Novel IgE crosslinking-induced luciferase expression method using human-rat chimeric IgE receptor-carrying mast cells.

BACKGROUND: The measurement of antigen-specific serum IgE is common in clinical assessments of type I allergies. However, the interaction between antigens and IgE won't invariably trigger mast cell activation. We previously developed the IgE crosslinking-induced luciferase expression (EXiLE) method using the RS-ATL8 mast cell line; however, the method may not be sensitive enough in some cases. METHODS: In this study, we introduced an NF-AT-regulated luciferase reporter gene into the RBL-2H3 rat mast cell line and expressed a chimeric high-affinity IgE receptor (FcεRI) α chain gene, comprising an extracellular domain from humans and transmembrane/intracellular domains from rats. RESULTS: We generated multiple clones expressing the chimeric receptor. Based on their responsiveness and proliferation, we selected the HuRa-40 clone. This cell line exhibited significantly elevated human α chain expression compared to RS-ATL8 cells, demonstrating a 10-fold enhancement of antigen-specific reactivity. Reproducibility across different batches and operators was excellent. Moreover, we observed a detectable response inhibition by an anti-allergy drugs (omalizumab and cyclosporin A). CONCLUSIONS: HuRa-40 cells-which carry the human-rat chimeric IgE receptor-comprise a valuable reporter cell line for the EXiLE method. Their versatility extends to various applications and facilitates high-throughput screening of anti-allergy drugs.

Solar urticaria involves rapid mast cell STAT3 activation and neutrophil recruitment, with FcεRI as an upstream regulator.

BACKGROUND: Solar urticaria is a rare photodermatosis characterized by rapid-onset sunlight-induced urticaria, but its pathophysiology is not well understood. OBJECTIVE: We sought to define cutaneous cellular and molecular events in the evolution of solar urticaria following its initiation by solar-simulated UV radiation (SSR) and compare with healthy controls (HC). METHODS: Cutaneous biopsy specimens were taken from unexposed skin and skin exposed to a single low (physiologic) dose of SSR at 30 minutes, 3 hours, and 24 hours after exposure in 6 patients with solar urticaria and 6 HC. Biopsy specimens were assessed by immunohistochemistry and bulk RNA-sequencing analysis. RESULTS: In solar urticaria specimens, there was enrichment of several innate immune pathways, with striking early involvement of neutrophils, which was not observed in HC. Multiple proinflammatory cytokine and chemokine genes were upregulated (including IL20, IL6, and CXCL8) or identified as upstream regulators (including TNF, IL-1ß, and IFN-γ). IgE and FcεRI were identified as upstream regulators, and phosphorylated signal transducer and activator of transcription 3 expression in mast cells was increased in solar urticaria at 30 minutes and 3 hours after SSR exposure, suggesting a mechanism of mast cell activation. Clinical resolution of solar urticaria by 24 hours mirrored resolution of inflammatory gene signature profiles. Comparison with available datasets of chronic spontaneous urticaria showed transcriptomic similarities relating to immune activation, but several transcripts were identified solely in solar urticaria, including CXCL8 and CSF2/3. CONCLUSIONS: Solar urticaria is characterized by rapid signal transducer and activator of transcription 3 activation in mast cells and involvement of multiple chemotactic and innate inflammatory pathways, with FcεRI engagement indicated as an early event.

Evaluation value and mechanism of ADRB2 and FCER1B gene polymorphisms in preterm infants with congenital respiratory diseases.

In this study, we observed the value of ADRB2 and FCER1B gene polymorphisms in evaluating congenital respiratory diseases in preterm infants (PTIs), analyzed their effects on airway smooth muscle cells (ASMCs), and preliminarily discussed the underlying mechanism. First, we placed 64 healthy PTIs (control group) and 45 PTIs with congenital respiratory diseases (research group) born at our hospital from April 2021 to June 2023 were selected as the research subjects. Through testing, we found that the carriers of AA genotype of the polymorphic marker rs1042713 of the ADRB2 gene and that of the rs569108 locus of the FCER1B gene were less in the research group compared with the control group (P<0.05). Preterm infants carrying the GG genotype had a 2.887-fold (P<0.05) increased risk of developing congenital respiratory disease under the recessive model at the rs1042713 locus of the ADRB2 gene. Under the dominant model, preterm infants who did not carry the AA genotype had a 3.070-fold (P<0.05) increased risk of developing congenital respiratory disease. Subsequently, the constructed abnormal expression vectors of ADRB2 and FCER1B were transfected into ASMCs to examine changes in cell activity and pyroptosis. We found that up-regulating ADRB2 and FCERIB expression promoted ASMC proliferation and inflammatory reactions, inhibited apoptosis, and accelerated pyroptosis (P<0.05); silencing their expression, however, led to the opposite effect. In conclusion, the ADRB2 and FCERIB gene polymorphisms are strongly correlated with congenital respiratory diseases, which can provide a reference for clinical evaluation of congenital respiratory diseases in PTIs.

CD3D and CD247 are the molecular targets of septic shock.

Septic shock is a serious systemic disease with circulatory failure and abnormal cell metabolism caused by sepsis. However, the relationship between CD3D and CD247 and septic shock remains unclear. The septic shock datasets GSE33118 and GSE142255 profiles were generated from the gene expression omnibus databases GPl570, GPl17586. Differentially expressed genes (DEGs) were screened and weighted gene co-expression network analysis was performed. The construction and analysis of protein-protein interaction (PPI) network, functional enrichment analysis, gene set enrichment analysis (GSEA) were performed. Gene expression heat map was drawn. Immune infiltration analysis was performed. Comparative toxicogenomics database (CTD) analysis were performed to find the disease most related to the core gene. Targets can was used to screen miRNAs regulating the hub DEGs. 467 DEGs were identified. According to the gene ontology analysis, they were mainly enriched in the regulation of immune response, cell activation, signaling receptor activity, enzyme binding. Kyoto encyclopedia of genes and genomes analysis showed that they were mainly enriched in the TCR signaling pathway, Fc epsilon RI signaling pathway. GSEA showed that the DEGs were mainly enriched in immune response regulation, cell activation, TCR signaling pathway, Fc epsilon RI signaling pathway. Positive regulation of Fc receptor signaling pathway, PID IL12 2 pathway, immune response was observed in go enrichment items in the enrichment items of metascape. PPI networks got 5 core genes. Gene expression heat map showed that 5 core genes (CD247, Lck, cd3e, cd3d, ITK) were lowly expressed in the sepsis shock samples and highly expressed in the normal samples. CTD analysis showed that 5 core genes (CD247, Lck, cd3e, cd3d, ITK) were found to be associated with hemorrhage and necrosis. Low expression of cd3d, CD247 was observed in septic shock, and the lower the level of cd3d, CD247, the worse the prognosis.

A Co-Expressed Natural Antisense RNA FCER1A-AS Controls IgE-Dependent Immunity by Promoting Expression of FcεRIα.

As the α-subunit of the high-affinity receptor for the Fc portion of immunoglobulin E (FcεRIα), FcεRIα plays a central role in IgE-mediated allergic disorders and in the immunity and immunopathology of some parasitic infections. FcεRIα is specifically expressed on basophils and mast cells, but the mechanism that controls FcεRIα expression in these cells is poorly understood. In this study, we found that the natural antisense transcript (NAT) of FcεRIα (FCER1A-AS) is co-expressed with the sense transcript (FCER1A-S) in both interleukin (IL)-3-induced FcεRIα-expressing cells and in the high FcεRIα-expressing cell line MC/9. When FCER1A-AS is selectively knocked down by the CRISPR/RfxCas13d (CasRx) approach in MC/9 cells, the expression of both FCER1A-S mRNA and proteins is markedly decreased. Furthermore, FCER1A-AS deficiency was also found to be associated with a lack of FCER1A-S expression in vivo. Correspondingly, homozygous mice deficient in FCER1A-AS demonstrated a similar phenotype to FCER1A knockout mice in Schistosoma japonicum infection and in IgE-FcεRIα-mediated cutaneous anaphylaxis. Thus, we uncovered a novel pathway for the control of FcεRIα expression by its co-expressed natural antisense transcript. IMPORTANCE FcεRIα is responsible for high-affinity binding with the Fc portion of IgE, which is critical for IgE-dependent disease responses such as allergy responses and anti-parasite immunity. FcεRIα is expressed on a few cell types, including mast cells and basophils. Although the expression of FcεRIα is known to be promoted by the IL-3-GATA-2 pathway during its differentiation, the mechanism by which FcεRIα expression is maintained remains unknown. In this study, we discovered that a natural antisense transcript, FCER1A-AS, is co-expressed with the sense transcript. The presence of FCER1A-AS is essential for sense transcript expression in mast cells and basophils, but not for the differentiation of these cells through cis-regulation. Like FcεRIα knockout mice, mice lacking FCER1A-AS also exhibit reduced survival after Schistosoma japonicum infection and a lack of IgE-mediated cutaneous anaphylaxis. Thus, a novel pathway for regulating IgE-mediated allergic diseases through noncoding RNAs has been revealed.

Natural isoflavone formononetin inhibits IgE-mediated mast cell activation and allergic inflammation by increasing IgE receptor degradation.

Immunoglobulin (Ig)E-associated mast cell (MC) activation triggers pro-inflammatory signals that underlie type I allergic diseases. Here, we examined the effects of the natural isoflavone formononetin (FNT) on IgE-mediated MC activation and associated mechanisms of high-affinity IgE receptor (FcεRI) signal inhibition. The effects of FNT on the mRNA expression of inflammatory factors, release of histamine and ß-hexosaminidase (ß-hex), and expression of signaling proteins and ubiquitin (Ub)-specific proteases (USPs) were analyzed in two sensitized/stimulated MC lines. FcεRIγ-USP interactions were detected by co-immunoprecipitation (IP). FNT dose-dependently inhibited ß-hex activity, histamine release, and inflammatory cytokine expression in FcεRI-activated MCs. FNT suppressed IgE-induced NF-κB and MAPK activity in MCs. The oral administration of FNT attenuated passive cutaneous anaphylaxis (PCA) reactions and ovalbumin (OVA)-induced active systemic anaphylaxis (ASA) reactions in mice. FNT reduced the FcεRIγ chain expression, via increased proteasome-mediated degradation, and induced FcεRIγ ubiquitination by inhibiting USP5 and/or USP13. FNT and USP inhibition may be useful for suppressing IgE-mediated allergic diseases.

Kaempferol Suppresses the Activation of Mast Cells by Modulating the Expression of FcεRI and SHIP1.

In the present study, we evaluated the effects of kaempferol on bone marrow-derived mast cells (BMMCs). Kaempferol treatment significantly and dose-dependently inhibited IgE-induced degranulation, and cytokine production of BMMCs under the condition that cell viability was maintained. Kaempferol downregulated the surface expression levels of FcεRI on BMMCs, but the mRNA levels of FcεRIα, ß, and γ-chains were not changed by kaempferol treatment. Furthermore, the kaempferol-mediated downregulation of surface FcεRI on BMMCs was still observed when protein synthesis or protein transporter was inhibited. We also found that kaempferol inhibited both LPS- and IL-33-induced IL-6 production from BMMCs, without affecting the expression levels of their receptors, TLR4 and ST2. Although kaempferol treatment increased the protein amount of NF-E2-related factor 2 (NRF2)-a master transcription factor of antioxidant stress-in BMMCs, the inhibition of NRF2 did not alter the suppressive effect of kaempferol on degranulation. Finally, we found that kaempferol treatment increased the levels of mRNA and protein of a phosphatase SHIP1 in BMMCs. The kaempferol-induced upregulation of SHIP1 was also observed in peritoneal MCs. The knockdown of SHIP1 by siRNA significantly enhanced IgE-induced degranulation of BMMCs. A Western blotting analysis showed that IgE-induced phosphorylation of PLCγ was suppressed in kaempferol-treated BMMCs. These results indicate that kaempferol inhibited the IgE-induced activation of BMMCs by downregulating FcεRI and upregulating SHIP1, and the SHIP1 increase is involved in the suppression of various signaling-mediated stimulations of BMMCs, such as those associated with TLR4 and ST2.

5-Hydroxymethylfurfural multimers induce pseudo-allergic reaction through FcεRI at high doses.

Under acidic and high temperature conditions, 5-hydroxymethylfurfural (5-HMF) converted from sugar further produces dimers (Compound II) and trimers (Compound III). The polymers were less reported, and sensitization effect of them was reported in this study. Compounds II and III induced the local and systemic anaphylaxis effect in passive cutaneous anaphylaxis mice model and activated RBL-2H3 cell inducing [Ca2+ ] mobilization, resulting in the release of ß-hexosaminidase and histamine in vitro. The gene knockdown assay figured out that Compounds II and III induced degranulation through FcεRI. Further, Compounds II and III had a certain affinity with FcεRI by cell membrane chromatography and may combine on the "proline sandwich" structure indicated by molecular docking. All above suggested Compounds II and III can induce pseudo-allergic reaction through FcεRI in vivo and in vitro. Our work provides basic research to prove that the newly discovered 5-HMF transformants, Compounds II and III, induce pseudo-allergic reaction in vitro and in vivo through FcεRI, which is different pathway from 5-HMF. In foods with high sugar content, the sensitization of Compounds II and III needs more attention. In high-sugar foods and medicines, especially traditional Chinese medicine injections, the content of transformants needs to be detected.

Identification of redundancy between human FcεRIß and MS4A6A proteins points toward additional complex mechanisms for FcεRI trafficking and signaling.

BACKGROUND: Allergic diseases are triggered by signaling through the high-affinity IgE receptor, FcεRI. In both mast cells (MCs) and basophils, FcεRI is a tetrameric receptor complex comprising a ligand-binding α subunit (FcεRIα), a tetraspan ß subunit (FcεRIß, MS4A2) responsible for trafficking and signal amplification, and a signal transducing dimer of single transmembrane γ subunits (FcεRIγ). However, FcεRI also exists as presumed trimeric complexes that lack FcεRIß and are expressed on several cell types outside the MC and basophil lineages. Despite known differences between humans and mice in the presence of the trimeric FcεRI complex, questions remain as to how it traffics and whether it signals in the absence of FcεRIß. We have previously reported that targeting FcεRIß with exon-skipping oligonucleotides eliminates IgE-mediated degranulation in mouse MCs, but equivalent targeting in human MCs was not effective at reducing degranulation. RESULTS: Here, we report that the FcεRIß-like protein MS4A6A exists in human MCs and compensates for FcεRIß in FcεRI trafficking and signaling. Human MS4A6A promotes surface expression of FcεRI complexes and facilitates degranulation. MS4A6A and FcεRIß are encoded by highly related genes within the MS4A gene family that cluster within the human gene loci 11q12-q13, a region linked to allergy and asthma susceptibility. CONCLUSIONS: Our data suggest the presence of either FcεRIß or MS4A6A is sufficient for degranulation, indicating that MS4A6A could be an elusive FcεRIß-like protein in human MCs that performs compensatory functions in allergic disease.

FcεRI deficiency alleviates silica-induced pulmonary inflammation and fibrosis.

Silicosis is one of the most important occupational diseases worldwide, caused by inhalation of silica particles or free crystalline silicon dioxide. As a disease with high mortality, it has no effective treatment and new therapeutic targets are urgently needed. Recent studies have identified FCER1A, encoding α-subunit of the immunoglobulin E (IgE) receptor FcεRI, as a candidate gene involved in the biological pathways leading to respiratory symptoms. FcεRI is known to be important in allergic asthma, but its role in silicosis remains unclear. In this study, serum IgE concentrations and FcεRI expression were assessed in pneumoconiosis patients and silica-exposed mice. The role of FcεRI was explored in a silica-induced mouse model using wild-type and FcεRI-deficient mice. The results showed that serum IgE concentrations were significantly elevated in both pneumoconiosis patients and mice exposed to silica compared with controls. The mRNA and protein expression of FcεRI were also significantly increased in the lung tissue of patients and silica-exposed mice. FcεRI deficiency significantly attenuated the changes in lung function caused by silica exposure. Silica-induced elevations of IL-1ß, IL-6, and TNF-α were significantly attenuated in the lung tissue and bronchoalveolar lavage fluid (BALF) of FcεRI-deficient mice compared with wild-type controls. Additionally, FcεRI-deficient mice showed a significantly lower score of pulmonary fibrosis than wild-type mice following exposure to silica, with significantly lower hydroxyproline content and expression of fibrotic genes Col1a1 and Fn1. Immunofluorescent staining suggested FcεRI mainly on mast cells. Mast cell degranulation took place after silica exposure, as shown by increased serum histamine levels and ß-hexosaminidase activity, which were significantly reduced in FcεRI-deficient mice compared with wild-type controls. Together, these data showed that FcεRI deficiency had a significant protective effect against silica-induced pulmonary inflammation and fibrosis. Our findings provide new insights into the pathophysiological mechanisms of silica-induced pulmonary fibrosis and a potential target for the treatment of silicosis.

Cyclic Hypoxia Induces Transcriptomic Changes in Mast Cells Leading to a Hyperresponsive Phenotype after FcεRI Cross-Linking.

Mast cells (MCs) play important roles in tumor development, executing pro- or antitumoral functions depending on tumor type and tumor microenvironment (TME) conditions. Cyclic hypoxia (cyH) is a common feature of TME since tumor blood vessels fail to provide a continuous supply of oxygen to the tumor mass. Here, we hypothesized that the localization of MCs in cyH regions within solid tumors could modify their transcriptional profile and activation parameters. Using confocal microscopy, we found an important number of MCs in cyH zones of murine melanoma B16-F1 tumors. Applying microarray analysis to examine the transcriptome of murine bone-marrow-derived MCs (BMMCs) exposed to interleaved cycles of hypoxia and re-oxygenation, we identified altered expression of 2512 genes. Functional enrichment analysis revealed that the transcriptional signature of MCs exposed to cyH is associated with oxidative phosphorylation and the FcεRI signaling pathway. Interestingly, FcεRI-dependent degranulation, calcium mobilization, and PLC-γ activity, as well as Tnf-α, Il-4, and Il-2 gene expression after IgE/antigen challenge were increased in BMMCs exposed to cyH compared with those maintained in normoxia. Taken together, our findings indicate that cyH causes an important phenotypic change in MCs that should be considered in the design of inflammation-targeted therapies to control tumor growth.

Time will tell about mast cells: Circadian control of mast cell activation.

Mast cell activation is crucial to the development of allergic disease. New studies have shown that both IgE-dependent and -independent mast cell activation is temporally regulated by the circadian clock, a time-of-day-keeping system that consists of transcriptional-translational feedback loops of several clock genes. For instance, the core clock gene Clock controls the expression of the high-affinity IgE receptor (FcεRI) and interleukin-33 (IL-33) receptor ST2 on mast cells in a time-dependent manner. As a result, the threshold of IgE-dependent or IL-33-dependent mast cell activation differs between daytime and nighttime. This mechanism may underlie the observation that allergic disease shows a marked day-night change in symptom occurrence and severity. Consistent with this novel concept, environmental and lifestyle factors that disturb the normal rhythmicity of the circadian clock, such as irregular eating habits, can lead to the loss of circadian control of mast cell activation. Consequently, the degree of mast cell activation becomes equally strong at all times of day, which might clinically result in worsening allergic symptoms. Therefore, further understanding of the association between mast cell activation and the circadian clock is important to better manage patients with allergic disease in the real world, characterized by a "24/7 society" filled with environmental and lifestyle factors that disturb the circadian clock rhythmicity.

Single-cell transcriptomics reveals the identity and regulators of human mast cell progenitors.

Mast cell accumulation is a hallmark of a number of diseases, including allergic asthma and systemic mastocytosis. Immunoglobulin E-mediated crosslinking of the FcεRI receptors causes mast cell activation and contributes to disease pathogenesis. The mast cell lineage is one of the least studied among the hematopoietic cell lineages, and controversies remain about whether FcεRI expression appears during the mast cell progenitor stage or during terminal mast cell maturation. Here, we used single-cell transcriptomics analysis to reveal a temporal association between the appearance of FcεRI and the mast cell gene signature in CD34+ hematopoietic progenitors in adult peripheral blood. In agreement with these data, the FcεRI+ hematopoietic progenitors formed morphologically, phenotypically, and functionally mature mast cells in long-term culture assays. Single-cell transcriptomics analysis further revealed the expression patterns of prospective cytokine receptors regulating development of mast cell progenitors. Culture assays showed that interleukin-3 (IL-3) and IL-5 promoted disparate effects on progenitor cell proliferation and survival, respectively, whereas IL-33 caused robust FcεRI downregulation. Taken together, we showed that FcεRI expression appears at the progenitor stage of mast cell differentiation in peripheral blood. We also showed that external stimuli regulate FcεRI expression of mast cell progenitors, providing a possible explanation for the variable FcεRI expression levels during mast cell development.

Regulation of Trafficking and Signaling of the High Affinity IgE Receptor by FcεRIß and the Potential Impact of FcεRIß Splicing in Allergic Inflammation.

Mast cells are tissue-resident immune cells that function in both innate and adaptive immunity through the release of both preformed granule-stored mediators, and newly generated proinflammatory mediators that contribute to the generation of both the early and late phases of the allergic inflammatory response. Although mast cells can be activated by a vast array of mediators to contribute to homeostasis and pathophysiology in diverse settings and contexts, in this review, we will focus on the canonical setting of IgE-mediated activation and allergic inflammation. IgE-dependent activation of mast cells occurs through the high affinity IgE receptor, FcεRI, which is a multimeric receptor complex that, once crosslinked by antigen, triggers a cascade of signaling to generate a robust response in mast cells. Here, we discuss FcεRI structure and function, and describe established and emerging roles of the ß subunit of FcεRI (FcεRIß) in regulating mast cell function and FcεRI trafficking and signaling. We discuss current approaches to target IgE and FcεRI signaling and emerging approaches that could target FcεRIß specifically. We examine how alternative splicing of FcεRIß alters protein function and how manipulation of splicing could be employed as a therapeutic approach. Targeting FcεRI directly and/or IgE binding to FcεRI are promising approaches to therapeutics for allergic inflammation. The characteristic role of FcεRIß in both trafficking and signaling of the FcεRI receptor complex, the specificity to IgE-mediated activation pathways, and the preferential expression in mast cells and basophils, makes FcεRIß an excellent, but challenging, candidate for therapeutic strategies in allergy and asthma, if targeting can be realized.

Bidirectional Transport of IgE by CD23 in the Inner Ear of Patients with Meniere's Disease.

Meniere's disease (MD) is a disorder of the inner ear characterized by episodes of spontaneous vertigo, fluctuating hearing loss, and tinnitus. Recent studies have demonstrated that IgE may play a role in the pathogenesis of MD. Patients with MD (n = 103), acoustic neuroma (n = 5), and healthy subjects (n = 72) were recruited into the study. Serum from the participants was analyzed for IgE and type 2-related cytokines. IgE and CD23 expression levels in vestibular end organs of patients, C57BL/6 mice, or mouse HEI-OC1 cells were analyzed. Finally, the role of CD23 in IgE transcytosis was assessed using HEI-OC1 cells. Serum IgE was elevated in patients with MD and positively correlated with clinical symptoms. IL-4, IL-5, IL-10, IL-13, and CD23 levels were increased in patients with MD compared with the control group. In the transcytosis assay, mouse IgE was found to be bidirectionally transported across the HEI-OC1 cell monolayer. Additionally, CD23 downregulation using a small interfering RNA approach significantly reduced the efficiency of IgE transcytosis, suggesting that IgE is transported by CD23. Furthermore, exposure to IL-4 increased CD23 expression and enhanced IgE transcytosis in the HEI-OC1 cells and primary vestibular end organs. Our study indicated that IgE may play a role in the pathophysiology of MD. In addition, CD23-mediated IgE transcytosis in the hair cells may play a critical role in initiating inflammation in the inner ear. Thus, reducing the level of IgE may be a potentially effective approach for MD treatment.

The role of immunoglobulin E and mast cells in hypertension.

AIMS: Hypertension is the major cause of cardiovascular diseases and global mortality. Immunoglobulin E (IgE), which plays crucial roles in allergic diseases, has been implicated in the pathogenesis of vascular and cardiac remodelling via its receptor (FcεR1). In this study, we aimed to reveal the role of IgE and FcεR1 in hypertension. METHODS AND RESULTS: Herein, we reported that IgE levels were significantly increased in hypertensive patients as well as in hypertensive mice induced by angiotensin II (Ang II). Ang II-induced vascular remodelling and hypertension were significantly alleviated in FcεR1 genetic knockout mice or in mice treated with anti-IgE monoclonal antibody. Similarly, treatment with omalizumab (a clinical IgE antagonist) also markedly inhibited Ang II-induced hypertension. Furthermore, the cellular contribution of IgE-FcεR1 in hypertension was evaluated in mice with FcεR1 conditional knockout in mast cell (MC), smooth muscle cell (SMC), or endothelial cell (EC). Our data revealed that IgE-mediated hypertension is largely dependent on FcεR1 in MCs but not SMCs and ECs. Finally, RNA-seq and signalling pathway analyses of mouse bone marrow-derived MCs suggested that interleukin 6 (IL-6) is one of critical mediators in IgE-mediated hypertension. IL-6 derived from IgE-stimulated MCs promoted reactive oxygen species production and decreased the levels of phosphorylated endothelial nitric oxide synthase in ECs, leading to endothelial dysfunction. CONCLUSION: Our findings reveal that IgE contributes to the pathogenesis of hypertension, at least partially through activating the IgE-FcεR1 signalling in MCs. Thus, IgE may represent a new therapeutic target for IgE-mediated hypertension.

E-Cadherin restricts mast cell degranulation in mice.

Crosslinking of FcεRI-bound IgE triggers the release of a large number of biologically active, potentially anaphylactic compounds by mast cells. FcεRI activation ought to be well-controlled to restrict adverse activation. As mast cells are embedded in tissues, adhesion molecules may contribute to limiting premature activation. Here, we report that E-Cadherin serves that purpose. Having confirmed that cultured mast cells express E-Cadherin, a mast-cell-specific E-Cadherin deficiency, Mcpt5-Cre E-Cdhfl/fl mice, was used to analyze mast cell degranulation in vitro and in vivo. Cultured peritoneal mast cells from Mcpt5-Cre E-Cdhfl/fl mice were normal with respect to many parameters but showed much-enhanced degranulation in three independent assays. Soluble E-Cadherin reduced the degranulation of control cells. The release of some newly synthesized inflammatory cytokines was decreased by E-Cadherin deficiency. Compared to controls, Mcpt5-Cre E-Cdhfl/fl mice reacted much stronger to IgE-dependent stimuli, developing anaphylactic shock. We suggest E-Cadherin-mediated tissue interactions restrict mast cell degranulation to prevent their precocious activation.

Mitochondria-targeted triphenylphosphonium-based compounds inhibit FcεRI-dependent degranulation of mast cells by preventing mitochondrial dysfunction through Erk1/2.

AIMS: FcεRI-dependent activation and degranulation of mast cells (MC) play an important role in allergic diseases. We have previously demonstrated that triphenylphosphonium (TPP)-based antioxidant SkQ1 inhibits mast cell degranulation, but the exact mechanism of this inhibition is still unknown. This study focused on investigating the influence of TPP-based compounds SkQ1 and C12TPP on FcεRI-dependent mitochondrial dysfunction and signaling during MC degranulation. MAIN METHODS: MC were sensitized by anti-dinitrophenyl IgE and stimulated by BSA-conjugated dinitrophenyl. The degranulation of MC was estimated by ß-hexosaminidase release. The effect of TPP-based compounds on FcεRI-dependent signaling was determined by Western blot analysis for adapter molecule LAT, kinases Syk, PI3K, Erk1/2, and p38. Fluorescent microscopy was used to evaluate mitochondrial parameters such as morphology, membrane potential, reactive oxygen species and ATP level. KEY FINDINGS: Pretreatment with TPP-based compounds significantly decreased FcεRI-dependent degranulation of MC. TPP-based compounds also prevented mitochondrial dysfunction (drop in mitochondrial ATP level and mitochondrial fission), and decreased Erk1/2 kinase phosphorylation. Selective Erk1/2 inhibition by U0126 also reduced ß-hexosaminidase release and prevented mitochondrial fragmentation during FcεRI-dependent degranulation of MC. SIGNIFICANCE: These findings expand the fundamental understanding of the role of mitochondria in the activation of MC. It also contributes to the rationale for the development of mitochondrial-targeted drugs for the treatment of allergic diseases.

FcER1: A Novel Molecule Implicated in the Progression of Human Diabetic Kidney Disease.

Diabetic kidney disease (DKD) is a key microvascular complication of diabetes, with few therapies for targeting renal disease pathogenesis and progression. We performed transcriptional and protein studies on 103 unique blood and kidney tissue samples from patients with and without diabetes to understand the pathophysiology of DKD injury and its progression. The study was based on the use of 3 unique patient cohorts: peripheral blood mononuclear cell (PBMC) transcriptional studies were conducted on 30 patients with DKD with advancing kidney injury; Gene Expression Omnibus (GEO) data was downloaded, containing transcriptional measures from 51 microdissected glomerulous from patients with DKD. Additionally, 12 independent kidney tissue sections from patients with or without DKD were used for validation of target genes in diabetic kidney injury by kidney tissue immunohistochemistry and immunofluorescence. PBMC DKD transcriptional analysis, identified 853 genes (p < 0.05) with increasing expression with progression of albuminuria and kidney injury in patients with diabetes. GEO data was downloaded, normalized, and analyzed for significantly changed genes. Of the 325 significantly up regulated genes in DKD glomerulous (p < 0.05), 28 overlapped in PBMC and diabetic kidney, with perturbed FcER1 signaling as a significantly enriched canonical pathway. FcER1 was validated to be significantly increased in advanced DKD, where it was also seen to be specifically co-expressed in the kidney biopsy with tissue mast cells. In conclusion, we demonstrate how leveraging public and private human transcriptional datasets can discover and validate innate immunity and inflammation as key mechanistic pathways in DKD progression, and uncover FcER1 as a putative new DKD target for rational drug design.