Otitis media with effusion in an allergic animal model: A functional and morphological study.

OBJECTIVES: Allergy is considered as one of important etiologic factor of otitis media with effusion (OME). In present study, we evaluated the causal effect of allergy on OME in an animal model, and investigated the secondary effect of bacterial infection. METHODS: Allergy and control animals were subdivided into groups with and without intratympanic injection of lipopolysaccharide (IT-LPS). Allergic otitis media was induced via intraperitoneal ovo-albumin injection with intranasal challenge. We assessed the occurrence of OME in allergic animals and the effect of IT-LPS on allergic otitis media. We also investigated the Th1 and Th2 responses in the middle-ear mucosa. Hearing of the animals was measured by ABR and DPOAE. RESULTS: OME was observed in 75% of the allergic animals. After IT-LPS, 100% of the control and allergy groups showed otitis media. Light microscopy revealed that the middle-ear mucosa of animals of both groups also was significantly increased after IT-LPS, and the Th1 response (IL-2 and IFN-γ) and Th2 response (IL-5 and IL-13) cytokines were expressed at higher levels in the allergy group with IT-LPS than in control group with IT-LPS. Hearing tests between the allergy and control group with IT-LPS did not reveal any differences. CONCLUSION: Our findings may be direct evidence of an allergic causal effect on OME. Th2 response cytokines were strongly expressed in allergic OME, and the inflammatory reaction to LPS was more intense in the allergic group, which indicates that otitis media related to allergy can be severely aggravated by an inflammatory reaction to bacterial infection.

Effect of Retinoic Acid in a Mouse Model of Allergic Rhinitis.

PURPOSE: All-trans retinoic acid (ATRA) modulates immune responses by affecting T cells. Several studies have revealed that allergic inflammation of the lower airways is negatively associated with the vitamin A concentration. However, the role of ATRA in allergic inflammation of the upper airways is unclear. We investigated the effects of ATRA in an allergic rhinitis mouse model. METHODS: BALB/c mice except control groups (CON group) were sensitized with and challenged intra-nasally with Dermatophagoides farina (AR group). The ATRA groups were administered ATRA intraperitoneally. The steroid groups were administered steroid intranasally (ST group). Allergic symptoms and the average eosinophil number were counted. Cytokines and transcription factors were measured by Real-Time PCR and Western blotting. Der f-specific immunoglobulin E (IgE) was measured. Flow cytometry results of CD4⁺CD25⁺Foxp3⁺ T cells were analyzed. RESULTS: The symptom scores were lower in the ATRA group than in the AR group and higher than in the CON group. The levels of IgE were lower in the ATRA group than in the AR group and higher than in the CON and ST groups. The levels of Foxp3, TGF-ß, and IL-10 mRNA, as well as the percentage of CD4⁺CD25⁺Foxp3⁺ T cells, were higher in the ATRA group than in theAR group. In the ATRA group the levels of IFN-γ mRNA were higher, and the levels of GATA-3 and IL-4 mRNA, and ROR-γt were lower. In Western blotting analyses, the expression patterns of all factors, except Foxp3, showed similar to those of mRNA expression. CONCLUSIONS: ATRA has anti-allergic effects in an allergic rhinitis model, and its underlying mechanisms mainly include the induction of regulatory T cells and the inhibition of Th2 responses.

The Serine Protease Inhibitor, 4-(2-aminoethyl) Benzene Sulfonyl Fluoride Hydrochloride, Reduces Allergic Inflammation in a House Dust Mite Allergic Rhinitis Mouse Model.

PURPOSE: Serine protease inhibitors are involved in immune development, anti-inflammatory mechanisms, and tissue repair. In the present study, the serine protease inhibitor 4-(2-aminoethyl) benzene sulfonyl fluoride hydrochloride (AEBSF) was evaluated for its prophylactic and therapeutic applications in a mouse model of allergic rhinitis (AR). METHODS: BALB/c mice were divided into 5 groups: contol (CON), Dermatophagoides farinae (Derf), AR mice treated with AEBSF before sensitization (S), AR mice treated with AEBSF after challenge (C), and steroid groups. Derf was used as an allergen. AEBSF was administered before S or after C. Allergic symptom scores, eosinophil counts, proteolytic activity, interferon-γ, interleukin (IL)-10 levels and serum Derf-specific IgE levels were measured. T-bet, GATA-3, Foxp3, IL-13, and transforming growth factor (TGF)-ß mRNA levels were determined using real-time polymerase chain reaction. CD4(+)CD25(+)Foxp3(+) T cells were assessed using flow cytometry. RESULTS: Symptom scores, serum Derf-specific IgE levels, GATA-3 mRNA levels, IL-13 mRNA levels, and tissue eosinophil counts decreased in both the S and C groups (P<0.05). Additionally, the percentage of CD4(+)CD25(+)Foxp3(+) T cells, IL-10 levels, and Foxp3 mRNA levels increased in the S and C groups compared with those in the Derf group (P<0.05). AEBSF treatment decreased the proteolytic activity in the S and C groups (P<0.05). CONCLUSIONS: Prophylactic and therapeutic treatment with AEBSF significantly reduces allergic airway inflammation and can induce regulatory T cells in a murine model of AR.

Human placental extract reduces allergic inflammation in a murine allergic rhinitis model.

OBJECTIVES/HYPOTHESIS: In this study, we addressed the immunotherapeutic potential of human placental extract (HPE) in a murine allergic rhinitis (AR) model and explored its immunological mechanisms. STUDY DESIGN: In vivo study using an animal model. METHODS: HPE was administered to BALB/c mice before sensitization with allergen (Dermatophagoides farinae [Derf]) (pre-S group) or after allergen challenge (post-C group). The groups were compared with Derf-treated mice that received no HPE (Derf group) and phosphate buffered saline (PBS)-treated mice (control). Allergic symptom scores, eosinophil counts, and serum Derf-specific IgE levels were measured. mRNA expression levels of interferon (IFN)-γ, T-bet, interleukin (IL)-4, GATA-3, and Foxp3 in nasal mucosa were determined by real-time polymerase chain reaction. IFN-γ, T-bet, IL-4, and GATA-3 were confirmed by Western blotting analysis. Spleen CD4(+) CD25(+) Foxp3(+) T cells were detected using flow cytometry. RESULTS: Rubbing motions, serum Derf-specific IgE, GATA-3 mRNA levels, IL-4 mRNA levels, and tissue eosinophil counts were decreased in both pre-S and post-C groups (all P < 0.05). Western blots showed decreased expression of GATA-3 and IL-4 in both pre-S and post-C groups as compared to the Derf group. An increased percentage of CD4(+) CD25(+) Foxp3(+) T cells and an increased level of Foxp3 mRNA were found in pre-S and post-C groups as compared to those in the Derf group (all P < 0.05). CONCLUSION: Both prophylactic and therapeutic treatments with HPE significantly reduced allergic inflammation in nasal mucosa and had the potential to induce regulatory T cells in a murine model of AR.

Comparison of antiallergic effects of pneumococcal conjugate vaccine and pneumococcal polysaccharide vaccine in a murine model of allergic rhinitis.

OBJECTIVES/HYPOTHESIS: Pneumococcal vaccines have been widely used, and Streptococcus pneumoniae has been suggested to be an effective therapeutic agent in allergic disease. OBJECTIVES: The present study was performed to evaluate the effects of pneumococcal polysaccharide vaccine (PV) and pneumococcal protein conjugate vaccine (PCV), and to examine differences between the vaccines in a murine model of allergic rhinitis. STUDY DESIGN: In vivo study using an animal model. SETTING: Catholic Research Institutes of Medical Science. METHODS: Allergic rhinitis was induced in 40 BALB/c mice by intraperitoneal sensitization and intranasal challenge with Dermatophagoides farinae (Derf). The animals were divided into four groups: control, Derf, PV, and PCV. Interferon-γ, interleukin-13, and interleukin-10 levels in nasal lavage fluid and Derf-specific immunoglobulin E levels in serum were measured. The levels of T-bet, GATA-3, and Foxp3 mRNA expression in splenic mononuclear cells were determined. The number of CD4(+) CD25(+) Foxp3(+) regulatory T cells in splenic mononuclear cells was compared between groups by flow cytometry. RESULTS: Allergic symptom scores, T-bet and GATA-3 mRNA levels, serum Derf-specific immunoglobulin E levels, and tissue eosinophil counts were lower in the PV and PCV groups than the Derf group (P < 0.05). The regulatory T (Treg) cell indicators, Foxp3 mRNA, and percentages of CD4(+) CD25(+) Foxp3(+) T cells were increased in the PV and PCV groups (P < 0.05). CONCLUSION AND CLINICAL RELEVANCE: Both PV and PCV suppressed the allergen-specific T helper 2 response and induced regulatory T cells in a murine model of allergic rhinitis. However, PV and PCV may activate Treg cells via different mechanisms. LEVEL OF EVIDENCE: N/A.

The effect of pneumococcal polysaccharide vaccine in a mouse model of allergic rhinitis.

OBJECTIVES: This study aimed to determine if pneumococcal polysaccharide vaccine (PPV) could suppress allergic inflammation in an allergic rhinitis mouse model and to explore whether differences exist regarding the effect of PPV according to timing of administration. STUDY DESIGN: In vivo study using an animal model. SETTING: Catholic Research Institutes of Medical Science. SUBJECTS AND METHODS: BALB/c mice were divided into control, Der f, Pre-S, and Post-S groups. The allergen was Dermatophagoides farinae (Der f). Pneumococcal polysaccharide vaccine was administered before (Pre-S) or after (Post-S) sensitization. Allergic symptoms and eosinophils in nasal mucosa, interferon-γ, interleukin (IL)-13, and IL-10 in nasal lavage fluid and serum Der f-specific IgE were measured. T-bet, GATA-3, and Foxp3 mRNA in spleen were determined by real-time polymerase chain reaction. Flow cytometry of CD4(+)CD25(+)Foxp3(+) T cells in spleen was analyzed. RESULTS: In the Pre-S group, symptom score, serum Der f-specific IgE, eosinophils, IL-13, and GATA-3 mRNA were decreased (P < .05), and IL-10, Foxp3 mRNA, and CD4(+)CD25(+)Foxp3(+) T cells were increased compared with those in Der f group (P < .05). In the Post-S group, symptom score, serum Der f-specific IgE, and GATA-3 mRNA were decreased (P < .05), and Foxp3 mRNA and CD4(+)CD25(+)Foxp3(+) T cells were increased compared with those in the Der f group (P < .05). CONCLUSION: These results suggest that PPV administered before or after sensitization suppresses Th2 response and enhanced induction of regulatory T cells in an allergic rhinitis model. In addition, there was no significant difference between the degrees of effects in these 2 conditions. In the future, we can consider PPV to be a preventative agent for allergic rhinitis.

Comparative proteomic analysis for the insoluble fractions of colorectal cancer patients.

We used label-free quantitative proteomics with the insoluble fractions from colorectal cancer (CRC) patients to gain further insight into the utility of profiling altered protein expression as a potential biomarker for cancer. The insoluble fractions were prepared from paired tumor/normal biopsies from 13 patients diagnosed with CRC (stages I to IV). Fifty-six proteins identified in data pooled from the 13 cases were differentially expressed between the tumor and adjacent normal tissue. The connections between these proteins are involved in reciprocal networks related to tumorigenesis, cancer incidence based on genetic disorder, and skeletal and muscular disorders. To assess their potential utility as biomarkers, the relative expression levels of the proteins were validated using personal proteomics and a heat map to compare five individual CRC samples with five normal tissue samples. Further validation of a panel of proteins (KRT5, JUP, TUBB, and COL6A1) using western blotting confirmed the differential expression. These proteins gave specific network information for CRC, and yielded a panel of novel markers and potential targets for treatment. It is anticipated that the experimental approach described here will increase our understanding of the membrane environment in CRC, which may provide direction for making diagnoses and prognoses through molecular biomarker targeting.

The effect of topical FK506 (tacrolimus) in a mouse model of allergic rhinitis.

BACKGROUND: The management of allergic rhinitis (AR) encompasses education, pharmacotherapy, immunotherapy, and surgery. FK506 (tacrolimus) is an immunosuppressant that inhibits allergic reactions. The purpose of this study was to reveal whether FK506 treatment reduces allergic inflammation in an AR mouse model and to elucidate the mechanisms. METHODS: Forty mice were divided into four groups: control, AR, FK (FK506), and dexamethasone (DEX). All mice except for the control group were sensitized by an i.p. injection of ovalbumin (OVA). After sensitization, the FK and DEX groups were treated with FK506 and DEX intranasally. All sensitized mice were challenged intranasally with OVA. Allergic symptoms and tissue eosinophil counts were recorded. Interleukin (IL)-5, interferon gamma, and IL-10 levels in nasal lavage fluid (NALF) and serum OVA-specific IgE levels were measured. T-bet, GATA-3, and Foxp3 mRNA expression in splenic mononuclear cells were determined by real-time polymerase chain reaction. RESULTS: In the FK group and DEX group, allergic symptoms, serum OVA-specific IgE, tissue eosinophil counts, IL-5 in NALF, and GATA-3 mRNAs expression decreased (p < 0.05), and IL-10 in NALF and Foxp3 mRNAs expression increased compared with the AR group (p < 0.05). No significant difference was observed between the FK group and the DEX group. CONCLUSION: These results suggest that topical FK506 may reduce allergic inflammation and have potency equal to DEX in the AR model. This mechanism may involve not only Th2 cells but also regulatory T cells. Additional studies are needed on FK506, but in the future, we can consider FK506 as an alternative to topical steroids in the treatment of AR.

In-depth analysis of cysteine oxidation by the RBC proteome: advantage of peroxiredoxin II knockout mice.

Peroxiredoxin II (Prdx II, a typical 2-Cys Prdx) has been originally isolated from erythrocytes, and its structure and peroxidase activity have been adequately studied. Mice lacking Prdx II proteins had heinz bodies in their peripheral blood, and morphologically abnormal cells were detected in the dense red blood cell (RBC) fractions, which contained markedly higher levels of reactive oxygen species (ROS). In this study, a labeling experiment with the thiol-modifying reagent biotinylated iodoacetamide (BIAM) in Prdx II-/- mice revealed that a variety of RBC proteins were highly oxidized. To identify oxidation-sensitive proteins in Prdx II-/- mice, we performed RBC comparative proteome analysis in membrane and cytosolic fractions by nano-UPLC-MSE shotgun proteomics. We found oxidation-sensitive 54 proteins from 61 peptides containing cysteine oxidation, and analyzed comparative expression pattern in healthy RBCs of Prdx II+/+ mice, healthy RBCs of Prdx II-/- mice, and abnormal RBCs of Prdx II-/- mice. These proteins belonged to cellular functions related with RBC lifespan maintain, such as cytoskeleton, stress-induced proteins, metabolic enzymes, signal transduction, and transporters. Furthermore, protein networks among identified oxidation-sensitive proteins were analyzed to associate with various diseases. Consequently, we expected that RBC proteome might provide clues to understand redox-imbalanced diseases.

Proteomic analysis of protein expression affected by peroxiredoxin V knock-down in hypoxic kidney.

Peroxiredoxin V, an atypical thioredoxin peroxidase, is widely expressed in mammalian tissues. In addition, Prdx V is localized in mitochondria, peroxisome, cytosol, and the nucleus. Prdx V has been reported to protect a wide range of cellular environments as an antioxidant enzyme, and its dysfunctions may be implicated in several diseases, such as cancer, inflammation, and neurodegenerative disease. Identification and relative quantification of proteins affected by Prdx V may help identify novel signaling mechanisms that are important for oxidative stress response. However, the role of Prdx V in the modulation of hypoxia-related cellular response is not studied yet. To examine the function of endogenous Prdx V in hypoxic condition in vivo, we generated a transgenic mouse model with Prdx V siRNA expression controlled by U6 promoter. Of many tissues, the knockdown of Prdx V expression was displayed in the kidney, lung, and liver but not the spleen and skin. We conducted on the basis of nano-UPLC-MS(E) proteomic study to identify the Prdx V-affected protein networks in hypoxic kidneys. In this study, we identified protein networks associated with oxidative stress, fatty acid metabolism, and mitochondrial dysfunction. Our results indicated that Prdx V affected to regulation of kidney homeostasis under hypoxia stress.