Nesprin1 deficiency is associated with poor prognosis of renal cell carcinoma and resistance to sunitinib treatment.

INTRODUCTION: Nuclear envelope spectrin repeat protein (Nesprin) 1 encoded by SYNE1, crucially regulates the morphology and functions of the cell. Mutations in the SYNE1 gene are associated with various diseases; however, their significance in renal cell carcinoma (RCC) remains unknown. In this study, we have investigated the association of SYNE1/Nesprin1 with the progression and prognosis of clear cell RCC (ccRCC). METHODS: In silico analyses of publicly available datasets of patients with RCC were performed. Based on the cohort data, Nesprin1 expression in nephrectomized tissue samples acquired from patients with ccRCC was analyzed using immunohistochemical staining. The invasion, migration, and proliferation of the SYNE1-knockdown human RCC cell lines were analyzed in vitro; moreover, RNA sequencing and Gene Set Enrichment Analysis were conducted to study the molecular mechanism underlying the association of SYNE1/Nesprin1 with prognosis of RCC. RESULTS: Patients with RCC-associated SYNE1 gene mutations exhibited significantly worse overall and progression-free survivals. Patients with Nesprin1-negative ccRCC tumors exhibit significantly poorer overall, cancer-specific, and recurrence-free survival rates than those recorded in the Nesprin1-positive group. SYNE1 knockdown enhanced the invasion and migration of RCC cells, however, it did not influence the proliferation of cells. RNA sequencing and Gene Set Enrichment Analysis revealed that SYNE1 knockdown significantly altered the expression of genes associated with oxidative phosphorylation. Consistently, patients with RCC exhibiting low SYNE1 expression, who were treated with the vascular endothelial growth factor receptor inhibitor sunitinib, had worse progression-free survival. CONCLUSIONS: The results indicate that the expression of SYNE1/Nesprin1 and SYNE1 mutations in patients with RCC are closely linked to their prognosis and responsiveness to sunitinib treatment.

Structure-activity relationship studies of pyrogallol as a calcineurin/NFAT signaling suppressor.

Previously, we have shown that pyrogallol alleviated nasal symptoms and suppressed IL-9 gene up-regulation in allergy model rats by inhibiting calcineurin/NFAT signaling. As pyrogallol has antioxidative activity, it may be responsible for inhibiting calcineurin/NFAT signaling-mediated IL-9 gene expression. However, the relationship between antioxidative activity and suppression of IL-9 gene expression has not been elucidated yet. Here, we conducted the structure-activity relationship studies of pyrogallol and its structurally related compounds to understand the mechanism of IL-9 gene suppression by pyrogallol. 2, 2-Diphenyl-1-picrylhydrazyl radical scavenging assay showed that the antioxidative activity of catechol, resorcinol, phloroglucinol, and gallic acid is 60.1%, 10.4%, 18.8%, and 113.5% of pyrogallol, respectively. Catechol, resorcinol, and phloroglucinol did not suppress NFAT dephosphorylation. Gallic acid suppressed dephosphorylation of NFAT. Gallic acid also suppressed ionomycin-induced up-regulation of IL-9 gene expression with the IC50 value of 82.6 µM. However, catechol, resorcinol and phloroglucinol showed no suppressive activity. In addition, using gallic acid-immobilized beads, we isolated and identified Poly(U)-binding-splicing factor 60 (PUF60) as a pyrogallol binding protein. These results suggest that the antioxidative activity of pyrogallol is not likely to be the mechanism of IL-9 gene suppression. Data also suggest that PUF60 is one of its target molecules responsible for the suppression of calcineurin/NFAT signaling by pyrogallol.

BACH1 promotes clear cell renal cell carcinoma progression by upregulating oxidative stress-related tumorigenicity.

The carcinogenesis and progression of renal cell carcinoma (RCC), a heterogeneous cancer derived from renal tubular epithelial cells, is closely related to oxidative stress responses (OSRs). Oxidative stress responses participate in various biological processes related to the metabolism and metastatic potential of cancer such as inflammation, epithelial-mesenchymal transition (EMT), and angiogenesis. In this study, we investigated the role of broad complex-tramtrack-bric-a-brac and cap 'n' collar homology 1 (BACH1), a key transcription factor for OSRs, in clear cell RCC (ccRCC) development and prognosis. The poor prognosis and elevation of serum inflammation markers in nephrectomized ccRCC patients were correlated with the intratumor expression of BACH1 accompanied by a downregulation of heme oxygenase-1. BACH1 contributes to the invasion and migration abilities of RCC cell lines without affecting their proliferation in vitro. In contrast, BACH1 contributes to tumor progression in vivo, in relation to OSRs with the activation of EMT-related pathways. BACH1 involvement in other OSR-linked pathways, including inflammatory responses, angiogenesis, and mTOR signaling, was further revealed by RNA sequencing analysis of BACH1-knockdown cells. In conclusion, the crucial role of BACH1 in the pathogenesis and poor prognosis of ccRCC through the promotion of OSRs is suggested.

Re-evaluation of over-the-counter histamine H1-receptor antagonists based on their effects on murine models of allergen-induced nasal hyperresponsiveness.

T cells play an essential role in the development of allergen-induced nasal hyperresponsiveness (NHR), a pathophysiological response in allergic rhinitis. The effects of histamine H1-receptor antagonists (antihistamines) on murine NHR models were investigated. Intragastric epinastine, fexofenadine, and loratadine administration suppressed allergen-induced immediate nasal response but not NHR in immunized mice. Regardless of the alleviation of stimulation-induced Th2 cytokine expression by loratadine and desloratadine in vitro, allergen-induced NHR and nasal eosinophil infiltration in Th2 cell-transferred mice were unaffected by loratadine in vivo. This influence on T cell-mediated NHR was excluded from the pharmacological effects of antihistamines.

Expression and Function of Nicotinic Acetylcholine Receptors in Induced Regulatory T Cells.

A contribution of the cholinergic system to immune cell function has been suggested, though the role of nicotine and its receptors in T cells, especially regulatory T (Treg) cells, is unclear. We herein investigated the expression and function of nicotinic acetylcholine receptors (nAChRs) in murine-induced Treg (iTreg) cells. Upon differentiation of naive BALB/c T cells into iTreg cells and other T-cell subsets, the effect of nicotine on cytokine production and proliferation of iTreg cells was examined. The expression of nAChRs and its regulatory mechanisms were comparatively analyzed among T-cell subsets. Stimulation-induced transforming growth factor-ß1 (TGF-ß1) production of iTreg cells was suppressed by nicotine, whereas interleukin (IL)-10 production and proliferation was not affected. α2-, α5-, α9-, and ß2-nAChRs were differentially expressed in naive, Th1, Th2, Th9, Th17, and iTreg cells. Among these cell types, the α9-nAChR was particularly upregulated in iTreg cells via its gene promoter, but not through tri-methylation at the 4th lysine residue of the histone H3-dependent mechanisms. We conclude that the immunoregulatory role of Treg cells is modified by the cholinergic system, probably through the characteristic expression of nAChRs.

Ablation of IL-17A leads to severe colitis in IL-10-deficient mice: implications of myeloid-derived suppressor cells and NO production.

IL-10 is an immune regulatory cytokine and its genetic defect leads to gastrointestinal inflammation in humans and mice. Moreover, the IL-23/Th17 axis is known to be involved in these inflammatory disorders. IL-17A, a representative cytokine produced by Th17 cells, has an important role for the pathological process of inflammatory diseases. However, the precise function of IL-17A in inflammatory bowel disease (IBD) remains controversial. In this study, we evaluated the effect of IL-17A on colitis in IL-10-deficient (Il10-/-) mice. Mice lacking both IL-10 and IL-17A (Il10-/-Il17a-/-) suffered from fatal wasting and manifested more severe colitis compared with Il10-/-Il17a+/- mice. Moreover, we found that CD11b+Gr-1+ myeloid-derived suppressor cells (MDSCs) accumulated in the bone marrow, spleen and peripheral blood of Il10-/-Il17a-/- mice. These MDSCs highly expressed inducible nitric oxide synthase (iNOS) (Nos2) and suppressed the T-cell response in vitro in a NOS-dependent manner. In correlation with these effects, the concentration of nitric oxide was elevated in the serum of Il10-/-Il17a-/- mice. Surprisingly, the severe colitis observed in Il10-/-Il17a-/- mice was ameliorated in Il10-/-Il17a-/-Nos2-/- mice. Our findings suggest that IL-17A plays suppressive roles against spontaneous colitis in Il10-/- mice in an iNOS-dependent manner and inhibits MDSC differentiation and/or proliferation.

Identification of novel interacting regions involving calcineurin and nuclear factor of activated T cells.

Nuclear factor of activated T cells (NFAT) leads to the transcription of diverse inducible genes involved in many biological processes; therefore, aberrant NFAT expression is responsible for the development and exacerbation of various disorders. Since five isoforms of NFAT (NFATc1-c4, NFAT5) exhibit distinct and overlapping functions, selective control of a part, but not all, of NFAT family members is desirable. By comparing the binding activity of each NFATc1-c4 with its regulatory enzyme, calcineurin (CN), using a quantitative immunoprecipitation assay, we found a new CN-binding region (CNBR) selectively functioning in NFATc1 and NFATc4. This region, termed CNBR3, is located between two preexisting CNBR1 and CNBR2, within the Ca2+ regulatory domain. The nuclear translocation of NFATc1 but not NFATc2 in T cells was suppressed by ectopic expression of CNBR3 and, accordingly, NFATc1-dependent cytokine expression was downregulated. Through competition assays using NFATc1-derived partial peptides and mass spectrometry with photoaffinity technology, we identified 18 amino acids in NFATc1 (Arg258 to Pro275 ) and 13 amino acids in CN catalytic subunit (CNA) (Asn77 to Gly89 ) responsible for CNA/CNBR3 binding in which Cys263 and Asp82 , respectively, played crucial roles. The possible selective regulation of NFAT-mediated biological processes by targeting this new CN/NFAT-binding region is suggested.

Isoform-Selective NFAT Inhibitor: Potential Usefulness and Development.

Nuclear factor of activated T cells (NFAT), which is the pharmacological target of immunosuppressants cyclosporine and tacrolimus, has been shown to play an important role not only in T cells (immune system), from which their name is derived, but also in many biological events. Therefore, functional and/or structural abnormalities of NFAT are linked to the pathogenesis of diseases in various organs. The NFAT protein family consists of five isoforms, and each isoform performs diverse functions and has unique expression patterns in the target tissues. This diversity has made it difficult to obtain ideal pharmacological output for immunosuppressants that inhibit the activity of almost all NFAT family members, causing serious and wide-ranging side effects. Moreover, it remains unclear whether isoform-selective NFAT regulation can be achieved by targeting the structural differences among NFAT isoforms and whether this strategy can lead to the development of better drugs than the existing ones. This review summarizes the role of the NFAT family members in biological events, including the development of various diseases, as well as the usefulness of and problems associated with NFAT-targeting therapies, including those dependent on current immunosuppressants. Finally, we propose a novel therapeutic strategy based on the molecular mechanisms that enable selective regulation of specific NFAT isoforms.

Downregulation of NFAT3 Due to Lack of T-Box Transcription Factor TBX5 Is Crucial for Cytokine Expression in T Cells.

The NFAT family transcription factors play crucial roles in immunological and other biological activities. NFAT3 is rarely expressed in T cells, and the mechanisms and significance of the specific NFAT3 downregulation in T cells have been unknown. In human CD4+ T cells, overexpression of NFAT1 and NFAT3 enhanced and suppressed IL-2 expression, respectively. NFAT3 downregulation in Jurkat cells using RNA interference technology augmented IL-2 expression, whereas a knockdown of NFAT1, NFAT2, and NFAT4 suppressed it. The promoter/enhancer activity of the NFAT-binding site in the IL-2 gene was upregulated and downregulated by NFAT1 and NFAT3, respectively. A study employing NFAT1/NFAT3 chimeric molecules revealed that the region in NFAT3 responsible for NFAT promoter activity inhibition was located within its N-terminal transactivation domain, Ca2+-regulatory domain, and DNA-binding domain. Downregulation of NFAT3 expression in T cells is mediated by lower chromatin accessibility and enhancer activity in its promoter in comparison with aortic smooth muscle cells expressing endogenous NFAT3. The binding sites of T-box transcription factor TBX5 and NK-2 transcription factor-related locus 5 Nkx2.5, which were expressed at higher levels in aortic smooth muscle cells than in T cells, were located within the -387 to +97 NFAT3 promoter region, exhibiting the maximum enhancer activity. Mutating the binding site of TBX5 but not Nkx2.5 diminished the NFAT3 promoter activity, whereas the overexpression of TBX5 enhanced it. Introduction of TBX5 into CD4+ T cells enhanced the expression of NFAT3 and suppressed that of IL-2. TBX5 deficiency-mediated downregulation of NFAT3 is crucial for the high cytokine-producing activity of T cells.

T Cell-Mediated Nasal Hyperresponsiveness in Allergic Rhinitis.

Allergic rhinitis patients suffer various symptoms such as sneezing, runny nose, and nasal congestion. As disease severity and chronicity progress, nasal hyperresponsiveness (NHR) develops in those patients. During the generation of a mouse allergic rhinitis model, we discovered that immunized mice developed NHR upon repeated nasal antigen challenge. Using genetically modified mice and an originally developed T cell-transferred mouse model, we confirmed the critical role of CD4+ T cells after differentiation into several helper subsets in NHR. On the other hand, immunoglobulin E/mast cell-dependent responses that are critical for evoking nasal symptoms and eosinophils that accumulate in allergic inflammation sites were dispensable. A steroid, but not drugs targeting mast cell-derived mediators, was effective in alleviating NHR. The possible generation of a new means to treat allergic rhinitis by targeting T cell-derived NHR-inducing factors is suggested.

Role of CD4+ T Cells in Allergic Airway Diseases: Learning from Murine Models.

The essential contribution of CD4+ T cells in allergic airway diseases has been demonstrated, especially by using various murine models of antigen-induced airway inflammation. In addition to antigen-immunized mouse models employing mast cell-deficient mice and CD4+ T cell-depleting procedure, antigen-specific CD4+ T cell transfer models have revealed the possible development of allergic inflammation solely dependent on CD4+ T cells. Regardless of the classical Th1/Th2 theory, various helper T cell subsets have the potential to induce different types of allergic inflammation. T cell receptor (TCR)-transgenic (Tg) mice have been used for investigating T cell-mediated immune responses. Besides, we have recently generated cloned mice from antigen-specific CD4+ T cells through somatic cell nuclear transfer. In contrast to TCR-Tg mice that express artificially introduced TCR, the cloned mice express endogenously regulated antigen-specific TCR. Upon antigen exposure, the mite antigen-reactive T cell-cloned mice displayed strong airway inflammation accompanied by bronchial hyperresponsiveness in a short time period. Antigen-specific CD4+ T cell-cloned mice are expected to be useful for investigating the detailed role of CD4+ T cells in various allergic diseases and for evaluating novel anti-allergic drugs.

Contribution of neuraminidase 3 to the differentiation of induced regulatory T cells.

Neuraminidase family enzymes that hydrolyze the terminal sialic acid linkage in biomolecules are involved in various immune responses. We previously showed that Th1 and Th2 cells differentially express several neuraminidases. Herein, the expression of neuraminidases in induced regulatory T (iTreg) cells was investigated in comparison with that in other T-cell subsets. Contrary to the tendency toward higher neuraminidase 1 mRNA expression in in vitro-differentiated Th2 cells, compared to Th1, Th17 and iTreg cells, we observed significantly higher expression of neuraminidase 3 (Neu3) in iTreg cells. Furthermore, the expression of Neu3 in FoxP3+ CD62L- spleen cells was higher than that in FoxP3+ CD62L+ and FoxP3- cells. Lentiviral expression of Neu3 in naïve CD4+ T cells during the stimulation culture led to upregulation of FoxP3 expression. On the basis of these findings, we conclude that Neu3 contributes to the differentiation of iTreg cells by upregulation of FoxP3.

Hyper-reactive cloned mice generated by direct nuclear transfer of antigen-specific CD4+ T cells.

T-cell receptor (TCR)-transgenic mice have been employed for evaluating antigen-response mechanisms, but their non-endogenous TCR might induce immune response differently than the physiologically expressed TCR Nuclear transfer cloning produces animals that retain the donor genotype in all tissues including germline and immune systems. Taking advantage of this feature, we generated cloned mice that carry endogenously rearranged TCR genes from antigen-specific CD4+ T cells. We show that T cells of the cloned mice display distinct developmental pattern and antigen reactivity because of their endogenously pre-rearranged TCRα (rTα) and TCRß (rTß) alleles. These alleles were transmitted to the offspring, allowing us to establish a set of mouse lines that show chronic-type allergic phenotypes, that is, bronchial and nasal inflammation, upon local administrations of the corresponding antigens. Intriguingly, the existence of either rTα or rTß is sufficient to induce in vivo hypersensitivity. These cloned mice expressing intrinsic promoter-regulated antigen-specific TCR are a unique animal model with allergic predisposition for investigating CD4+ T-cell-mediated pathogenesis and cellular commitment in immune diseases.

Potential Mechanisms of T Cell-Mediated and Eosinophil-Independent Bronchial Hyperresponsiveness.

Bronchial asthma is a chronic disease characterized by reversible airway obstruction, mucus production, and bronchial hyperresponsiveness (BHR). Although Th2 cell-mediated eosinophilic inflammation is an important disease mechanism in the majority of patients with bronchial asthma, recent studies suggest the possible development of Th2-independent airway inflammation and BHR. These non-Th2 endotype patients seem to consist of multiple subgroups, and often do not respond to inhaled corticosteroids. Therefore, to understand the pathogenesis of asthma, it is important to characterize these non-Th2 subgroups. Recently, we demonstrated that Th9 cells induce eosinophil infiltration and eosinophil-independent BHR, and Th9 cells-mediated BHR may be resistant to glucocorticoid. In this review, we summarize the contribution of several T cell subsets in the development of bronchial asthma and introduce our recent study demonstrating Th9 cell-mediated and eosinophil-independent BHR.

Identification of biomarker sets for predicting the efficacy of sublingual immunotherapy against pollen-induced allergic rhinitis.

Sublingual immunotherapy (SLIT) is effective against allergic rhinitis, although a substantial proportion of individuals is refractory. Herein, we describe a predictive modality to reliably identify SLIT non-responders (NRs). We conducted a 2-year clinical study in 193 adult patients with Japanese cedar pollinosis, with biweekly administration of 2000 Japanese allergy units of cedar pollen extract as the maintenance dose. After identifying high-responder (HR) patients with improved severity scores and NR patients with unchanged or exacerbated symptoms, differences in 33 HR and 34 NR patients were evaluated in terms of peripheral blood cellular profiles by flow cytometry and serum factors by ELISA and cytokine bead array, both pre- and post-SLIT. Improved clinical responses were seen in 72% of the treated patients. Pre-therapy IL-12p70 and post-therapy IgG1 serum levels were significantly different between HR and NR patients, although these parameters alone failed to distinguish NR from HR patients. However, the analysis of serum parameters in the pre-therapy samples with the Adaptive Boosting (AdaBoost) algorithm distinguished NR patients with high probability within the training data set. Cluster analysis revealed a positive correlation between serum Th1/Th2 cytokines and other cytokines/chemokines in HR patients after SLIT. Thus, processing of pre-therapy serum parameters with AdaBoost and cluster analysis can be reliably used to develop a prediction method for HR/NR patients.

Effects of anti-allergic drugs on T cell-mediated nasal hyperresponsiveness in a murine model of allergic rhinitis.

BACKGROUND: We have recently demonstrated that T cell-mediated nasal hyperresponsiveness (NHR) is a representative pathophysiological feature of allergic rhinitis (AR). Although several anti-allergic drugs are used for the treatment of AR, the efficacy of these drugs on T cell-mediated NHR have not been elucidated. In these studies we investigated the effects of dexamethasone (Dex), montelukast (Mk), and chlorpheniramine (Chl) on NHR in antigen-immunized and antigen-specific Th2 cell-transferred mice. METHODS: OVA-immunized BALB/c mice were treated with Dex, Mk, or Chl and challenged intranasally with OVA. We then assessed NHR, the number of inflammatory cells in the nasal lavage fluid (NALF), mRNA expression of Th2 cytokines in the nasal tissue, the population of CD3+CD4+ cells in the nasal lymphoid tissue (NALT), and antigen-specific serum IgE and IgG levels. Antigen-induced NHR and changes in antigen-specific T cells in the NALT were investigated in OVA-specific Th2 cell-transferred mice. RESULTS: Dex significantly suppressed antigen-induced NHR, inflammatory cell infiltration, and IL-4, IL-5, IL-6, and IL-13 expression in immunized mice. Chl was completely ineffective, and only IL-13 expression was suppressed by Mk. None of these drugs affected IgE and IgG production. Antigen-induced NHR and the increase in antigen-specific T cells in the NALT of Th2 cell-transferred mice were inhibited by Dex, but not by Mk or Chl. CONCLUSIONS: Steroids are effective for the reduction of NHR in AR by suppressing the accumulation of inflammatory cells, especially antigen-specific T cells.

Toll-like receptor 4 antagonist TAK-242 inhibits autoinflammatory symptoms in DITRA.

BACKGROUND: IL36RN encodes the IL-36 receptor antagonist (IL-36Ra), and loss-of-function mutations in IL36RN define a recessively inherited autoinflammatory disease named "deficiency of IL-36Ra" (DITRA). DITRA causes systemic autoinflammatory diseases, including generalized pustular psoriasis (GPP), an occasionally life-threatening disease that is characterized by widespread sterile pustules on the skin, fever and other systemic symptoms. GPP can present at any age, and provocative factors include various infections, medicines and pregnancy. OBJECTIVE: We aimed to elucidate the role of toll-like receptor 4 (TLR4) signaling in DITRA and to innovate an efficient treatment for DITRA. METHODS: We generated Il36rn-/- mice and treated them with TLR4 agonist to establish DITRA model mice. Furthermore, we administrated TLR4 antagonist TAK-242 to the model mice to inhibit the DITRA symptoms. RESULT: Il36rn-/- mice treated by TLR4 agonist showed autoinflammatory symptoms in skin, articulation and liver. Thus, we established model mice for DITRA or GPP that show cutaneous, articular, and hepatic autoinflammatory symptoms typical of DITRA or GPP: sterile pustules on the skin, liver abscesses and enthesitis of the hind paws. Additionally, these symptoms were canceled by TAK-242 administration. We demonstrated the inhibitory effects of the TLR4 antagonist TAK-242 on the autoinflammatory symptoms exhibited by the DITRA models. CONCLUSION: We suggested that blockage of TLR4 signaling is a promising treatment for DITRA and GPP.

Differential Contribution of Adhesion Molecules to Th1 and Th2 Cell-Mediated Lung and Bowel Inflammation.

CD4+ T cells play a critical role in the development of allergic inflammation in several target organs. Various adhesion molecules are involved in the local recruitment of T cells and other inflammatory cells. We investigated the differential contribution of adhesion molecules to T helper 1 (Th1) and Th2 cell-mediated allergic lung and bowel inflammation by employing their neutralizing antibodies. BALB/c mice transferred with in vitro-differentiated antigen-specific Th1 and Th2 cells were intratracheally or intrarectally challenged with a relevant antigen. Infiltration of infused T cells occurred, along with the accumulation of neutrophils and eosinophils in the lungs of Th1 and Th2 cell-transferred recipients, respectively. Th1-mediated neutrophil and Th2-mediated eosinophil accumulation in the large intestine, which occurred after intrarectal challenge with the antigen, was indicated by the significant elevation of myeloperoxidase (MPO) and eosinophil peroxidase (EPO) activity. Blocking experiments with neutralizing antibodies indicated that intercellular cell adhesion molecule (ICAM)-1; vascular cell adhesion molecule (VCAM)-1; and αL, ß2, and ß7 integrins participate in the accumulation of Th2 cells and eosinophils in the lungs. In contrast, the migration of Th1 cells and neutrophils was diminished by blockage of αL/ß2-integrin and ICAM-1, respectively. Mucosal addressin cell adhesion molecule (MadCAM)-1, vascular cell adhesion molecule (VCAM)-1, α4, ß1, and ß7 contributed to Th1-mediated neutrophilic inflammation in the bowel, though only MadCAM-1, α4, αL, and ß2 were involved in Th2-mediated eosinophilic inflammation. We conclude that distinct sets of adhesion molecules are involved in Th1- and Th2-mediated allergic lung and bowel inflammation.

Oral administration of ovalbumin after sensitization attenuates symptoms in a mouse model of food allergic enteropathy.

Oral immunotherapy (OIT) is a promising treatment of food allergy. To administer an appropriate oral dose of an allergenic component as OIT to individuals sensitized with a food allergen may prevent inducing food allergic inflammation in them. So we attempted to establish a mouse model to evaluate efficacy for oral administration of food allergen after sensitization. In BALB/c mice sensitized by injecting ovalbumin (OVA) with alum twice, OVA was administered before inducing inflammation by feeding the mice with egg white (EW) diet. Severe inflammatory responses, such as enteropathy, weight loss, IL-4 production, and increase of IgE antibody levels, were suppressed by administration with 4 mg of OVA 7 times before feeding EW diet. OVA administration alone induced a slight Th2 response, but no symptoms. The current study demonstrated that severe food allergic enteropathy could be prevented by pre-administration with appropriate dose of OVA to sensitized mice.