Inhibitory function of NKT cells during early induction phase of nickel allergy.

Until now, metal allergies have been regarded as a Th1-type immune response.　However, because the contribution of a Th2-type immune response has been suggested by clinical findings, we previously examined the Th2-type immune response during the development of metal allergies using a GATA-3 transgenic (GATA-3 Tg) mouse model. As a result, a Th2-type immunization reaction was suggested to be involved in the early phase of metal allergies. Recently, the involvement of NKT cells in metal allergies has been suggested. We examined this possibility using the activation of NKT cells and an NKT cell-deficient mouse model to determine the contribution of NKT cells to nickel allergy in the present study. In NKT cell-deficient mice, ear swelling was remarkably increased, compared with that in control mice. Also, in mice that had been treated with α-galactosylceramide (α-GalCer) to activate NKT cells, the ear swelling response was remarkably inhibited, compared with that in untreated mice. These facts show that NKT cells are involved in the inhibition of nickel allergy-induced ear swelling responses.

Release from Th1-type immune tolerance in spleen and enhanced production of IL-5 in Peyer's patch by cholera toxin B induce the glomerular deposition of IgA.

We examined the pathogenesis of glomerular damage in Th2 type-dependent GATA-3 transgenic (GATA-3 Tg) mice with IgA nephropathy (IgAN). GATA-3 Tg mice were immunized orally using OVA plus cholera toxin B (CTB), and measurement of the serum IgA antibody level and histopathological examination were performed. Marked increases in the serum levels of OVA-specific IgA antibody, IgA and IgG, C3 deposits analogous to those seen in IgAN, and expansion of the matrix in association with mesangial cell proliferation were observed. Furthermore, glomerular IgA deposits were co-localized with mannan-binding lectin (MBL) deposits, which might actually have been abnormal IgA deposits. In GATA-3/TCR-Tg mice that had been orally sensitized with CTB plus OVA and were re-stimulated with OVA in vitro, cultured Peyer's patch cells showed the enhanced production of IL-5 and supernatants from cultures of spleen cells showed a reduction of TGF-ß production with a simultaneous increase in IL-2 production and the recovery of IFN-γ formation. The amount of TGF-ß produced by the spleen cells was found to be correlated with the amount of IFN-γ and IL-IL-2 produced by the cells. Also, the percentage of regulatory T cells (Treg) in the spleens of mice sensitized with OVA plus CTB was lower than that in mice orally sensitized with OVA alone. These results suggest that the increased production of IL-5 from Peyer's patch cells (PPc) and the restored Th1-type immune response might cause the production of abnormal IgA and might induce the deposition of IgA in glomeruli.

GATA-3 regulates contact hyperresponsiveness in a murine model of allergic dermatitis.

Contact hypersensitivity (CHS) is thought to be associated mainly with the activation of T helper (Th) type 1 cells. However, evidence also suggests that Th type 2 cells (Th2) and cytokines play roles in the development of CHS in humans. The present study examines the Th2 response during the development of CHS in response to 2,4,6-trinitrochlorobenzene (TNCB) in GATA-3-transgenic (GATA-3 Tg) mice. GATA-3 Tg mice were immunized with 7% TNCB applied to abdominal shaved skin. Seven days later, the mice were challenged with 1% TNCB applied to the left ear. Ear swelling, cytokine production in the skin of the ear, and the levels of IgE, IgG1 and IgG2a were measured. Furthermore, we examined the effects of medical treatment on TNCB-induced contact dermatitis using this model. The ear-swelling responses of TNCB-sensitized/challenged GATA-3 Tg mice were significantly greater than those of similarly treated wild-type (WT) mice. The expression of both IL-5 and IL-13 in TNCB sensitized/challenged skin tissues and the IgE response after challenge were obviously increased in the GATA-3 Tg mice, whereas the expression of IFN-γ was identical in the challenged skin tissues of GATA-3-Tg and WT mice. When TNCB-sensitized GATA-3 Tg mice were treated with a high dose of tacrolimus, ear swelling was not significantly decreased, compared with the results in WT mice. These results suggest that GATA-3-induced Th2-dominant responses play a critical role in the pathogenesis of allergic types of dermatitis, such as atopic dermatitis, and may lead to useful new drug development in the future.

IL-10 controls Th2-type cytokine production and eosinophil infiltration in a mouse model of allergic airway inflammation.

Interleukin-10 was originally described as a factor that inhibits cytokine production by murine Th1 clones. Recent studies have since shown that IL-10 can also downregulate Th2 clones and their production of IL-4 and IL-5. Because of its immuno-suppressive properties, IL-10 has been suggested as a potential therapy for allergic inflammation and asthma. However, the pathophysiological role of IL-10 in vivo has not been clearly elucidated. We investigated the effects of IL-10 administration on the production of IgE, cytokine and allergen-induced Th2 cytokine production as well as its effects on eosinophilic inflammation. We established GATA-3/TCR double transgenic (GATA-3/TCR-Tg) mice by crossing GATA-3 transgenic mice with ovalbumin (OVA)-specific TCR transgenic mice; these mice were then sensitized using an intraperitoneal injection of OVA adsorbed to alum and challenged with the intratracheal instillation of an allergen. When GATA-3/TCR-Tg mice sensitized with OVA and alum were injected with C57-IL-10 cells before OVA inhalation, the levels of IL-5, IL-13, and IL-4 decreased by 40-85% and number of eosinophils decreased by 70% (P<0.03) in the murine bronchoalveolar lavage fluid (BALF). These results suggest that IL-10 plays an important role downstream of the inflammatory cascade in the Th2 response to antigens and in the development of BALF eosinophilia and cytokine production in a murine model of asthma. These immunosuppressive properties in animal models indicate that IL-10 could be a potential clinical therapy for the treatment of allergic inflammation.

Th2 immune response plays a critical role in the development of nickel-induced allergic contact dermatitis.

BACKGROUND: The precise roles of T helper (Th)1-type and Th2-type cytokine responses in nickel (Ni)-induced allergic contact dermatitis have not yet been clearly defined. We investigated the involvement of Th2 cytokines in Ni-induced contact hypersensitivity reaction using GATA-3 transgenic (Tg) mice. METHODS: A Ni-titanium (Ti) alloy was implanted under the skin of GATA-3 Tg mice. A Ni solution was then injected 1 month after sensitization. The ear swelling response was measured at several time points after the injection; the cytokine levels in the skin were measured at 48 h after injection, and the serum levels of IgE were measured 1 month after injection. In addition, purified CD4+ splenic cells obtained from the GATA-3 Tg mice sensitized with the Ni-Ti alloy were infused into Rag-2(-/-) mice, and the ear swelling response of these mice after a further challenge with Ni solution was also measured. RESULTS: Marked ear swelling and elevated serum IgE levels and skin tissue levels of IL-4 were observed in Ni-Ti-sensitized GATA-3 Tg mice. The Rag-2(-/-) mice transfused with the CD4+ splenic cells from the Ni-Ti alloy sensitized GATA-3 Tg mice showed a significantly more pronounced ear swelling response than the control mice. CONCLUSION: We confirmed the participation of Th2-type immune reactions in Ni-induced allergy using GATA-3 Tg mice.

Involvement of GATA-3-dependent Th2 lymphocyte activation in airway hyperresponsiveness.

The pathophysiological characteristics of bronchial asthma consist of chronic inflammation of airways, airway hyperresponsiveness, and bronchoconstriction. Studies have shown that T helper type 2 (Th2) cytokines produced by both T cells and mast cells in the airway contribute substantially to the initiation of inflammation in both experimental and human bronchial asthma. GATA-3 is a transcription factor essential to the production of Th2 cytokines by T lymphocytes. To clarify the role of GATA-3-expressing T cells in the pathophysiology of bronchial asthma, we utilized transgenic (Tg) mice carrying the GATA-3 gene and the ovalbumin (OVA)-specific T cell receptor gene (GATA-3-Tg/OVA-Tg). Mice were intranasally administrated OVA without systemic immunization. Airway responses were analyzed with noninvasive and invasive whole body plethysmographs. GATA-3-Tg/OVA-Tg mice exhibited significantly higher IL-13 and IL-4 protein expression in the airway. Although there were no differences in the types of infiltrating cells between GATA-3-Tg/OVA-Tg and GATA-3-non-Tg/OVA-Tg mice and no significant increase in IgE level in either group compared with nontreated mice, the response after ACh inhalation was significantly elevated in GATA-3-Tg/OVA-Tg on the seventh day of intranasal treatment with OVA. This hyperresponsiveness was inhibited by 5-lipoxygenase inhibitor and IL-13 neutralization, suggesting that airway responses were induced through IL-13 and leukotriene pathway. In conclusion, airway hyperresponsiveness, a characteristic of bronchial asthma, is regulated at the level of GATA-3 transcription by T lymphocytes in vivo.

Immune responses to Nippostrongylus brasiliensis and tuberculin protein in GATA-3-transgenic mice.

GATA-3 appears to be key to the Th2 response. However, few in vivo experiments have examined the function of GATA-3 in Th1 and Th2 immune responses. We developed two lines of GATA-3-transgenic (Tg) mice harboring the SRalpha or lck promoters and examined the Th2 immune responses of mice infected with the intestinal nematode Nippostrongylus brasiliensis and the Th1 responses with purified derivative of tuberculin (PPD) immunization. Numbers of peripheral blood eosinophils in all GATA-3-Tg mice increased 10- to 20-fold after primary infection with N. brasiliensis and 25-100-fold after secondary infection. The number of eosinophils in infected GATA-3-Tg mice was significantly higher than that in infected control littermates. Total IgE levels after primary infection in GATA-3-Tg mice were 8-450-fold increased, which was significantly higher than those of control mice. Mesenteric lymph node cells of infected GATA-3-Tg mice upon stimulation with N. brasiliensis antigen secreted more IL-5 and IL-13 than those of control mice. However, production of IL-4 and IFN-gamma were comparable between GATA-3-Tg and controls. Mice immunized with PPD were intradermally challenged with PPD to induce delayed type hypersensitivity (DTH). The amount of footpad swelling caused by the DTH reaction in GATA-3-Tg mice was significantly smaller than that of control littermates. Inguinal lymph node cells from GATA-3-Tg mice stimulated with PPD in vitro secreted more IL-5, IL-10 and less IFN-gamma than those of control littermates. These results suggested that Th1 and Th2 driven conditions enhance IL-5 production in GATA-3-Tg mice through the direct binding of GATA-3 to the IL-5 promoter region. The influence of GATA-3 on IL-13, IFN-gamma and IL-10 production varied according to the stimulating conditions. However, IL-4 production was not significantly elevated in GATA-3-Tg mice, indicating that IL-4 and IL-5 production was differentially regulated in these mice.

Evidence of GATA-3-dependent Th2 commitment during the in vivo immune response.

The transcription factor GATA-3 has been shown to play an important role for the in vitro induction of T(h)2 cells. To clarify how the in vivo immune response is governed under GATA-3 function, we generated double-transgenic mice by crossing GATA-3 transgenic mice with ovalbumin (OVA)-specific TCR transgenic mice. After immunization with OVA, the double-transgenic mice showed increased expression of GATA-3 in antigen-reactive fresh CD4(+) T cells, and higher production of IL-5 and IL-13 in cultured spleen cells in the presence of cognate antigen without any polarizing conditions for T(h)2 cells. Moreover, the immunized double-transgenic mice showed a higher increase of in vivo secretion of IL-5 and IL-13 in bronchoalveolar lavage fluid after OVA aerosol challenging. The serum levels of OVA-specific IgG1, IgE and IgA antibodies were much higher in the immunized double-transgenic mice than TCR transgenic mice. These findings provide direct evidence that antigen-stimulated CD4(+) T cells in the immunized mice have already been committed into T(h)2 cells producing IL-5 and IL-13 selectively through enhanced GATA-3 expression in vivo, thereby inducing higher production of antigen-specific antibody for three isotypes other than IgM.