Alteration of Immune Responses by N-acetylglucosaminyltransferase V during Allergic Airway Inflammation

Background: p-1,6-N-acetylglucosaminyltransferase V (Mgat5 or GlcNac-TV), which is involved in the glyco- sylation of proteins, is known to be important for down-regulation of TCR-mediated T-cell activation and negatively regulates induction of contact dermatitis and experimental autoimmune encephalomyelitis. However, the role of Mgat5 in the induction of allergic airway inflammation remains unclear. Methods: To elucidate the role of Mgat5 in the pathogenesis of allergic airway inflammation, ovalbumin (OVA)-induced airway inflammation was induced in Mgat5-deficient mice. The OVA-specific lymphocyte proliferation and cytokine production levels, OVA-specific IgG1, IgG2a and IgE levels in the serum, and the number of leukocytes and cytokine levels in the bronchoalveolar lavage (BAL) fluid were compared between wild-type and Mgat5-deficient mice. Results: OVA-specific lymphocyte proliferation and production of IFN-y and IL-10, but not IL-4, were increased in Mgat5-deficient mice, suggesting that Th2-type immune responses are seemed to be suppressed by increased IFN-y and IL-10 production in these mice. However, Th2-type responses such as OVA-specific IgG1, but not IgE, and IL-5 levels in BAL fluids were increased in Mgat5-deficient mice. Meanwhile, the number of eosinophils was normal, but the numbers of neutrophils, macrophages and lymphocytes were reduced, in these mutant mice during OVA-induced airway inflammation. Conclusions: Mgat5-dependent glycosylation of proteins can modulate acquired immune responses, but it is not essential for the development of OVA-induced eosinophilic airway inflammation

IL-33, but not IL-25, is crucial for the development of house dust mite antigen-induced allergic rhinitis.

Both interleukin (IL)-33 and IL-25 induce Th2 cytokine production by various cell types, suggesting that they contribute to development of allergic disorders. However, the precise roles of IL-33 and IL-25 in house dust mite (HDM)-induced allergic rhinitis (AR) remain unclear. Both IL-33 and IL-25 were produced mainly by nasal epithelial cells during HDM-induced AR. Eosinophil and goblet cell counts in the nose and IL-5 levels in lymph node cell culture supernatants were significantly decreased in IL-33-deficient, but not IL-25-deficient, mice compared with wild-type mice during HDM-induced AR, but the serum IgE and IgG1 levels did not differ. On the other hand, HDM-induced AR developed similarly in wild-type mice transferred with either IL-33-deficient BM cells or wild-type BM cells. IL-33, but not IL-25, produced by nasal epithelial cells was crucial for the development of murine HDM-induced AR. These observations suggest that IL-33 neutralization may be a potential approach for treatment of HDM-induced AR in humans

TIM-3 is not essential for development of airway inflammation induced by house dust mite antigens

Background: T cell immunoglobulin domain and mucin domain-containing molecule 3 (TIM-3), which is preferentially expressed on Th1 cells rather than Th2 cells, is considered to be a negative regulator of Th1 cell function. This suggests that TIM-3 indirectly enhances Th2-type immune responses by suppressing Th1 cell function. Methods: To investigate TIM-3's possible involvement in Th2-type acute and chronic airway inflammation, wild-type and TIM-3-deficient (TIM-3−/−) mice were sensitized and challenged with a house dust mite (HDM) extract. Airway inflammation and the number of inflammatory cells in bronchoalveolar lavage fluids (BALFs) in the mice were determined by histological analysis and with a hemocytometer, respectively. Expression of mRNA in the lungs was determined by quantitative PCR, while the levels of cytokines in the BALFs and IgE in sera were determined by ELISA. Results: Despite constitutive expression of TIM-3 mRNA in the lungs, the number of eosinophils in bronchoalveolar lavage fluids (BALFs) and the score of pulmonary inflammation were comparable between wild-type and TIM-3−/− mice during both acute and chronic HDM-induced airway inflammation. On the other hand, the number of lymphocytes in the BALFs of TIM-3−/− mice was significantly increased compared with wild-type mice during HDM-induced chronic, but not acute, airway inflammation, while the levels of Th2 cytokines in the BALFs and HDM-specific IgG1 and IgG2a and total IgE in the sera were comparable in both groups. Conclusions: Our findings indicate that, in mice, TIM-3 is not essential for development of HDM-induced acute or chronic allergic airway inflammation, although it appears to be involved in reduced lymphocyte recruitment during HDM-induced chronic allergic airway inflammation

Fundamental Characterization of Antibody Fusion-Single-Chain TNF Recombinant Proteins Directed against Costimulatory TNF Receptors Expressed by T-Lymphocytes

The costimulatory signal regulated by the members of the tumor necrosis factor receptor (TNFR) superfamily expressed by T cells plays essential roles for T cell responses and has emerged as a promising target for cancer immunotherapy. However, it is unclear how the difference in TNFR costimulation contributes to T cell responses. In this study, to clarify the functional significance of four different TNFRs, OX40, 4-1BB, CD27 and GITR, we prepared corresponding single-chain TNF ligand proteins (scTNFLs) connected to IgG Fc domain with beneficial characteristics, i.e., Fc−scOX40L, Fc−sc4-1BBL, Fc−scCD27L (CD70) and Fc−scGITRL. Without intentional cross-linking, these soluble Fc−scTNFL proteins bound to corresponding TNFRs induced NF-kB signaling and promoted proliferative and cytokine responses in CD4+ and CD8+ T cells with different dose-dependencies in vitro. Mice injected with one of the Fc−scTNFL proteins displayed significantly augmented delayed-type hypersensitivity responses, showing in vivo activity. The results demonstrate that each individual Fc−scTNFL protein provides a critical costimulatory signal and exhibits quantitatively distinct activity toward T cells. Our findings provide important insights into the TNFR costimulation that would be valuable for investigators conducting basic research in cancer immunology and also have implications for T cell-mediated immune regulation by designer TNFL proteins

IL-25, IL-33 and TSLP receptor are not critical for development of experimental murine malaria

IL-25, IL-33 and TSLP, which are produced predominantly by epithelial cells, can induce production of Th2-type cytokines such as IL-4, IL-5 and/or IL-13 by various types of cells, suggesting their involvement in induction of Th2-type cytokine-associated immune responses. It is known that Th2-type cytokines contribute to host defense against malaria parasite infection in mice. However, the roles of IL-25, IL-33 and TSLP in malaria parasite infection remain unclear. Thus, to elucidate this, we infected wild-type, IL-25−/−, IL-33−/− and TSLP receptor (TSLPR)−/− mice with Plasmodium berghei (P. berghei) ANKA, a murine malaria strain. The expression levels of IL-25, IL-33 and TSLP mRNA were changed in the brain, liver, lung and spleen of wild-type mice after infection, suggesting that these cytokines are involved in host defense against P. berghei ANKA. However, the incidence of parasitemia and survival in the mutant mice were comparable to in the wild-type mice. These findings indicate that IL-25, IL-33 and TSLP are not critical for host defense against P. berghei ANKA

IL-33, but not IL-25, is required for IL-5 production, but not Th2 cell differentiation, during HDM-induced AR in mice.

<p>(A, B) HDM-specific LN cell proliferative responses and cytokine production of wild-type and IL-25^−/− mice (A) and of wild-type and IL-33^−/− mice (B) 48 hours after the last inhalation of HDM or PBS. Data show the mean + SEM (PBS, n = 3–4; HDM, n = 6–7), and the results generated in one of two independent experiments, each of which gave similar results. * P<0.05. (C–F) The proportion of cytokine-producing CD3⁺ CD4⁺ T cells in cervical LN cells cultured with and without HDM <i>in vitro</i>, as in (A) and (B). (C, E) FACS data show the results for LN cells pooled from 5 mice in each group, and a representative result of 5 independent experiments. (D, F) Data show the mean + SEM of 5 independent experiments in (C) and (E). Med =  Medium alone.</p

IL-33 produced by immune cells derived from bone marrow stem cells is not crucial for development of HDM-induced AR.

<p>(A) H&E staining of nasal mucosa, (B) the number of eosinophils in the nasal mucosa and (C) cytokine levels in the culture supernatants of HDM-stimulated LN cells from wild-type mice injected with wild-type bone marrow (BM) cells or IL-33^−/− BM cells, 48 hours after the last inhalation of HDM (n = 10) or PBS (n = 5). Histological data show a representative result from each group (bar  = 20 µm). Data show the mean + SEM. Med =  Medium alone.</p

Histological analysis for infiltration of leukocytes and hyperplasia of goblet cells.

<p>Histological analysis of nasal mucosa (maxillary turbinate) from wild-type, IL-25^−/− and IL-33^−/− mice 48 hours after the last inhalation of HDM or PBS. (A) H&E staining. After inhalation of HDM, but not PBS, eosinophils were observed in the nasal mucosa of wild-type and IL-25^−/− mice but hardly detectable in mucosa from IL-33^−/− mice. Arrowheads indicate eosinophils. (B) PAS staining. After inhalation of HDM, but not PBS, goblet cell hyperplasia was similarly observed in the nasal mucosa of both wild-type and IL-25^−/− mice, but it was markedly reduced in IL-33^−/− mice compared with wild-type mice. Arrowheads indicate PAS-positive goblet cells. Data show a representative result from 3–7 mice in each group (bar  = 20 µm).</p

IL-25 and IL-33 may somehow contribute to the development of HDM-induced AR.

<p>(A) Scheme of intranasal treatment of mice with HDM or PBS. (B) Detection of IL-25 in nose of wild-type and IL-25^−/− mice treated with HDM or PBS on Day 30 as in (A). The sections were stained with anti-mouse IL-25 Ab. IL-25 was detected in the cytoplasm of epithelial cells (brown) from HDM-treated wild-type mice, but not PBS-treated wild-type mice or PBS- and HDM-treated IL-25^−/− mice. Arrowheads indicate IL-25-positive epithelial cells (×400). (C) Detection of IL-33 in nose from wild-type and IL-33^−/− mice treated with HDM or PBS on Day 30, as in (A). The sections were stained with anti-mouse IL-33 Ab. IL-33 was detected in the nuclei of epithelial cells (red) from PBS-treated wild-type mice, but not HDM-treated wild-type mice or PBS- and HDM-treated IL-25^−/− mice. Arrowheads indicate IL-33-positive epithelial cells. Red shows IL-33 staining, and blue shows DAPI staining (×400). Data show a representative result from 3–7 mice in each group.</p