Prophylactic effects of isomaltodextrin in a Balb/c mouse model of egg allergy.

The aim of this study was to evaluate the potential effects of isomaltodextrin (IMD), a dietary saccharide polymer derived from enzymatically produced from starch, on the ability to alter immune response (IR) bias to hen egg ovalbumin (Ova) induced allergic inflammation in mice. Groups of Balb/c mice were pre-treated with various doses of IMD in drinking water (1.0, 2.5, and 5.0% w/v) for 6 weeks and subsequently sensitized to the Ova together with continuous administration of IMD. To evaluate changes in immune response bias, immunoglobulin isotype-associated antibody activity, concentrations of type 1 and 2 cytokines and the percentage of T-regulatory cells (T-regs) in blood were measured. Clinical signs of allergy were assessed after oral challenge with Ova. Treatment with IMD did not significantly alter the frequency of clinical signs, however there was a trend in the overall reduction of clinical signs. Effect on IR bias was observed in the treatment groups as reflected by reduction in a type 1-biased phenotype as evident by decrease in isotype-specific IgE, IgG and increase in IL-12 cytokine production and a high proportion of T-regs. This study revealed that IMD could be a useful prophylactic candidate for alteration of allergic IR bias in mice and an immune-stimulator for reducing egg induced allergic reactions.

Comparison of Glycated Ovalbumin-Monosaccharides in the Attenuation of Ovalbumin-Induced Allergic Response in a BALB/C Mouse Model.

The aim of the present study was to compare various glycated ovalbumin (OVA)-monosaccharides, including OVA-mannose (Man), -glucose, -ribose, and -fructose, in the attenuation of OVA-induced allergic response in a BALB/C mouse model and the potential mechanisms of immunological modulation. The glycated OVA forms were prepared by Maillard reactions. OVA-Man significantly reduced the frequency of allergic signs. Mouse mast cell protease enzyme concentration was significantly reduced in the OVA-Man group (549.80 ± 84.67 ng/mL, p < 0.05). The OVA-Man group also had a lower histamine concentration (30.96 ± 1.12 ng/mL) as compared with the positive control OVA group (44.43 ± 0.71 ng/mL, p < 0.05). Both specific IgG and IgE were significantly reduced in the OVA-Man-treated group (p < 0.05). The OVA-Man group exhibited decreased concentrations of IL-4 (67.98 ± 3.11 pg/mL) and IL-17 (67.98 ± 3.11 pg/mL) and an increased concentration of IL-12 (336.70 ± 18.69 pg/mL, p < 0.05) compared with the positive control. Mannosylation played a vital role in allergen recognition, implicating deleterious downstream Th2 cell activation, cytokine secretion, and IgE production. This result indicates that different glycans target specific DC receptors, and differential DC processing, antigen presentation, and T cell response leads to altered variation in allergic response. OVA-Man exhibited minimal DC internalization, DC processing, MHC antigen presentation, and antigen-specific T cell activation, resulting in an attenuated allergic response and validating its efficacy as a potential immunotherapeutic candidate to treat egg allergy.

Oral administration of hen egg white ovotransferrin attenuates the development of colitis induced by dextran sodium sulfate in mice.

Ovotransferrin (OVT), one of the major hen egg white proteins, was shown to possess antimicrobial and antioxidant activities in vitro. However, there is no information regarding the in vivo preventative effect in chronic inflammatory diseases such as inflammatory bowel disease (IBD). The aim of the present study is to evaluate the anti-inflammatory effects of OVT in a mouse model of dextran sodium sulfate (DSS)-induced colitis. OVT (50 or 250 mg/kg BW) was given orally for 14 days to female BALB/c mice, and 5% DSS (MW 36-50 kDa) was used to induce acute colitis (days 7-14) via drinking water. The current in vivo study demonstrated that OVT significantly reduced clinical signs, weight loss, shortening of the colon, and inflammatory cytokine markers of disease. The histopathological analysis of the colon revealed that OVT reduced histological scores. These results indicate that the use of OVT may be a potential promising candidate for the prevention of IBD.

Hydrolysate from eggshell membrane ameliorates intestinal inflammation in mice.

Inflammatory bowel diseases (IBD) comprises of ulcerative colitis (UC) and Cohn's disease (CD) as two main idiopathic pathologies resulting in immunologically mediated chronic inflammatory conditions. Several bioactive peptides and hydro lysates from natural sources have now been tested in animal models of human diseases for potential anti-inflammatory effects. Eggshell membrane (ESM) is a well-known natural bioactive material. In this study, we aim to study the anti-inflammatory activity of ESM hydro lysate (AL-PS) in vitro and in vivo. In vitro, AL-PS was shown to inhibit pro-inflammatory cytokine IL-8 secretion. In vivo treatment with AL-PS was shown to reduce dextran sodium sulphate (DSS)-induced weight loss, clinical signs of colitis and secretion of interleukin (IL)-6 (p < 0.05). In addition, treatment with AL-PS also attenuated the severity of intestinal inflammation via down-regulation of IL-10 an anti-inflammatory cytokine. This validates potential benefits of AL-PS as a novel preventative target molecule for treatment of IBD.

Attenuation of allergic immune response phenotype by mannosylated egg white in orally induced allergy in BALB/c mice.

Food allergies are attributed to an imbalance in immune response to ubiquitous antigens. A previous study demonstrated that mannose glycation (mannosylation) of ovalbumin decreased allergenicity in vivo. The proposed research targets mannosylation of various common allergens that may help prevent food allergy. Balb/c mice (n = 8) were sensitized toxin egg white, peanut, and whey and treated with mannosylated forms of the test antigens. Glucosylated peanut and cholera toxins were used as controls. Allergic status was assessed as clinical signs, serum histamine, mouse mast cell protease (MMCP), antibody activity, cytokines, and T regulatory cells (T-regs). Significant preventative effects were observed with mannosylated egg white treatment such as reduced clinical signs, histamine, MMCP, specific G, G1, and E antibody activities, and IL-4 and increased IL-10 and CD25(+) Foxp3(+) cells. Other groups did not differ significantly. It was concluded that mannosylated egg white provides a powerful tool to prevent allergic phenotypes with possible relevance to control human egg allergy.

In vitro and ex vivo uptake of glutathione (GSH) across the intestinal epithelium and fate of oral GSH after in vivo supplementation.

Glutathione (GSH) is the most prevalent low-molecular-weight thiol in mammalian cells and is crucial for antioxidant defense, nutrient metabolism, and the regulation of pathways essential for whole body homeostasis. GSH transport systems have been identified in the membranes of various tissues and organs, including the small intestine. However, little is known regarding GSH transport across intestinal epithelial cells. The current in vitro and ex vivo uptake study of GSH demonstrated that intact GSH can be transported across intestinal epithelial cells, suggesting that GSH uptake is not proton-dependent. It would appear that the initial uptake of GSH into cells is a rapid process. Furthermore, the visualized GSH after 60 min of transport by MALDI-MS imaging showed localization of intact GSH inside the intestinal wall. In vivo study found that ingested (13)C-GSH was rapidly converted to GSSG and accumulated in red blood cells and liver, but was little present in plasma. The ingested GSH has potent nutraceutical benefits for human health to improve oxidative stress and defense in human.

Effects of ovalbumin glycoconjugates on alleviation of orally induced egg allergy in mice via dendritic-cell maturation and T-cell activation.

SCOPE: Glycation of allergens via Maillard reaction or chemical conjugation has been shown to influence susceptibility to food-induced allergies. It is hypothesized that mucosal immune response bias can be favorably altered by orally administering various forms of glycated ovalbumin (OVA). METHODS AND RESULTS: Groups of Balb/c mice (n = 10) were orally sensitized to OVA and administered various forms of glycated OVA (glucose, mannose, glucomannan, galactomannan, and a mixture containing OVA and glucomannan). Outcomes post oral challenge were measured as clinical allergic signs, serum histamine, mouse mast cell protease 1 (MMCP-1), antibody activity, type-1/2 cytokines, percentage of T-regulatory cells (T-regs) and in vitro dendritic cell, and T-cell-related mechanisms. Clinical signs and specific IgE were decreased (p ≤ 0.05), and T-reg cell percentage was increased in the mannose and glucomannan treated groups. The OVA-mannose group also had less histamine, MMCP-1, specific IgG, IL-4 and IL-17, and more IL-12p70 (p ≤ 0.05). Other parameters measured did not differ significantly among groups. Also, OVA-glycated mannose reduced maturation and uptake by dendritic cells. Less activation of T cells and type-2 cytokine response in DC-T-cell cocultures were observed with OVA-glycated mannose stimulation. CONCLUSION: This study validates, for the first time, the use of OVA-glycated mannose and glucomannan for potential beneficial dietary interventions for allergy.

Immune response phenotype of allergic versus clinically tolerant pigs in a neonatal swine model of allergy.

The prevalence of childhood food allergy and the duration of these allergies, particularly those considered to be transient, like egg and milk allergy, are increasing. The identification of allergic individuals using minimally invasive, non-anaphylaxis-threatening methods is therefore of increasing importance. In this experiment, correlates were sought of an allergic immune response (IR) phenotype in pigs. Using pigs pre-treated with heat-killed bacteria or bacterial components before allergic sensitization with the egg white protein ovomucoid (Ovm), differences were determined in IR phenotype of pigs in the categories treated-allergic, treated-tolerant, control-allergic (CA) and control-tolerant. Phenotype was established by measuring immunoglobulin (Ig)-associated antibody activity (AbA), cytokine profiles and the proportion of blood T-regulatory cells (T-regs) and observing late-phase allergen-specific skin tests (ST). Although 100% of pigs became sensitized to Ovm, only 33% of pigs had clinical signs of allergy after oral challenge with egg white. Pigs without clinical signs were classified as clinically tolerant. Sixty-seven percent of allergic pigs had a positive, late-phase ST classified as very strong or strong, while 84% of clinically tolerant pigs did not have late-phase ST. Treated-allergic pigs and CA pigs had greater total antibody IgG (H+L), IgE and IgG1 AbA than clinically tolerant pigs. Cytokine profiles of allergic pigs and the proportion of circulating T-regs, did not differ significantly between allergic and clinically tolerant pigs. Therefore, measurement of allergen-specific IgG, IgG1 and/or IgE activity and evaluation of late-phase ID ST may be useful in identifying allergic IR phenotypes in swine models of food allergy, which may be extended toward human use.

Prophylaxis of intranasally induced pollen allergy in a BALB/C mouse model using a potential prebiotic ß-1, 4 mannobiose.

BACKGROUND: Dietary supplementation with unique prebiotic nondigestible carbohydrates has been shown to suppress allergy. In the present study, the prophylactic efficacy of a disaccharide ß-1, 4 mannobiose (MNB) in a BALB/C mouse model of intranasally-induced pollen allergy was characterized. METHODS: Balb/c mice were pretreated with MNB orally and sensitized with pollen extract intraperitoneally and intranasally and challenged with histamine and crude pollen extract. Outcomes were measured as clinical signs, antibody isotypes, cytokine gene and protein expression patterns. RESULTS: The MNB-treated mice had lower sneezing frequency as compared to the positive control mice (P < 0.05). The low dose MNB-treated mice had less histamine (P < 0.05). However, the Cry j1 and Cry j 2-specific IgE, IgG, IgG1 and IgG2a antibody activity did not differ between groups (P > 0.05). The MNB-treated mice had increased IFN-γ (P < 0.05), and decreased IL-4 (P < 0.05). Mice in the high dose group had increased IL-10 (P < 0.05). However, TGF-ß and IL-17 concentration did not differ between groups (P > 0.05). Both total and Cry j1 and Cry j 2-specific IgA were increased in the high dose group. Real-time RT-PCR analysis indicated that IL-4 and IL-17 mRNA expression were lower in MNB-treated mice (P < 0.05). CONCLUSIONS: This work provides insights into using MNB as a potential prebiotic immunomodulator via decreased clinical signs, improved type1/type 2 balance, and IgA production, thus validating the potential use of MNB as a prophylactic prebiotic candidate to attenuate allergic response.

Therapeutic effects of ß1, 4 mannobiose in a Balb/c mouse model of intranasally-induced pollen allergy.

BACKGROUND: Nutritional prebiotic supplementation represents an attractive approach for interventions of allergy. In this study, the potential therapeutic effect of ß-1, 4 mannobiose (MNB) in a murine model of cedar pollinosis was investigated. METHODS: Groups of Balb/c mice were intranasally sensitized to Japanese cedar pollen extract, and subsequently administered with low or high dose MNB. Both intraperitoneal and intranasal challenges were performed to monitor for clinical signs. Frequency of sneezing was recorded. Serum, spleen and Peyer's patches were collected for various biomarker analyses. Anti-allergic activity of MNB using RBL-2H3 cells was also evaluated. RESULTS: Significant decrease in sneezing frequency, histamine, interleukin (IL)-4 and IL-17A and increase in TGF-ß and IL-10 concentration were exhibited by the MNB-treated mice. However, Cry j1 and Cry j 2-specific IgE activity remained unaltered. The high dose MNB treatment increased total IgA activity and IL-10, TGF-ß and FoxP3 and decreased IL-4, IL-17A, and RORγT mRNA expression. Inhibition of activation of RBL-2H3 cells was observed via decrease in histamine, intracellular Ca2+ concentration, and FcεRI mRNA expression. CONCLUSIONS: We demonstrated the immunomodulatory effects of MNB and conclude that MNB is a potential therapeutic molecular nutritional supplement candidate for treatment of pollen allergy.

Effect of heat-killed Escherichia coli, lipopolysaccharide, and muramyl dipeptide treatments on the immune response phenotype and allergy in neonatal pigs sensitized to the egg white protein ovomucoid.

Predisposition to food allergies may reflect a type 2 immune response (IR) bias in neonates due to the intrauterine environment required to maintain pregnancy. The hygiene hypothesis states that lack of early environmental stimulus leading to inappropriate development and bias in IR may also contribute. Here, the ability of heat-killed Escherichia coli, lipopolysaccharide (LPS), or muramyl dipeptide (MDP) to alter IR bias and subsequent allergic response in neonatal pigs was investigated. Three groups of three litters of pigs (12 pigs/litter) were given intramuscular injections of E. coli, LPS, MDP, or phosphate-buffered saline (PBS) (control) and subsequently sensitized to the egg white allergen ovomucoid using an established protocol. To evaluate change in IR bias, immunoglobulin isotype-associated antibody activity (AbA), concentrations of type 1 and 2 and proinflammatory cytokines released from mitogen-stimulated blood mononuclear cells, and the percentage of T-regulatory cells (T-regs) in blood were measured. Clinical signs of allergy were assessed after oral challenge with egg white. The greatest effect on IR bias was observed in MDP-treated pigs, which had a type 2-biased phenotype by isotype-specific AbA, cytokine production, and a low proportion of T-regs. LPS-treated pigs had decreased type 1- and type 2-associated AbA. E. coli-treated pigs displayed increased response to Ovm as AbA and had more balanced cytokine profiles, as well as the highest proportion of T-regs. Accordingly, pigs treated with MDP were more susceptible to allergy than PBS controls, while pigs treated with LPS were less susceptible. Treatment with E. coli did not significantly alter the frequency of clinical signs.

Recent advances in the role of probiotics in human inflammation and gut health.

The gastrointestinal (GI) tract provides residence to an astounding number of bacterial species, which have profound effects on host biology, function, physiology, and immune response. Discovery of "symbiosis factors" from symbionts that facilitate the peaceful coexistence of microbiota and the host immune system are of interest. Symbionts synthesize immunomodulatory molecules that guide maturation of the immune system and have pivotal roles in many biological processes; however, individuals differ in the makeup of their GI microbiota, which is influenced by many external and internal factors such as diet, antibiotic use, and host genetics, which in turn influences health and disease outcomes. Various endogenous, genetic, and environmental factors influence GI development including species composition and health status of neonates, resulting in interactions that occur between the bacteria and the host. Mechanisms of probiotics involved in homeostasis of a balanced immune system have been inconclusive. The probable mechanism of action may be postulated as direct competition between pathogenic bacteria in the gut and/or immune modulation. This review focuses on probiotics in health and disease prevention, especially the biological importance of intestinal regulation of inflammatory processes that may be beneficial in a multitude of disorders both inside and outside the GI tract.

Practical immunoregulation: neonatal immune response variation and prophylaxis of experimental food allergy in pigs.

The importance of environment in immune response is identified and the increase in prevalence of allergic, autoimmune and chronic inflammatory diseases reviewed. In particular, altered opportunity to acquire evolutionarily anticipated commensal microbiota is associated through the "hygiene hypothesis" with defective developmental and response signals to the innate and adaptive immune systems. Evidence of the detrimental effects of such environments is reviewed as is evidence for remediation using controlled exposure to bacteria or their active components such as LPS or peptidoglycan ligands for TLR and NOD-like receptors. Occurrence of major environmentally associated changes in porcine immune response phenotype are described. The prophylactic effects of heat-killed Escherichia coli given intramuscularly or of oral Lactococcus lactis on experimental ovomucoid-induced allergy in piglets are described in the context of altered immune response bias favouring reduced type-2 phenotypes. The high frequency of clinical tolerance to developing allergic signs even in the face of classical sensitization indicates possible function in this pig model of regulatory effectors such as Treg cells.

Immunomodulatory effects of heated ovomucoid-depleted egg white in a BALB/c mouse model of egg allergy.

Oral immunotherapy (OIT) is a promising therapeutic approach for treating food allergy. The treatment with heated ovomucoid-depleted egg white (HOMEW) in egg-allergic patients is noteworthy; however, OIT protocols are still experimental, and a better knowledge of the underlying mechanism is required. The objective of this work was to investigate the immunomodulatory effects of HOMEW and characterize the underlying mechanism in a BALB/c mouse model of egg allergy. Mice were sensitized with EW and treated with HOMEW. Post treatment, mice were challenged with EW and euthanized for collecting blood and spleen. Markers of allergic clinical outcomes were measured as histamine concentration, serum antibody activity, and cytokine production from cultured splenocytes. Digestibility of HOMEW was assessed mimicking physiological conditions in vitro. The HOMEW demonstrated high digestibility. The treatment induced a marked increase of the Th1/Th2 ratio in the high-dose treatment group. Treated mice had significantly less histamine, EW-specific IgE, and IL-4 and more IFN-γ and IL-10. This study confirms the mechanisms involved in successful tolerance induction with OIT using HOMEW and allows understanding of the vital role of surrogate allergy markers involved in immune modulation.

Porcine allergy and IgE.

Anaphylaxis was reported in 1963 in pigs experimentally sensitized with ovalbumin and was subsequently associated indirectly with IgE-related antibodies by functional assays to confirm heat-labile passive cutaneous anaphylaxis (PCA), reverse passive anaphylaxis (RPA) and Prausnitz-Küstner (PK) reactions to this and other allergens. The immunoglobulin mediating immediate hypersensitivity could be cross-adsorbed with anti-human IgE. Porcine IgE epsilon chain has been cloned and sequenced. Rabbit anti-pig IgE has been described by two groups, as has cross reactivity with pig IgE of various heterologous polyclonal and monoclonal anti-IgEs. Pigs develop transient post-weaning food allergy to soy allergens which can be prevented by pre-weaning feeding of soy proteins in sufficient quantity. Natural hypersensitivity also occurs to nematodes. Recently, experimental allergy has been induced in outbred pigs to peanut and to egg allergens which manifest as respiratory, cutaneous and enteric signs similar to those of human food allergy. These models are platforms for comparative allergy research as realistic alternatives to use of inbred mice or humans for investigation of pathogenesis, prophylaxis and therapy.

Attenuation of allergy to ovomucoid in pigs by neonatal treatment with heat-killed Escherichia coli or E. coli producing porcine IFN-gamma.

Food allergy is epidemic and prompts investigation to reduce allergic predisposition. It was hypothesized that heat-killed Escherichia coli injected intramuscularly (im) with or without interferon gamma (IFN-gamma), reduces neonatal susceptibility to experimental egg allergy. Two litters of Yorkshire pigs were assigned to three intramuscular treatment groups (four/group): control (PBS), heat-killed E. coli with or without IFN-gamma-expressing plasmid. Pigs were sensitized to ovomucoid (Ovm) by intraperitoneal injection with cholera toxin. To assess induction of allergy, pigs were fed egg white in yoghurt and assigned scores for allergic signs. Significantly fewer pigs developed allergy and passive cutaneous anaphylaxis in E. coli and E. coli+IFN-gamma vs control groups. E. coli-treated pigs also had significantly lower frequency of mean clinical scores. E. coli and E. coli+IFN-gamma groups did not differ. Serum antibody associated with IgG (H & L), IgG(1), IgG(2) or IgE all correlated but did not differ by treatment groups. Thus, treatment of neonatal pigs with heat-killed E. coli by im injection reduced susceptibility to allergic sensitization with Ovm. Inclusion of the type-1 cytokine, IFN-gamma, had no additional effect. Results indicate a method for prophylaxis of allergy and suggest support for the "hygiene hypothesis".

Porcine IgE in the context of experimental food allergy: purification and isotype-specific antibodies.

Measurement of allergen-specific immunoglobulin E (IgE) is a common practice in the investigation of allergy. It has not been possible to measure porcine IgE due to unavailability of anti-porcine IgE. This study was undertaken to purify and characterize porcine IgE from sera of allergic pigs, identify heterologous anti-IgE reactive with pig IgE and to use purified heavy (H) chain of porcine IgE to generate rabbit anti-IgE. A four-step process for the purification of porcine IgE is reported using ammonium sulphate precipitation, Protein G affinity chromatography, DEAE cellulose anion-exchange chromatography and sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) to obtain IgE H chain. The resultant IgE was evaluated for purity using SDS-PAGE and immunoreactivity was detected by Prausnitz-Küstner (PK) tests and passive cutaneous anaphylaxis with the allergen, crude peanut extract, used to induce experimental allergy. Cross-reactivity with anti-mouse and anti-human IgE antibodies were confirmed in western blot and enzyme-linked immunosorbent assays (ELISA). The H chain of IgE was excised from SDS-PAGE gels and used to develop rabbit anti-porcine IgE antisera. Antiserum obtained from rabbits immunized with porcine IgE, as well as heterologous murine and human-specific anti-IgE, induced reverse cutaneous anaphylaxis in pig skin and detected allergen-specific IgE in ELISA but did not react with IgG H chain in western blots. These results confirm allergy-associated bioactivity of porcine IgE and describe both homologous and heterologous anti-pig IgE suitable for use in allergen-specific and other assays. This will enhance utility of pig allergy models and provide an additional measure of type-2 immune response in pigs.

Expression of bioactive porcine interferon-gamma by recombinant Lactococcus lactis.

Lactococcus lactis is an immunomodulator and candidate live mucosal delivery vehicle for vaccine antigens and for biologically active molecules, including immunoregulatory cytokines such as interferon-gamma (IFN-gamma). To provide a tool for investigating downregulation of allergic predisposition of pigs to experimental food allergy, porcine IFN-gamma was cloned and expressed as a fusion protein with the usp45 secretion signal. Immunoblot analysis with polyclonal anti-pIFN-gamma-antibody demonstrated that the recombinant porcine IFN-gamma (rpIFN-gamma) protein was expressed in the L. lactis transformants as a secreted product. Activity of rpIFN-gamma was confirmed by ability to upregulate class II major histocompatibility complex (MHC) on cells of the porcine monocytic cell line 3D4/31. The L. lactis producing biologically active rpIFN-gamma will be used to investigate its possible ability to modulate the allergic immune response phenotype of pigs.

A neonatal swine model of allergy induced by the major food allergen chicken ovomucoid (Gal d 1).

BACKGROUND: Food allergy is a serious health problem for which a validated outbred large animal model would be useful in comparative investigations of immunopathogenesis and treatment and in testing hypotheses relevant to complex host-environmental interactions in predisposition to and expression of food allergy. OBJECTIVE: To establish a neonatal swine model of IgE-mediated allergy to the egg protein ovomucoid (Ovm) that may mimic human allergy. METHODS: In order to induce Ovm sensitivity, piglets at days 14, 21 and 35 of age were sensitized by intraperitoneal injection of 100 microg of crude Ovm and cholera toxin (50, 25 or 10 microg). Controls received 50 microg of cholera toxin in phosphate-buffered saline. The animals were challenged orally on day 46 with a mixture of egg white and yoghurt. Outcomes were reported as direct skin tests, clinical signs, IgG-related antibody and passive cutaneous anaphylaxis. RESULTS: Sensitized pigs developed immediate wheal and flare reactions, and after oral challenge, sensitized but not control animals displayed signs of allergic hypersensitivity. Serum IgG-related, Ovm-specific antibodies were detected only in the sensitized pigs and IgE-mediated antibody response to Ovm was confirmed by positive passive cutaneous anaphylaxis reactions induced by sera of sensitized but not by heat-treated sera from Ovm-sensitized pigs or sera of unsensitized control pigs. CONCLUSION: The present results confirm induction of Ovm-specific allergy in pigs and provide opportunity to investigate allergic predisposition and immunopathogenesis of IgE-induced Ovm allergy using outbred neonatal swine. This may better simulate allergic disease in humans and allow investigation of candidate prophylactic and therapeutic approaches.

Engineered recombinant ovomucoid third domain can modulate allergenic response in Balb/c mice model.

IgE-mediated allergic reactions to egg white are a serious health problem and ovomucoid being the dominant egg white allergen has been on focus in the past decade. Engineered hypoallergens with reduced reactivity for IgE antibodies are being examined to modulate the allergic response and develop prophylactic allergen vaccines. In this study, we evaluated the immunomodulatory effect of a genetic variant of the third domain of ovomucoid (GMFA) which showed reduced IgE binding with egg allergic patient's sera in comparison to the native form of the third domain of ovomucoid (DIII) in a murine model system. Balb/c mice were injected intraperitoneally with DIII and GMFA antigens. Allergen-specific serum IgG, IgG1, IgG2a, and IgE responses were evaluated using enzyme-linked immunosorbent assay. Splenocyte cytokine levels in the medium of the cultured cells were examined by ELISA and levels of IL-4, INF-gamma, and IL-12 (p70) cytokines were quantified. Neutralization with anti-IL-12 monoclonal antibody was assayed and cytokine levels with respect to GMFA mutant antigen stimulation were measured. GMFA mutant form was found to have significantly reduced levels of specific IgE when compared to the DIII suggesting a mutation-induced abrogation of the IgE binding epitope in mice. The increase in IgG2a levels in GMFA together with the decline of IgE and IgG1 points to a shift from a Th2 response to a Th1 dominated response. The cytokine profile showed a modulation of anti-allergic Th1 phenotype in GMFA from a proallergic Th2 response observed with DIII. Low levels of IL-4 and increased levels of INF-gamma and IL-12 were observed and anti-IL-12 monoclonal antibody restored the levels of IL-4 and suppressed levels of INF-gamma and IL-12 in the GMFA sensitized group. These results indicate that GMFA has a marked suppressive effect on the allergic response of ovomucoid and caused a shift towards a Th1 pathway, thereby modulating the Th1/Th2 cytokine balance and could be used as a potential hypoallergenic candidate for allergen-immunotherapy in the treatment of egg white allergy.