Rat IgE Research Articles

Prevalence of rat and mouse sensitization in Asian patients with respiratory allergy.

Skin prick test (SPT) is useful in identifying rat and mouse sensitization. To determine the prevalence of rat and mouse sensitization by using local and commercial allergen extracts. Patients with allergic rhinitis or asthma were recruited. SPT of local and commercial rat and mouse allergen extracts were performed. The level of rat and mouse specific IgE (sIgE) was quantified in all patients with positive SPT and randomized patients with negative SPT. Two hundred and thirty patients, 108 male (47%) and median age 14 years (3.2-63.5 years), were enrolled. Rat sensitization by SPT was 11.7% and mouse sensitization was 17.8%. SPT result to local rat and commercial rat allergen extracts were moderately correlated (rs = 0.51, p < 0.001), while SPT result to local mouse and commercial mouse allergen extracts showed low correlation (rs = 0.38, p < 0.001). The concordance of SPT results between local rat and commercial rat allergen extracts was 90.4%. Concordance between the local mouse and commercial mouse allergen extracts was 85.2%. When compared with rat and mouse sIgE, the concordance of local rat, commercial rat and commercial mouse allergen extract were > 80% while that of local mouse was 54.4%. No adverse effect was observed in SPT with any allergen or extract. The prevalence of rat and mouse sensitization was low compared to the study in USA. SPT with local rat and mouse allergen extract was safe and showed good concordance with the SPT result of commercial allergen extracts and rat and mouse sIgE levels.

The role of skin inflammation, barrier dysfunction, and oral tolerance in skin sensitization to gluten-derived hydrolysates in a rat model.

Adverse reactions to wheat-containing skin care products have been linked to food allergy development. To determine the role of skin barrier dysfunction and inflammation in sensitization to gluten-derived hydrolysates via the skin in Brown Norway rats with and without oral tolerance to wheat. Skin barrier defect was induced by mechanical disruption, and skin inflammation was induced by topical application of SLS or MC903. Unmodified, enzyme hydrolyzed, or acid hydrolyzed gluten products were applied to the skin three times per week for 5 weeks. Subsequently, rats were orally gavaged with unmodified gluten. Wheat-naïve rats were readily sensitized to gluten hydrolysates via the skin. Skin barrier defect and skin inflammation had little effect on the skin sensitization and hydrolysate-specific IgE levels. Oral administration of unmodified gluten promoted the production of unmodified gluten-specific IgE in rats sensitized via the skin. Sensitization through intact skin, disrupted skin barrier, or inflamed skin was unable to break tolerance to unmodified gluten in rats on a wheat-containing diet. Mechanical skin barrier disruption and skin inflammation play a limited role in experimental skin sensitization to gluten-derived hydrolysates.

Development and characterization of a unique anti-IgE mouse monoclonal antibody cross-reactive between human and canine IgE.

BackgroundThe efficacy assessment of human anti‐IgE monoclonal antibodies (mAbs) in animal models before clinical trials is hampered due to the lack of cross‐reactivity of anti‐IgE mAbs between species.ObjectiveWe developed CRE‐DR (an anti‐dog IgE monoclonal antibody), an anti‐IgE mouse mAb that recognizes canine and human IgE, and then examined its IgE specificity and cross‐reactivity between three animal and human species.MethodsAfter mouse immunization with a synthetic peptide derived from canine IgE (282NTNDWIEGETYYC294), we generated a hybridoma producing CRE‐DR. The CRE‐DR purified from the ascites of hybridoma‐inoculated mice was used for ELISA and Western blot analysis to examine reactivity to dog, human, and rodent IgEs as well as recombinant bovine serum albumin (BSA)‐conjugated to canine, human, and rodent IgE amino acid peptides corresponding to the immunizing sequence. We then performed enzyme‐linked immunosorbent assays (ELISAs) for dog IgE using sera from dogs with atopic dermatitis (AD) after inhibition with canine IgE and IgG. The amino acid sequence recognized by CRE‐DR was identified by ELISA using synthetic peptides.ResultsCRE‐DR is a monoclonal mouse IgG1κ specific for dog IgE, and the ELISA values in atopic dog sera were inhibited by dog IgE, but not dog IgG. The binding of CRE‐DR to human IgE was relatively maintained, but not to rodent IgEs, which results were confirmed with the BSA‐conjugated IgE peptides of the various species. The CRE‐DR reactivity was supported by the comparison of amino acid sequence of CRE‐DR epitope, DWIEGETYYC, in dog IgE; one, two, and three amino acids were substituted in the human, rat, and mouse IgE epitopes, respectively.Conclusions and Clinical RelevanceCRE‐DR is a mAb cross‐reactive to dog and human IgEs, which can allow the use of a dog model of allergy to test the efficacy of a CRE‐DR‐derived anti‐IgE therapeutic mAb before human clinical trials.

Desloratadine Ameliorates Olfactory Disorder and Suppresses AMPA Receptor GluA1 Expression in Allergic Rhinitis Rat.

Allergic rhinitis (AR) is an IgE-mediated inflammation which causes olfactory dysfunction. Antihistamines have been widely used to treat AR while few studies have investigated the effect of antihistamines on improving the sense of smell. In addition, the underlying mechanisms are not well elucidated. We established the ovalbumin (OVA)-induced allergic rhinitis rat model and administrated desloratadine to AR rats. The AR symptoms, serum level of OVA-specific IgE and IL-17, and expression of IL-4, IL-5 and IL-13 in nasal mucosa were measured. The olfactory dysfunction was monitored by buried food test and the expression of GluR1 was measured. Desloratadine treatment alleviated AR symptoms, decreased serum level of OVA-specific IgE and IL-17 in AR rats. Desloratadine decreased IL-4, IL-5, and IL-13 expression in nasal mucosa of AR rats. Desloratadine ameliorated olfactory dysfunction in AR rats and decreased GluR1 expression in AR rats. Desloratadine treatment alleviated AR symptoms and ameliorated olfactory dysfunction in AR rats. The expression of AMPA receptor subunit GluR1 in olfactory bulb was associated with olfactory disorder.

Effects of Palm Oil Tocotrienol-Rich Fraction (TRF) and Carotenes in Ovalbumin (OVA)-Challenged Asthmatic Brown Norway Rats.

Synthetic therapeutic drugs for asthma, a chronic airway inflammation characterised by strong eosinophil, mast cell, and lymphocyte infiltration, mucus hyper-production, and airway hyper-responsiveness, exhibit numerous side effects. Alternatively, the high antioxidant potential of palm oil phytonutrients, including vitamin E (tocotrienol-rich fractions; TRF) and carotene, may be beneficial for alleviating asthma. Here, we determined the therapeutic efficacy of TRF, carotene, and dexamethasone in ovalbumin-challenged allergic asthma in Brown Norway rats. Asthmatic symptoms fully developed within 8 days after the second sensitization, and were preserved throughout the time course via intranasal ovalbumin re-challenge. Asthmatic rats were then orally administered 30 mg/kg body weight TRF or carotene. TRF-treated animals exhibited reduced inflammatory cells in bronchial alveolar lavage fluid. TRF- and carotene-treated rats exhibited notable white blood cell reduction comparable to that from dexamethasone. TRF- and carotene-treatment also downregulated pro-inflammatory markers (IL-β, IL-6, TNF-α), coincident with anti-inflammatory marker IL-4 and IL-13 upregulation. Treatment significantly reduced asthmatic rat plasma CRP and IgE, signifying improved systemic inflammation. Asthmatic lung histology displayed severe edema and inflammatory cell infiltration in the bronchial wall, whereas treated animals retained healthy, normal-appearing lungs. The phytonutrients tocotrienol and carotene thus exhibit potential benefits for consumption as nutritional adjuncts in asthmatic disease.

An immunologically relevant rodent model demonstrates safety of therapy using a tumour-specific IgE.

BackgroundDesigning biologically informative models for assessing the safety of novel agents, especially for cancer immunotherapy, carries substantial challenges. The choice of an in vivo system for studies on IgE antibodies represents a major impediment to their clinical translation, especially with respect to class‐specific immunological functions and safety. Fcε receptor expression and structure are different in humans and mice, so that the murine system is not informative when studying human IgE biology. By contrast, FcεRI expression and cellular distribution in rats mirror that of humans.MethodsWe are developing MOv18 IgE, a human chimeric antibody recognizing the tumour‐associated antigen folate receptor alpha. We created an immunologically congruent surrogate rat model likely to recapitulate human IgE‐FcεR interactions and engineered a surrogate rat IgE equivalent to MOv18. Employing this model, we examined in vivo safety and efficacy of antitumour IgE antibodies.ResultsIn immunocompetent rats, rodent IgE restricted growth of syngeneic tumours in the absence of clinical, histopathological or metabolic signs associated with obvious toxicity. No physiological or immunological evidence of a “cytokine storm” or allergic response was seen, even at 50 mg/kg weekly doses. IgE treatment was associated with elevated serum concentrations of TNFα, a mediator previously linked with IgE‐mediated antitumour and antiparasitic functions, alongside evidence of substantially elevated tumoural immune cell infiltration and immunological pathway activation in tumour‐bearing lungs.ConclusionOur findings indicate safety of MOv18 IgE, in conjunction with efficacy and immune activation, supporting the translation of this therapeutic approach to the clinical arena.

Construction and identification of the recombinant hFcεRIα/RBL-2H3 cells

IgE/FcεRI signal pathway plays a crucial role in triggering allergic reactions, and there is no cross-recognition between IgE and FcεRI in human and rats. In order to obtain the hFcεRIα/ RBL-2H3 cell line, total RNA was extracted from U937 cells, and the human FcεRIα gene was obtained by RT-PCR technology. Then the amplified product was digested and inserted into the pIRES2-EGFP vector. After the plasmid was transfected into the RBL-2H3 cells using lipofectamine, and the RBL-2H3 cell lines of stable expression were screened by G418. The transfection efficiency reached 60.45% with optimizing transfection parameters. The last the expression of hFcεRIα was detected by RT-PCR, western blotting and fluorescent microscopy. The present results demonstrated that the pIRES2-EGFP-hFcεRIα vector was constructed and a stable cell line of hFcεRIα/ RBL-2H3 cells was established successfully. This cell line is promising tools for further research on the pathogenesis and drug development of allergic diseases.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Anti-Folate Receptor-α IgE but not IgG Recruits Macrophages to Attack Tumors via TNFα/MCP-1 Signaling.

IgE antibodies are key mediators of antiparasitic immune responses, but their potential for cancer treatment via antibody-dependent cell-mediated cytotoxicity (ADCC) has been little studied. Recently, tumor antigen-specific IgEs were reported to restrict cancer cell growth by engaging high-affinity Fc receptors on monocytes and macrophages; however, the underlying therapeutic mechanisms were undefined and in vivo proof of concept was limited. Here, an immunocompetent rat model was designed to recapitulate the human IgE-Fcε receptor system for cancer studies. We also generated rat IgE and IgG mAbs specific for the folate receptor (FRα), which is expressed widely on human ovarian tumors, along with a syngeneic rat tumor model expressing human FRα. Compared with IgG, anti-FRα IgE reduced lung metastases. This effect was associated with increased intratumoral infiltration by TNFα+ and CD80+ macrophages plus elevated TNFα and the macrophage chemoattractant MCP-1 in lung bronchoalveolar lavage fluid. Increased levels of TNFα and MCP-1 correlated with IgE-mediated tumor cytotoxicity by human monocytes and with longer patient survival in clinical specimens of ovarian cancer. Monocytes responded to IgE but not IgG exposure by upregulating TNFα, which in turn induced MCP-1 production by monocytes and tumor cells to promote a monocyte chemotactic response. Conversely, blocking TNFα receptor signaling abrogated induction of MCP-1, implicating it in the antitumor effects of IgE. Overall, these findings show how antitumor IgE reprograms monocytes and macrophages in the tumor microenvironment, encouraging the clinical use of IgE antibody technology to attack cancer beyond the present exclusive reliance on IgG. Cancer Res; 77(5); 1127-41. ©2017 AACR.

Berberine improves airway inflammation and inhibits NF-κB signaling pathway in an ovalbumin-induced rat model of asthma

Objectives: Berberine has been reported for its various activities including anti-inflammatory effects and has been used in treating many diseases. However, its effects on airway inflammation in asthma have not been investigated. This study mainly aimed to detect its effects on the airway inflammation and the nuclear factor-κB (NF-κB) signaling pathway activity in a rat model of asthma. Methods: Asthma was induced by ovalbumin (OVA) sensitization and challenge. The asthmatic rats were respectively treated with vehicle PBS or berberine (100 mg/kg or 200 mg/kg) for 28 days. The control rats were treated with PBS. Inflammatory cells in bronchoalveolar lavage fluid (BALF) were counted and the lung inflammation was scored. Levels of NF-κB p65 (mRNA and protein), phosphorylated NF-κB p65 (p-NF-κB p65), inhibitory κB alpha (IκBα) (mRNA and protein) and phosphorylated IκBα (p-IκBα), as well as NF-κB p65 DNA-binding activity, were measured to assess the activity of NF-κB signaling pathway. Levels of the downstream inflammatory mediators of NF-κB signaling, IL-1β, IL-4, IL-5, IL-6, IL-13 and IL-17 in BALF, were measured. Besides, the serum levels of OVA-specific immunoglobulin (Ig)E were measured. Results: Results showed that OVA increased the number of inflammatory cells in BALF, elevated lung inflammation scores, enhanced the NF-κB signaling activity and promoted the production of IgE in rats. Berberine dose-dependently reversed the alterations induced by OVA in the asthmatic rats. Conclusions: The findings suggested a therapeutic potential of berberine on OVA- induced airway inflammation. The ameliorative effects on the OVA-induced airway inflammation might be associated with the inhibition of the NF-κB signaling pathway.

THE LEVEL OF GENERAL IMMUNOGLOBULIN E (IGE) IN RATS OF DIFFERENT AGE WITH ATOPIC DERMATITIS.

In the article we studied the level of general immunoglobulin E in rats of different age with atopic dermatitis. On the basis of the obtained results characteristics are given of levels the general IgE in rats of different age and it’s increasing with age. In rats which had been experimentally reproduced atopic dermatitis the average value of general IgE was increased in prepubertal rats, mature and old animals significantly (р<0,001). That can have a diagnostic and predictive value of this indicator at atopic dermatitis.

Effects of Ethanol Extract of Liriope platyphylla on Allergic Inflammation

This study investegated the effect of Liriope platyphylla (LP) on allergic reactions and its mechanism of action. We investigated the effect of LP on Evans Blue (EB) extravasation induced by anti-dinitrophenyl (DNP)-IgE in rats. We tested whether the ethanol extract of LP reduced ear skin thickness and historical changes induced by topical application of 2,4-dinitrofluorobenzene (DNFB) to ears of mice. We evaluated compound 48/80-induced release of histamine in rats peritoneal mast cell (RPMCs). We also investigated the regulatory effect of LP on the level of inflammatory mediators in PMACI-induced human mast cell (HMC-1); cytokine IL-6, IL-8, TNF-α in HMC-1, MAPKs (ERK, JNK and p38) in HMC-1. The ethanol extract of LP (81.3 ㎎/100 g body weight) significantly inhibited the PCA reaction compared with the control (P ＜ 0.05). However, LP did not prevent topical applications of DNFB-induced ear skin thickening and histological changes. In RPMCs, histamine release induced by compound 48/80 was significantly attenuated by LP at 100 ㎍/ml (P ＜ 0.05). LP extract (100 ㎍/ml) significantly reduced the PMACI-induced IL-6, IL-8, and TNF-α secretion via inhibition of ERK phosphorylation in HMC-1. In conclusion, the ethanol extract of LP inhibited mast cell-derived, immediate-type allergic reactions, and the result suggest the potential of LP for preventing allergic inflammatory disorders.

Effects of Angelica acutiloba on mast cell-mediated allergic reactions in vitro and in vivo

The root of Angelica acutiloba is a widely used herbal medicine which has been used as a typical therapeutic for allergic diseases in traditional medicine. This study was aimed to investigate the effects of A. acutiloba on allergic reactions in in vitro and in vivo models and its mechanism of action. A. acutiloba was extracted by maceration with 80% ethanol (AAE) and standardized by high-performance liquid chromatography. We investigated the effect of AAE on phorbol-12-myristate-13-acetate plus calcium ionophore A23187 (PMACI)-induced cytokine release; phosphorylation of JNK, ERK, and p38 in human mast cell-1 (HMC-1); and compound 48/80-induced release of histamine in rat peritoneal mast cells (RPMCs). We also investigated the effects on Evans blue (EB) extravasation induced by anti-DNP IgE in rats. Treatment with 1, 10 and 100 μg/ml AAE concentration-dependently inhibited the release of cytokines (tumor necrosis factor-α, interleukin (IL) -6, and IL-8) and phosphorylation of ERK and JNK induced by PMACI in HMC-1 cells, but it did not inhibit the phosphorylation of p38. It also inhibited compound 48/80-induced histamine release in RPMCs. Oral administration of 271 mg/kg AAE inhibited EB extravasation in a passive cutaneous anaphylaxis rat model. In conclusion, AAE inhibited mast cell-derived allergic reactions by inhibiting the release of histamine, the production of pro-inflammatory cytokines, and the phosphorylation of ERK and JNK.

Enhancement of the pulmonary allergic granulocyte recruitment in rats exposed to DMTI-II, a Kunitz-type inhibitor isolated from Dimorphandra mollis seeds

DMTI-II (23-kDa trypsin inhibitor purified from Dimorphandra mollis seeds) promotes acute inflammation accompanied by an early infiltration of eosinophils, a critical cell type involved in allergic diseases. We have evaluated here the capacity of DMTI-II to enhance the allergic pulmonary inflammation, looking over time to the leukocyte trafficking from bone marrow to peripheral blood, and their recruitment into the allergic airways. Male Wistar rats were sensitized and challenged with ovalbumin (OVA). At 2 to 16 h prior to OVA challenge, animals were exposed to DMTI-II (10 μg). Bronchoalveolar lavage fluid (BAL), circulating blood and bone marrow were examined at 24 h post-OVA challenge. Challenge with OVA significantly increased the influx of total inflammatory cells, neutrophils and eosinophils in BAL and lung tissue. Pre-exposure to DMTI-II potentiated total inflammatory cell and neutrophil recruitment ( p < 0.05). Neutropoiesis and neutrophilia accompanied pulmonary cell influx. Pre-exposure to DMTI-II also significantly increased eosinophil recruitment to BAL, an effect starting at 4 h, remaining markedly elevated at 16 h ( p < 0.05). Eosinopoiesis and eosinophilia (seen within 2 to 4 h) were also observed. Exposure to DMTI-II alone increased the IL-4 levels, and further increased the IL-4 levels in OVA-challenged rats. The levels of IgE, LTB 4 and eotaxin in OVA-challenged rats were greater compared with non-sensitized rats, but DMTI-II exposure failed to further enhance such levels. In summary, our study shows that DMTI-II itself presents granulocytopoietic activity, and enhances allergen-induced neutrophil and eosinophil mobilization from bone marrow to lung tissues that is accompanied by enhanced IL-4 production.

Novel IgE peptide‐based vaccine prevents the increase of IgE and down‐regulates elevated IgE in rodents

Immunotherapy with anti-IgE antibodies for treatment of allergy is promising but a short half-life and extremely high cost limit its application. We sought to develop IgE vaccines that induce longer-lasting auto-antibodies to neutralize self-IgE as an alternative therapy. The vaccine was made by conjugating three synthetic peptides corresponding to human IgE receptor-binding sites to a carrier, hepatitis B surface antigen. To test the immunogenicity of the vaccine, rats were immunized with the vaccine or hepatitis B surface antigen as control. Serum IgG titres to human IgE and the IgE of other species were measured. The inhibition by rat antisera of the binding of human IgE to its receptor was assessed by ELISA, flow cytometry analysis, and passive cutaneous anaphylaxis (PCA), and its ability to recognize receptor-bound IgE was examined. The in vivo effect of the vaccine was evaluated in trichosanthin-sensitized mice and rats. In the preventative study, vaccination started before sensitization commenced, while in the treatment study, vaccination started after sensitization. Sensitized mice and rats receiving injections of the carrier served as controls. Trichosanthin-specific IgE was measured using PCA. Sera from vaccine-immunized rats contained high titre antibodies that reacted with soluble and plate-bound but not with receptor-bound human IgE; they also reacted with mouse, rat, and dog IgE. Furthermore, the sera inhibited the binding of human IgE to its receptor in a dose-dependent manner. In preventative and treatment studies, serum trichosanthin-specific IgE levels were significantly reduced in vaccinated groups compared with controls. Antibodies against self-IgE can be induced by IgE peptide-based vaccines, which are effective in preventing the increase of IgE and in down-regulating IgE in sensitized animals.

Changes in expression of Hyaluronidase genes during immunization-induced IgE formation

Hyaluronan (HA) is a glycosaminoglycan that associates with various molecules to assemble a specific HA-rich extracellular matrix. Low and intermediate-molecular-weight HA induces cytokines, chemokines, and other mediators of inflammation and extracellular matrix (ECM) degradation. Hyaluronidases are a group of enzymes that degrade HA. As a model of patients with allergic diseases, we used egg albumin to induce high levels of IgE in rats and examined changes in the expression of hyaluronidase genes (Hyal1 and Hyal2) using reverse transcription-polymerase chain reaction (RT-PCR). The expression of Hyal2 was not affected by the overproduction of IgE induced with egg albumin, whereas the expression of the Hyal1 gene was suppressed. High-molecular-weight HA might be degraded by Hyal2 into intermediate-molecular-weight HA, which can trigger inflammation.

Efficient anti-IgE vaccination without anaphylactogenic properties

Rationale An allergen-independent therapeutic vaccine has been developed to reduce levels of IgE. The aim of this study was to characterize the immune response raised upon anti-IgE vaccination of rats. Methods The active vaccine component is a chimeric protein consisting of the constant domain 3 from rat IgE (Ce3) flanked by Ce2 and Ce4 domains from the American opossum. Wistar F rats were immunised subcutaneously at weeks 0, 3, 5 and 7 with vaccine adsorbed onto aluminum hydroxide or emulsified in the oil-based adjuvant Montanide ISA 51. The anti-IgE response and serum IgE concentrations were measured. The effect of concurrent ovalbumin (OVA) sensitization on IgE levels was also investigated. Furthermore, we evaluated potential crosslinking activity by anti-IgE from a number of rats on IgE bound to a mast cell line. Results The vaccine induced a pronounced anti-IgE response and reduced IgE levels with more than 90% upon administration with aluminum hydroxide or Montanide ISA 51. The vaccine also prevents increased levels of serum IgE by OVA. No sign of crosslinking activity by anti-IgE on receptor-bound IgE could be detected. Conclusions The present study demonstrates that the vaccine generates high anti-IgE titres and strongly reduced IgE levels also with an aluminum-based adjuvant. Moreover, this immune response does not crosslink receptor-bound IgE. Thus, the present study supports the concept of therapeutic IgE vaccination.

Mast Cell Degranulation Induced by Lectins: Effect on Neutrophil Recruitment

The mammalian lectin macrophage-derived neutrophil chemotactic factor (MNCF) and the plant lectin KM+ were characterized for their ability to activate and degranulate mast cells. The association between mast cell activation and the induction of neutrophil migration was also investigated. Incubation of rat peritoneal mast cells with these lectins resulted in degranulation and mediator release. By confocal microscopy, both lectins were evenly distributed on the cell surface. MNCF activated RBL-2H3 mast cells only if the cells had been sensitized with IgE. KM+ was able to activate either unsensitized or IgE sensitized RBL-2H3 cells. In microplate assays MNCF, but not KM+, bound to rat IgE. In rats that were depleted of mast cells, neutrophil recruitment by MNCF and KM+ were significantly reduced indicating that mast cell activation provides an amplification loop for the neutrophil recruitment induced by these lectins. The present study supports the concept that mammalian lectins play a fundamental role in innate immunity.

Immunization with cathepsin L proteinases CL1 and CL2 secreted by Fasciola hepatica elicit a preferential type 1 response based on IgG2a antibodies in rats.

Cathepsin L proteinases (CL1 and CL2), the major components of Fasciola hepatica excretion/secretion products (E/S) are considered potential antigens of a vaccine against fascioliasis. The humoral response elicited by CL1 and CL2 in rats either immunized with the enzymes or infected with F. hepatica has been analysed, examining specific IgE and IgG subclass dynamics. The experiment was continued for 10 weeks and peripheral blood eosinophilia was also determined. Infected rats presented peaks of eosinophilia at weeks 3 and 7 post-infection, while those immunized with CL1 and CL2 were no different from controls. Total IgE in infected rats increased up to week 5, reaching 30 microg(-1) in some cases, then decreased slowly and rising again towards the end of the experiment. Determination of specific IgE, carried out in sera previously absorbed with Protein G-Sepharose, reached a peak in infected rats between weeks 2 and 5, depending on the individual. In immunized rats both total and specific IgE levels remained around the pre-immunization values. With regard to the IgG subclass responses to E/S products, in infected rats IgG1 predominated over IgG2a, and the reverse was true in rats immunized with CL1 and CL2 and tested against the respective antigens. In all cases an increase in IgG1 and IgG2a antibody titres was seen, with maximum levels being reached later (weeks 6-7) in infected rats than in immunized ones (weeks 4-5). No IgG2b or IgG2c responses were detected in any of the groups studied.

Mast Cell-Derived VEGF Enhances the Passage of IgE FE-3 through the Rat Aortic Endothelial Cell Monolayer

Background: It is well known that the IgE-mediated allergic reaction in various extravascular tissues is induced by the mutual interaction of IgE, target cells (mast cells, MCs) and allergens. However, so far little has been known about the detailed mechanism whereby IgE in the circulating blood is transferred to the extravascular tissue. To examine whether or not MCs are involved in the permeability of IgE across rat aortic endothelial cells (RAECs) in vitro, we determined the permeability constant (PC) of dinitrophenyl-specific rat IgE (IgE FE-3) using a dual chamber system. Methods: Isolated RAECs obtained by a primary explant technique were seeded on a collagen-coated membrane in the upper chamber. MCs were collected from the peritoneal cavity of Wistar rats and suspended in the lower chamber. The time-dependent changes in concentration of IgE FE-3 (IgE) in the upper and lower chambers were measured by IgE capture ELISA after addition of IgE (10 µg/ml) to the upper chamber. Results: The cultured medium of IgE-stimulated MCs significantly increased the PC of IgE (9.86 ± 0.46 × 10^–6 cm/s), as compared to the value to which calcium ionophore A 23187-stimulated MCs increased the PC of IgE (7.97 ± 0.21 × 10^–6 cm/s). The increase of PC by IgE-stimulated MCs was most strongly inhibited by suramin (92.3% ± 1.89), and was weakly inhibited by tranexamic acid, cimetidine and diphenhydramine. In addition, the PC of IgE was increased with the increase in the MC-derived vascular endothelial growth factor/permeability factor (VEGF). Conclusions: After IgE is transferred from the circulating blood to the extravascular tissue, it may bind to FcΕRI of the MC and the other FcΕRI-bearing cells. The MC is then activated through the interaction of IgE and FcΕRI. Release of VEGF from the activated MC increases, and the VEGF enhances the permeability of IgE.