Yoshitaka Tanaka

Critical role for OX40 ligand in the development of pathogenic Th2 cells in a murine model of asthma

Bronchial asthma is characterized by massive infiltration of eosinophils and airway hyperreactivity (AHR), which are caused by overproduction of Th2 cytokines (IL‐4, IL‐5, and IL‐13) by allergen‐specific T cells. We recently demonstrated a critical contribution of OX40 ligand (OX40L) to the development of Th2‐mediated experimental leishmaniasis. In this study, we have examined the role of OX40L in the development of Th2‐mediated pulmonary inflammation by utilizing OX40L‐deficient mice and a neutralizing anti‐OX40L mAb in a murine model of asthma. Sensitization and airway challenge with ovalbumin in wild‐type BALB/c mice induced a typical allergic asthma characterized by AHR, accumulation of eosinophils, increased mucus production, and high levels of Th2 cytokines in the lung.All these asthmatic responses were not induced in OX40L‐deficient BALB/c mice. Administration of neutralizing anti‐OX40L mAb in wild‐type BALB/c mice during the sensitization period also abolished the induction of asthmatic responses. In contrast, administration of anti‐OX40L mAb during the challenge period did not inhibit the asthmatic responses. These results indicate a critical role for OX40L in the induction phase, which leads to the development of pathogenic Th2 cells, but not in the effector phase, which includes migration and activation of pathogenic Th2 cells in the lung.

Murine NKT cells produce Th17 cytokine interleukin-22

Natural killer T (NKT) cells are known to produce Th17 cytokine IL-17 in addition to Th1/2 cytokines. In this study, the ability of NKT cells to produce IL-22, another Th17 cytokine, was examined in mice. When murine spleen cells were stimulated with alpha-galactosyl ceramide, a ligand for NKT cells, not only Th1/2 cytokines (IFN-gamma, IL-4) but Th17 cytokines (IL-17, IL-22) were produced. NKT cells isolated from splenocytes released IL-17 and IL-22 following CD3, CD3/IL-2 or CD3/CD28 stimulation, in which CD3/CD28 costimulation was most effective. Production of IL-17 and IL-22 in CD4+ and CD8+ T cells from splenocytes was little, if any, even after CD3/CD28 costimulation. Treatment with IL-6/TGF-beta decreased CD3/CD28-stimulated production of IL-22, but not that of IL-17, in NKT cells. These findings show for the first time that NKT cells are a cell source of IL-22, and that expression of two Th17 cytokines might be regulated in NKT cells by different mechanisms.

A Critical Role for Mouse CXC Chemokine(s) in Pulmonary Neutrophilia During Th Type 1-Dependent Airway Inflammation

Ag-specific Th1 and Th2 cells have been demonstrated to play a critical role in the induction of allergic diseases. Here we have investigated the precise mechanisms of Th1-induced airway inflammation. Airway inflammation was induced in BALB/c mice by transfer of freshly induced OVA-specific Th1 or Th2 cells followed by OVA inhalation. In this model, both Th1 and Th2 cells induced airway inflammation. The former induced neutrophilia in airways, whereas the latter induced eosinophilia. Moreover, we found that Th1 cells induced more severe airway hyperresponsiveness (AHR) than Th2 cells. The eosinophilia induced by Th2 cell infusion was almost completely blocked by administration of anti-IL-5 mAb, but not anti-IL-4 mAb. In contrast, Th1-induced AHR and pulmonary neutrophilia were inhibited by the administration of anti-human IL-8R Ab, which blocks the function of mouse CXC chemokine(s). These findings reveal a critical role of mouse CXC chemokine(s) in Th1-dependent pulmonary neutrophilia and AHR.

ASB16165, a novel inhibitor for phosphodiesterase 7A (PDE7A), suppresses IL-12-induced IFN-γ production by mouse activated T lymphocytes

Phosphodiesterase 7A (PDE7A) has been suggested to be involved in activation of T lymphocytes. In the present study, a possible involvement of PDE7A in function of preactivated T cells (i.e. T lymphoblasts) was investigated using ASB16165, an inhibitor for PDE7A. ASB16165, which has an IC50 value of 15 nM for human PDE7A, suppressed IL-12-induced IFN-gamma production by T lymphoblasts which have been prepared by stimulating mouse T cells with anti-CD3 antibody. In the same experiment, rolipram, a PDE4-specific inhibitor, showed similar effect, while calcineurin antagonist FK506 did not. Forskolin (an adenylyl cyclase activator) and dibutyryl cAMP also inhibited the IL-12-induced IFN-gamma synthesis. Rp-8-Br-cAMPS, an inhibitor of protein kinase A (PKA), reduced the suppressive effect of ASB16165 on the IFN-gamma production by T lymphoblasts. The rescue of IFN-gamma production by Rp-8-Br-cAMPS was also observed in the inhibition by rolipram and forskolin. These findings suggest that PDE7A may regulate function of activated T cells in a cAMP/PKA-dependent manner, and that PDE4 might share the role. The data in our study also indicate that PDE7 inhibitors such as ASB16165 will be beneficial for the patients with immunological disorders.

Effect of phosphodiesterase 7 inhibitor ASB16165 on development and function of cytotoxic T lymphocyte

In the present study, possible role of phosphodiesterase 7 (PDE7) in development and function of cytotoxic T lymphocyte (CTL) was examined using ASB16165, a specific inhibitor for PDE7. ASB16165 inhibited generation of CTL activity in mixed lymphocyte reaction (MLR), in which splenocytes from C57BL/6N mice were stimulated with those from BALB/c mice. Flow cytometric analysis revealed that ASB16165 suppressed induction of activated CD4+ as well as CD8+ T cells in MLR. In cell division analyses using 5-carboxyfluorescein diacetate succinimide ester (CFSE), ASB16165 was shown to markedly inhibit proliferation of CD4+ and CD8+ T cells. In addition, ASB16165 reduced effector function of CTL, while the effect was less than that observed in CTL induction in MLR. Forskolin and dibutyryl cAMP also inhibited both the induction and effector function of CTL. PDE4 inhibitor rolipram showed similar but weaker inhibition for the development and proliferation of CD8+ T cells compared with ASB16165, and failed to impair effector function of CTL. These findings suggest that PDE7 but not PDE4 has the major role in induction and function of CTL in mice, and that the effect might be mediated by elevation of intracellular cAMP level. ASB16165 may be useful for treatment of the diseases in which CTL has a pathogenic role (e.g. autoimmune diseases).

Establishment of a T cell-dependent nude mouse liver injury model induced by Propionibacterium acnes and LPS.

Normal ICR mice developed severe liver injury when they were given intravenous injections of Propionibacterium acnes and lipopolysaccharide (LPS) with a 7 day interval. In contrast, T cell-deficient ICR nude mice were resistant to P. acnes and LPS-induced liver injury. However, athymic ICR nude mice, which were treated with cell transfer of normal ICR mouse spleen cells (10(8) cells) or ICR mouse nylon-wool passed splenic T-enriched cells (over 10(7) cells), showed severe liver injury as assessed by elevation of serum transaminase activities. Histological analyses also demonstrated that the transferred cells migrated into the liver of nude mice to induce liver injury. However, depletion of both CD4+ T cells and CD8+ T cells from transferred cell populations caused a marked decrease in the elevation of serum transaminase, indicating the actual involvement of T cells in liver injury. Moreover, in vivo administration of anti-LFA-1 mAb blocked P. acnes and LPS-induced liver injury in nude mice following T cell transfer. Thus, this model will provide a new strategy to investigate T cell-dependent cell-cell interaction during the induction of liver damage.