Shuichiro Endo

Suppressive effect of locally produced interleukin‐10 on respiratory syncytial virus infection

Interleukin (IL)‐10 is known to be a multifunctional cytokine. This study was designed to evaluate the role of IL‐10 during respiratory syncytial virus (RSV) infection using a C57BL/6 transgenic (TG) mouse model in which the expression of murine IL‐10 cDNA was regulated by a human salivary amylase promoter (IL‐10 TG mice). These mice expressed a large amount of IL‐10 in the nasal mucosa and in salivary glands. Viral replication in the respiratory tract after intranasal infection with RSV was suppressed significantly in IL‐10 TG mice compared to non‐transgenic controls. This suppression was IL‐10 specific, because it was prevented by treating mice with neutralizing anti‐IL‐10 antibodies. We also found that IL‐10‐stimulated T cells displayed cytotoxic activity against infected murine nasal epithelial cells. Previous data indicated that IL‐10 induces Fas ligand (L) expression on mouse T cells. Taken together, these data suggest that Fas/Fas L mediated cytotoxicity is involved in the suppression of RSV replication observed in IL‐10 TG mice after intranasal infection.

Cetirizine decreases interleukin-4, interleukin-5, and interferon-γ gene expressions in nasal-associated lymphoid tissue of sensitized mice

Although the action of cetirizine dihydrochloride (cetirizine), a potent histamine H1 receptor antagonist, has been well known, its effect on the cytokine profiles in the nasal immune inductive site has not been elucidated yet. We studied the effect of cetirizine on the cytokine profiles in the nasal-associated lymphoid tissue (NALT), which is a principal mucosal lymphoid tissue of the respiratory tract in rodents. Two different doses of cetirizine were given intraorally for 5 days before the nasal challenge of ovalbumin in sensitized mice. The sensitized group was given normal saline instead of cetirizine, and the nonsensitized group had no sensitization or medication. The cytokine gene expressions in the NALT taken from the mice were investigated with real-time quantitative reverse-transcription polymerase chain reaction. The effect of cetirizine on the allergic symptom score, histamine threshold, and the eosinophil count in the nasal septal mucosa were examined also. Compared with the normal mice, the sensitized mice showed significantly increased levels of interleukin (IL)-4 and IL-5 gene expression although the increase of interferon (INF)-γ gene expression was not significant. In the cetirizine groups, the levels of expression of IL-4, IL-5, and INF-γ in the NALT were significantly decreased compared with the sensitized group. The cetirizine groups also showed decreased allergic symptom score, histamine threshold, and eosinophil count in the nasal septal mucosa compared with the sensitized group. In conclusion, cetirizine reduced the levels of expression of IL-4, IL-5, and INF-γ in the NALT of ovalbumin-sensitized mice. Cetirizine also reduced the acute allergic symptom, histamine sensitivity, and eosinophil count in the nasal septal mucosa.

Mountain cedar pollen induces IgE-independent mast cell degranulation, IL-4 production, and intracellular reactive oxygen species generation

Cedar pollens cause severe allergic disease throughout the world. We have previously characterized allergenic pollen glycoproteins from mountain cedar (Juniperus ashei) that bind to allergen-specific immunoglobulin E (IgE). In the present report, we investigated an alternative pathway of mast cell activation by mountain cedar pollen extract through IgE-independent mechanisms. We show that mountain cedar pollen directly induces mast cell serotonin and IL-4 release and enhances release induced by IgE cross-linking. Concomitant with mediator release, high levels of intracellular reactive oxygen species (ROS) were generated, and both ROS and serotonin release were inhibited by anti-oxidants. These findings suggest that alternative mechanisms exist whereby pollen exposure enhances allergic inflammatory mediator release through mechanisms that involve ROS. These mechanisms have the potential for enhancing the allergenic potency of pollens.

Analysis of Helper T Cell Responses to Cry j 1-Derived Peptides in Patients with Nasal Allergy: Candidate for Peptide-Based Immunotherapy of Japanese Cedar Pollinosis

BACKGROUND Allergen specific immunotherapy is highly effective, but adverse events may occur during treatment. Peptide-based immunotherapy has been proposed as one of new strategies for reduction of allergic adverse reactions. We examined the possibility of candidate peptides for the development of peptide-based immunotherapy for Japanese cedar pollinosis. METHODS Twelve Cry j 1-specific T-cell lines were established from peripheral blood mononuclear cells (PBMC) of 12 patients with Japanese cedar pollinosis. Using these T-cell lines, 37 Cry j 1-derived overlapping peptides were assessed for their proliferative responses and cytokine production. RESULTS Four peptides corresponding to the Cry j 1 sequence were able to induce proliferative responses to more than one T-cell line: p61-80 (3/12; 25.0%); p115-132 (2/12; 16.6%); p206-225 (4/12; 33.3%); and p337-353 (5/12; 41.7%). Furthermore, T-cell lines generated from 11 of 12 donors (91.7%) responded to at least one of these four peptides. On the other hand, the pattern of cytokine production from Cry j 1-specific T-cell lines varied. Moreover, cytokine production patterns by stimulation with Cry j 1 peptide did not reflect those by stimulation with Cry j 1 protein. CONCLUSIONS Our results suggest four Cry j 1-derived peptides (p61-80, p115-132, p206-225 and p337-353) may be considered to be the immunodominant T-cell epitopes of the Cry j 1 molecule, and can be useful for the design of peptide-based immunotherapy for the management of Japanese cedar pollinosis.

Trial of pranlukast inhibitory effect for cedar exposure using an OHIO chamber

Abstract Objective: In practical guidelines for management of allergic rhinitis in Japan, pranlukast is a leukotriene receptor antagonist recommended for the treatment of pollinosis. However, the effect of pranlukast on nasal symptoms for cedar pollinosis has not been thoroughly investigated. The aim of this study is to examine this effect in a double-blind controlled crossover study using a pollen challenge chamber (the OHIO Chamber) developed in Japan. Research design and methods: A total of 39 patients with cedar pollinosis were targeted. The subjects were exposed to a specific amount of cedar pollen (8000/m3) in the OHIO Chamber during the non-cedar pollen season. Efficacy of pranlukast for the treatment of artificially induced nasal symptoms was compared with that of a placebo using the crossover method. Pranlukast was administered orally for 3 days, after dinner on the day before cedar pollen exposure, after breakfast and after dinner on the day of cedar pollen exposure, and after breakfast on the following day. Pollen testing was carried out twice, with a 1-week wash-out interval. Clinical trial registration: The University Hospital Medical Information Network in Japan (UMIN), number UMIN000001282. Main outcome measures: The effect of pranlukast was evaluated using self-rating of nasal symptoms by the subjects. Results: All 39 subjects demonstrated a positive skin reaction to cedar pollen by a positive CAP-RAST score (class 2 or higher) within the last 3 years, and experienced aggravated congestion during the cedar pollen season for more than 2 years. Nasal congestion was inhibited significantly in the pranlukast group compared to the placebo group during cedar pollen exposure. Furthermore, pranlukast significantly inhibited nasal congestion compared to the placebo on the day after exposure and on the following day. Conclusions: The effect of pranlukast on cedar pollinosis indicates immediate action, and such an effect could take place continuously after cedar pollen exposure. These results demonstrate that pranlukast is effective for the relief of congestion due to pollinosis.

Immune Regulation by CD4+CD25+ Regulatory T Cells in Patients with Japanese Cedar Pollinosis

Background: Evidence indicating that CD4+CD25+ regulatory T (Treg) cells play a crucial role in the maintenance of peripheral T cell tolerance to allergens has been accumulated. To explore the functional role of Treg cells in patients with Japanese cedar pollinosis, we performed an in vitro investigation of the regulation of immune responses to allergens by Treg cells. Methods: CD4+ and CD4+CD25– T cells obtained from 12 patients with Japanese cedar pollinosis were stimulated with Cry j 1 protein and Cry j 1-derived peptide. On day 6, T cells were tested for allergen-specific reactivity using a CFSE-based proliferation assay and cytokine ELISA assays. The frequency of Cry j 1-specific interleukin (IL)-10-producing Treg cells was assessed by ELISPOT assays. Results: The proportion of proliferated cells induced by allergen stimulation was similar in both CD4+ and CD4+CD25– cell cultures. The production of interferon (IFN)-γ, but not that of IL-5 was significantly enhanced in CD4+CD25– cell cultures compared to that in CD4+ cell cultures. Interestingly, the production of IL-10 was decreased in CD4+CD25– cell cultures. Moreover, Cry j 1-specific IL-10-producing Treg cells were detected in pollen-allergic patients. Conclusion: Our findings suggest that in pollen-allergic patients, Treg cells predominantly suppresses Th1 responses rather than Th2 responses, where allergen-specific IL-10-producing Treg cells may also be responsible for the downregulation of allergen-specific immune responses.

Expression of cysLT1 and cysLT2 receptor in chronic hyperplastic eosinophilic sinusitis.

Elevated production of cysteinyl leukotrienes (cysLTs) from sinus tissues and abundant sinus eosinophils are characteristic features of chronic hyperplastic eosinophilic sinusitis (CHS). CysLTs exert their action through G-protein-coupled receptors named cysLTs receptor type I (cysLT1R) and type II (cysLT2R). These expressions of cysLT receptors in the sinus mucosa have yet to be clarified and the relationship between eosinophilia and the expression of these receptors remains obscure. We compared the expressions of cysLT1R and cysLT2R in the sinus mucosa in patients with CHS, non-eosinophilic chronic sinusitis (NECS), and control sinus tissues; and analyzed the correlation between the expression of CysLTRs and the presence of sinus eosinophils by immunohistochemistry and real-time PCR. A significantly higher percentage of eosinophils expressing cysLT2R protein was observed in patients with CHS compared with NECS and controls. In addition, cysLT2R mRNA expression in CHS was significantly higher than in NECS and controls. Furthermore, a positive correlation was observed between cysLT2R mRNA expression and the number of infiltrated eosinophils. In contrast, the cysLT1R mRNA expression did not differ significantly among these groups. The effect of cysLTs on sinus eosinophils may be mediated through the cysLT2R in patients with CHS. These results may suggest the therapeutic benefit of cysLT2R antagonists in CHS.

Analysis of T-helper Responses and FOXP3 Gene Expression in Patients with Japanese Cedar Pollinosis:

Background Evidence has been accumulated indicating that regulatory T (T-reg) cells play a crucial role in the maintenance of peripheral T-cell tolerance to allergens. To explore the role of FOXP3, which is required for the development of T-reg cells, in allergen-specific immune responses, we examined the relationship between the alteration of FOXP3 gene expression and in vitro immune responses against allergens. Methods Peripheral blood mononuclear cells obtained from 19 human histocompatibility leukocyte antigens (HLA)-DPB1*0501 donors, including patients with Japanese cedar pollinosis and nonallergic healthy donors, were stimulated with Cry j 1 p61-75 peptide. On day 7, T cells were tested for peptide-specific reactivity in IFN-γ and interleukin (IL)-5 enzyme-linked immunospot (ELISPOT) assays. Real-time quantitative RT-PCR was performed to assess relative change of FOXP3 gene expression before and after in vitro stimulation. Neutralization assays using anti-glucocorticoid-induced tumor necrosis factor receptor family-related protein (GITR) and anti-IL-10 monoclonal antibody were also performed. Results Of 14 patients with allergic pollinosis tested, 10 responders displayed T-helper type 2 (Th2)-polarized reactivity to Cry j 1 p61-75, and 2 donors showed Th0 responses. Notably, the change of FOXP3 gene expression in donors showing peptide-specific T-helper responses was significantly lower than that in nonresponders, regardless of allergic pollinosis. Conclusion Our data indicate that FOXP3 is functional in nonallergic healthy donors as well as allergic patients, and FOXP3-expressing T cells may be responsible for the down-regulation of allergen-specific T-helper responses in individuals. A better understanding of the nature and specificity of FOXP3-expressing T cells in a suppressive mechanism is necessary to develop new immunotherapies against allergic rhinitis.

Proposed method for assessing audible hazard alarms using simulated hearing impairment

The need for sound design schemes for alarm systems that can be easily heard by both people with hearing impairments and with those with normal hearing has been expressed from many quarters. While sound designs for various types of hazard alarm systems, different classifications of hearing impairments, and even ambient sound are required, no solutions have appeared thus far. This paper proposes to fill this need by using simulated impaired hearing based on a sound design assessment scheme applying the Amplitude bandwidth Compression/Expansion method (ACE) for modeling the characteristics of nonlinear aural fields. By having people with normal hearing listen to hazard alarm systems that employ this simulated hearing impairment, they can re-create the experience of people with hearing impairments, enabling them to identify the necessary aural elements of existing hazard alarm systems (car horns/alarms, railroad crossing warning sounds). This methodology proved to be effective for universal sound design.