurokinin-1 receptor (NK1R) plays a role in the activation of RBL-2H3 mast cells after FcεRΙ aggregation.

Materials and methods

NK1R expression in RBL-2H3 cells was inhibited by small hairpin RNA (shRNA) against NK1R, and determined by western blotting. For activation, both NK1R knockdown and control RBL-2H3 cells were sensitized by dinitrophenol (DNP)-specific IgE and stimulated with the antigen DNP-bovine serum albumin (BSA). Following the activation of RBL-2H3 cells, monocyte chemoattractant protein (MCP-1) production and intracellular calcium flux were monitored by ELISA and confocal microscopy assay, respectively. For investigation of the signaling mechanism, phosphorylation of mitogen-activated protein kinases (MAPKs) after RBL-2H3 cell activation was assessed by western blotting.

Results

shRNA-NK1R mediated an effective inhibition of NK1R expression in RBL-2H3 cells. Protein production of MCP-1 was reduced by more than 55?% in NK1R knockdown RBL-2H3 cells compared with control RBL-2H3 cells. In addition, both calcium mobilization and phosphorylation levels of MAPKs (Erk1/2, JNK, and p38) after DNP-BSA stimulation (via FcεRΙ) were decreased due to the inhibition of NK1R expression.

Conclusion

NK1R is required for the activation of RBL-2H3 cells following FcεRΙ engagement and involved in the regulation of MAPK signaling pathways.     

高亲和力FcεRI可介导IgE依赖的变应原提呈

高亲和力ＦｃεＲＩ可介导ＩｇＥ依赖的变应原提呈（文摘）［英／ＭａｕｒｅｒＤ…／／ＪＩｍｍｕｎｏｌ１９９５；１５４：６２８５」近来一些研究提示，ＩｇＥ可能在迟发型特应性免疫应答的发病机理中发挥一定的作用，这一认识基于以下的事实：①迟发型超敏反应的发生是...