Effect of Mandarin Orange Yogurt on Allergic Conjunctivitis Induced by Conjunctival Allergen Challenge.

Purpose: To evaluate the effects of mandarin orange yogurt containing nobiletin and ß-lactoglobulin on the allergic conjunctivitis induced by a conjunctival allergen challenge (CAC). Methods: Experiment 1 was performed on 26 asymptomatic patients (age, 25.3 ± 5.3 years) with proven seasonal allergic conjunctivitis due to cedar pollen. We compared the degree of conjunctivitis induced by CAC before and after ingesting mandarin orange yogurt for 2 weeks. Experiment 2 was a double-blind, placebo-controlled trial performed on 31 patients (age, 32.5 ± 12.2 years). A diet containing mandarin orange yogurt was compared to a diet containing yogurt lacking the mandarin orange on the conjunctivitis induced by CAC. The temperature of the inferior bulbar conjunctiva was measured before and 20 minutes after the CAC with an ocular surface thermographer (OST). The degree of conjunctival injection and chemosis was graded by slit-lamp biomicroscopy. The changes in the symptoms were evaluated by a questionnaire. Results: In experiment 1, the scores of redness (3.07 ± 3.03 vs. 1.05 ± 1.70), chemosis (2.84 ± 2.27 vs. 0.81 ± 1.11), itching (4.34 ± 3.05 vs. 1.39 ± 2.12), and temperature (0.73 ± 0.42°C vs. 0.45 ± 0.43°C) were significantly lower (P < 0.001) after a diet of mandarin orange yogurt for 2 weeks. In experiment 2, the scores of redness (1.03 ± 0.18 vs. 1.28 ± 0.52; P = 0.0156), itching (1.93 ± 1.92 vs. 2.82 ± 2.21; P = 0.0133), and surface temperature (0.54 ± 0.21°C vs. 0.31 ± 0.25°C; P < 0.001) were significantly lower in the mandarin orange yogurt group than in the control yogurt group. Conclusions: Mandarin orange yogurt can be an effective nutritional intervention for allergic conjunctivitis.

Methoxychlor enhances degranulation of murine mast cells by regulating FcεRI-mediated signal transduction.

Methoxychlor, an organochlorine insecticide developed to replace DDT (dichlorodiphenyltrichloroethane), has been reported to induce mast cell degranulation and to enhance IgE-mediated allergic responses. However, the mechanisms underlying these effects are not clear. To clarify potential mechanisms, the effects of methoxychlor on degranulation of mast cells were examined. Degranulation responses were evaluated using RBL-2H3 cells and mouse bone marrow-derived mast cells with either the antigen-induced or calcium ionophore-induced stimulation. Phosphorylation of enzymes related to signaling events associated with mast cell degranulation was analyzed by immunoblotting. Effects on vascular permeability in the passive cutaneous anaphylaxis reaction were evaluated following oral administration of methoxychlor to BALB/c mice. The results indicated that methoxychlor caused increased mast cell degranulation in the presence of antigen, whereas it had no effect on calcium ionophore-induced degranulation of RBL-2H3 cells. Immunoblot analyses demonstrated that the phosphorylation level of phosphoinositide 3-kinase (which plays a central role in mast cell signaling) was increased by methoxychlor during antigen-induced degranulation. In addition, methoxychlor activated the signaling pathway via the high-affinity IgE receptor by inducing phosphorylation of Syk and PLCγ1/2, which transfer the signal for degranulation downstream. Lastly, oral administration of methoxychlor exhibited a tendency to promote vascular permeability in passive cutaneous anaphylaxis model mice. Taken together, the results here suggested that methoxychlor enhanced degranulation through FcεRI-mediated signaling and promoted allergenic symptoms involved in mast cell degranulation.

CTLA-4 recombinant protein genetically fused to canine Fcepsilon receptor Ialpha enhances allergen specific lymphocyte responses in experimentally sensitized dogs.

Vaccination with a recombinant antigen fused to a targeting molecule is a potential strategy for inducing efficient immune responses. For the therapeutic purpose of allergic diseases in dogs, a DNA construct which expresses recombinant fusion protein with two functional domains, cytotoxic T lymphocyte antigen (CTLA-4) and Fcepsilon receptor Ialpha, was developed to bridge antigen-presenting cells and IgE-allergen complex. The recombinant fusion protein expressed by the DNA construct was demonstrated to retain the ability to bind monocytes in PBMC and dog IgE, respectively. Additionally, the recombinant protein induced enhancement of allergen-induced lymphoproliferation in experimentally sensitized dogs under conditions of suboptimal allergen stimulation. These results indicated that the DNA construct could enhance allergen-induced immune responses in vivo, implying its usefulness for perspective application in immunotherapy in dogs.

Immunological findings in 3 dogs clinically diagnosed with allergic rhinitis.

Three dogs clinically diagnosed with allergic rhinitis (AR) were examined for their immunological findings. House dust mites (HDM) such as Dermatophagoides farinae (DF) and D. pteronyssinus (DP) were identified as positive allergens in the 3 dogs with both intradermal skin test and serum antigen-specific IgE test. Lymphocyte blastogenic response of peripheral blood mononuclear cells (PBMCs) under stimulation with DF antigen in dogs with AR was higher than that in 4 healthy control dogs. Expression level of IL-4 mRNA in PBMCs obtained from the 3 AR dogs was higher than that in PBMCs obtained from 4 healthy control dogs before and after stimulation with DF antigen. Expression level of IFN-gamma mRNA in PBMCs was not different between the AR and control dogs before and after stimulation with DF antigen. These results suggested that allergic reaction to HDM antigen and T(H)2-type immune response were associated with the development of AR in 3 dogs examined in this study.

Cytokine profiles of peripheral blood mononuclear cells from dogs experimentally sensitized to Japanese cedar pollen.

Japanese cedar (Cryptomeria japonica, CJ) pollinosis is mediated by type-I hypersensitivity and induces seasonal rhinitis and conjunctivitis in humans. Previous studies showed that dogs could be experimentally sensitized with CJ pollen. In this study, we carried out quantitative analysis of mRNA levels of various cytokines in the peripheral blood mononuclear cells (PBMC) of 12 dogs experimentally sensitized to Japanese cedar pollen. Experimental sensitization was carried out by injection of crude CJ pollen extract with aluminium hydroxide gel. The expression levels of interleukin (IL)-1beta, IL-2, IL-4, IL-6, IL-8, IL-10, IL-18, interferon (IFN)-gamma, transforming growth factor (TGF)-beta(1), and tumor necrosis factor (TNF)-alpha mRNAs in the PBMC were quantified using a real-time sequence detection system. In the PBMC tested without culture, the expression levels of IL-8 and TNF-alpha mRNAs in experimentally sensitized dogs were significantly higher than those in control dogs. The expression level of IFN-gamma mRNA in the sensitized group was significantly lower than that in the control group. When the PBMCs were cultured in the presence of CJ pollen extract, the level of IL-4 mRNA expression was markedly increased in the PBMC from the experimentally sensitized dogs. In the PBMC stimulated with the CJ pollen extract, the expression level of IL-2 mRNA in the sensitized group was also significantly higher than that in the control group. Our data indicated that a Th2 response and proliferation of PBMC occur in response to the sensitizing antigen in dogs experimentally sensitized with CJ pollen, and revealed the presence of antigen-specific Th2 cells in this canine model. In addition, the expression levels of the mRNAs encoding proinflammatory cytokines were shown to be elevated after CJ pollen sensitization, indicating the activation of monocytes and macrophages.

Antigen-specific enhancements of CD80 mRNA expression in experimentally sensitized dogs with Japanese cedar pollen.

CD80, CD86, CD28 and Cytotoxic T lymphocyte antigen-4 (CTLA-4) are well-known co-stimulatory molecules that form the major co-stimulatory pathway essential for full activation of T cells. To investigate their role in pathogenesis of immune-mediated diseases, 12 dogs were sensitized experimentally to Japanese cedar pollen antigen (CPAg) as models of allergic diseases in dogs. After sensitization, lymphocyte stimulation test (LST) was carried out to evaluate reactivity to CPAg, and semi-quantitative real-time RT-PCR analysis of CPAg-stimulated peripheral blood mononuclear cells (PBMCs) to evaluate the expression of co-stimulatory molecules. As a result, CPAg-specific enhancements of CD80 expression were detected in all sensitized dogs. Furthermore, two different kinetics of its enhancements according to the blastgenic responses to CPAg were also observed. Expression of CD28, CTLA-4 and CD86 were suppressed following CPAg-stimulation. The result of the present study indicated the potential role of the CD28-CD80 co-stimulation pathway in pathogenesis of allergic diseases in dogs.