Intranasal ovalbumin immunotherapy with mycobacterial adjuvant promotes regulatory T cell accumulation in lung tissues

Allergen‐specific immunotherapy to induce T regulatory cells in the periphery has been used to treat allergic diseases. Mycobacteria can be used as an adjuvant for inducing T regulatory cells. However, it is unclear whether intranasal immunotherapy in combination with Mycobacteria adjuvant induces regulatory T cell differentiation and attenuates allergic responses in vivo. To investigate the role of intranasal ovalbumin (OVA) treatment alone and in combination with Mycobacteria vaccae, proportions of FoxP3⁺ regulatory T cells and anti‐inflammatory responses were evaluated in a murine model of asthma that was established in three groups of bicistronic Foxp3^EGFP reporter BALB/c mice. Before establishment of the asthma model, two groups of mice received intranasal OVA immunotherapy and one also received simultaneous s.c. M. vaccae. Expression of CD4⁺CD25⁺Foxp3^+EGFP+ T cells in the lung and spleen was analyzed by flow cytometry and the cytokine profiles of allergen‐stimulated lung and spleen lymphocytes assessed. The intranasal OVA immunotherapy group showed greater expression of CD4⁺CD25⁺Foxp3^+EGFP+ T cells in the spleen whereas in the group that also received M. vaccae such greater expression was demonstrated in the lung. Additionally, the proportion of IL‐10 and IFN‐γ‐secreting splenocytes was greater in the intranasal OVA + M. vaccae group. CD25 neutralization decreased CD4⁺Foxp3⁺ cells more than other groups. In parallel with this finding, production of IL‐10 and IFN‐γ was down‐regulated. Mucosal administration of OVA antigen results in a greater proportion of CD4⁺Foxp3⁺ T cells in the spleen. IL‐10 and IFN‐γ induced by intranasal OVA immunotherapy and M. vaccae administration is down‐regulated after CD25 neutralization.

     

EPO modified MSCs can inhibit asthmatic airway remodeling in an animal model

There was no effective measures can be obtained at present to reverse or prevent airway remodeling. We investigated the therapeutic effect of Erythropoietin (EPO) gene modified mesenchymal stem cells (MSCs) on asthmatic airway remodeling and the possible underlied molecular mechanisms. EPO gene was transfected into MSCs via lentivirus vector. The transfected cells (EPO‐MSCs) were identified by flow cytometry and the EPO secreting function was detected by PCR and Western blot. MSCs or EPO‐MSCs were administrated to albumin (OVA)‐induced chronic asthmatic mouse model via tail veins. The asthmatic phenotype was analyzed. Number of cells in bronchoalveolar lavage fluid (BALF) was counted using a hemocytometer. Histological findings of airways were evaluated by microscopic examination. The concentrations of interleukin 4(IL‐4), interleukin 5(IL‐5), and interleukin 13(IL‐13) in lung homogenate were determined by ELISA. The activation state of transforming growth factor‐β 1 (TGF‐β1), Transforming growth factor beta‐activated kinase 1 (TAK1), and p38 Mitogen Activated Protein Kinase (p38MAPK) signaling was detected by Real‐Time PCR and Western blotting. EPO‐MSCs were successfully constructed. EPO‐MSCs showed a more potently suppressive effect on local asthmatic airway inflammation and the level of IL‐4, IL‐5, and IL‐13 in lung tissue than MSCs. Moreover, the numbers of goblet cells, the thicknesses of smooth muscle layer, collagen density, percentage of proliferating cell nuclear antigen positive (PCNA⁺) mesenchymal cells, and von Willebrand factor positive(vWF⁺) vessels were also significantly inhibited by EPO‐MSCs. Furthermore, EPO‐MSCs could downregulate the expression of TGF‐β1, TAK1, and p38MAPK in lung tissue both in mRNA level and in protein level. EPO gene modified MSCs may more efficiently attenuate asthmatic airway remodeling, which maybe related with the downregulation of TGF‐β1‐TAK1‐p38MAPK pathway activity.     

CD4+CD25+ T cells regulate airway eosinophilic inflammation by modulating the Th2 cell phenotype

We used a TCR-transgenic mouse to investigate whether Th2-mediated airway inflammation is influenced by Ag-specific CD4+CD25+ regulatory T cells. CD4+CD25+ T cells from DO11.10 mice expressed the transgenic TCR and mediated regulatory activity. Unexpectedly, depletion of CD4+CD25+ T cells before Th2 differentiation markedly reduced the expression of IL-4, IL-5, and IL-13 mRNA and protein when compared with unfractionated (total) CD4+ Th2 cells. The CD4+CD25--derived Th2 cells also expressed decreased levels of IL-10 but were clearly Th2 polarized since they did not produce any IFN-gamma. Paradoxically, adoptive transfer of CD4+CD25--derived Th2 cells into BALB/c mice induced an elevated airway eosinophilic inflammation in response to OVA inhalation compared with recipients of total CD4+ Th2 cells. The pronounced eosinophilia was associated with reduced levels of IL-10 and increased amounts of eotaxin in the bronchoalveolar lavage fluid. This Th2 phenotype characterized by reduced Th2 cytokine expression appeared to remain stable in vivo, even after repeated exposure of the animals to OVA aerosols. Our results demonstrate that the immunoregulatory properties of CD4+CD25+ T cells do extend to Th2 responses. Specifically, CD4+CD25+ T cells play a key role in modulating Th2-mediated pulmonary inflammation by suppressing the development of a Th2 phenotype that is highly effective in vivo at promoting airway eosinophilia. Conceivably, this is partly a consequence of regulatory T cells facilitating the production of IL-10.     

Influences of PON1 on airway inflammation and remodeling in bronchial asthma

This study aims to explore the influences of Paraoxonase‐1 (PON1) involved in airway inflammation and remodeling in asthma. Mice were divided into control, asthma, asthma + PON1 and asthma + NC groups, and asthma models were established via aerosol inhalation of ovalbumin (OVA). HE, Masson, and PAS stains were used to observe airway inflammation and remodeling, Giemsa staining to assess inflammatory cells in bronchoalveolar lavage fluid (BALF), qRT‐PCR and Western blot to detect PON1 expression, lipid peroxidation and glutathione assays to quantify malondialdehyde (MDA) activity and glutathione peroxidase (GSH) levels, ELISA to determine inflammatory cytokines and immunoglobulin, and colorimetry to detect PON1 activities. Additionally, mice lung macrophages and fibroblasts were transfected with PON1 plasmid in vitro; ELISA and qRT‐PCR were performed to understand the effects of PON1 on inflammatory cytokines secreted by lung macrophages, MTT assay for lung fibroblasts proliferation and qRT‐PCR and Western blot for the expressions of PON1, COL1A1, and fibronectin. After overexpression of PON1, the asthma mice had decreased inflammatory cell infiltration, fibrosis degree, and airway wall thickness; inflammatory cells and inflammatory cytokines in BALF were also reduced, expressions of OVA‐IgE and IgG1, and MDA activity were decreased, but the expressions of OVA‐IgG2a and INF‐γ and GSH levels were increased. Besides, PON1 significantly inhibited microphage expression of LPS‐induced inflammatory cytokines, lung fibroblast proliferation, and COL1A1 and fibronectin expression. Thus, PON1 could relieve airway inflammation and airway remodeling in asthmatic mice and inhibit the secretion of LPS‐induced macrophage inflammatory cytokines and the proliferation of lung fibroblasts.     

Aged regulatory T cells protect from autoimmune inflammation despite reduced STAT3 activation and decreased constraint of IL-17 producing T cells

Regulatory T cells (Tregs) are specialized CD4⁺ T lymphocytes helping defend against autoimmunity and inflammation. Although age is associated with increased inflammation and autoimmunity, few reports address age effects of immune regulation or auto‐aggressive T cells. We show here that young and aged naïve CD4⁺ T cells are equivalently auto‐aggressive in vivo in T cell‐driven autoimmune colitis. Young and aged CD4⁺ Tregs equally suppressed age‐matched T cell proliferation in vitro and controlled clinical and pathologic T cell‐driven autoimmune colitis, suggesting equivalent regulatory function. However, whereas young and aged CD4⁺ Tregs suppressed interferon (IFN)‐γ⁺ T cells equivalently in this model, aged CD4⁺ Tregs unexpectedly failed to restrain interleukin (IL)‐17⁺ T cells. Nonetheless, young and aged CD4⁺ Tregs equally restrained IL‐17⁺ T cells in vivo during acute inflammation, suggesting a chronic inflammation‐related defect in aged CD4⁺ Tregs. In support, aged Tregs expressed reduced STAT3 activation, a defect associated with poor IL‐17‐producing T cell restraint. Aged naïve mice had markedly increased programmed death (PD)‐1⁺ T cells, but these exhibited no significant auto‐aggressive or regulatory functions in T cell‐driven colitis. Young CD8⁺ CD122^? T cells induce autoimmune bone marrow failure, but we show that aged CD8⁺ CD122^? T cells do not. These data demonstrate no apparent age‐related increase in auto‐aggressive T cell behavior, but disclose previously unrecognized functional defects in aged CD4⁺ Tregs during chronic inflammation. IL‐17 can be inflammatory and contributes to certain autoimmune disorders. Reduced aged Treg function during chronic inflammation and reduced IL‐17 restraint could contribute to age‐related inflammation or autoimmunity.     

Effects of Cystamine on antioxidant activities and regulatory T cells in lupus‐prone mice

Attenuated antioxidant activities, irregular cytokines expressions and reduced regulatory T cells, are strongly associated with the pathogenesis of systemic lupus erythematosus (SLE). Despite the well‐established beneficial effects of cystamine on lupus‐prone mice, the extent to which cystamine contributes to antioxidant activity and the reduction of regulatory T cells has seldom been investigated. Therefore, this study elucidates how cystamine affects anti‐oxidant activities in NZB/W F1 mice by performing assays of Glutathione (GSH), 1,1‐diphenyl‐2‐ picryl‐hydrazyl (DPPH) and malondialdehyde thiobarbituric acid (MDA). In addition, investigations of the effects of cystamine on CD4⁺/CD25⁺ regulatory T cells and interleukin‐6 (IL6)/STAT‐3 signalling were performed with flow cytometry and immunoblots. Experimental results reveal more significantly reduced MDA and increased GSH and DPPH in NZB/W F1 mice receiving cystamine than in those mice receiving PBS. Meanwhile, CD4⁺/CD25⁺ regulatory T cells more significantly increase in NZB/W F1 mice receiving cystamine than in those mice receiving PBS, accompanied by significantly reduced IL‐6/phosphorylated STAT‐3 expression. The above findings suggest the beneficial effects of cystamine in terms of increasing antioxidant activities and CD4⁺/CD25⁺ regulatory T cells in lupus‐prone mice by suppressing IL‐6/STAT3 signalling.     

High‐dose wogonin exacerbates DSS‐induced colitis by up‐regulating effector T cell function and inhibiting Treg cell

Wogonin exerts anti‐tumour activities via multiple mechanisms. We have identified that high‐dose wogonin (50 or 100 mg/kg) could inhibit the growth of transplanted tumours by directly inducing tumour apoptosis and promoting DC, T and NK cell recruitment into tumour tissues to enhance immune surveillance. However, wogonin (20–50 μM) ex vivo prevents inflammation by inhibiting NF‐κB and Erk signalling of macrophages and epithelial cells. It is elusive whether high‐dose wogonin promotes or prevents inflammation. To investigate the effects of high‐dose wogonin on murine colitis induced by dextran sodium sulphate (DSS), mice were co‐treated with DSS and various doses of wogonin. Intraperitoneal administration of wogonin (100 mg/kg) exacerbated DSS‐induced murine colitis. More CD4⁺ CD44⁺ and CD8⁺ CD44⁺ cells were located in the inflamed colons in the wogonin (100 mg/kg) treatment group than in the other groups. Frequencies of CD4⁺ CD25⁺ CD127^? and CD4⁺ CD25⁺ Foxp3⁺ cells in the colons and spleen respectively, were reduced by wogonin treatment. Ex vivo stimulations with high‐dose wogonin (50–100 μg/ml equivalent to 176–352 μM) could synergize with IL‐2 to promote the functions of CD4⁺ and CD8⁺ cells. However, regulatory T cell induction was inhibited. Wogonin stimulated the activation of NF‐κB and Erk but down‐regulated STAT3 phosphorylation in the CD4⁺ T cells. Wogonin down‐regulated Erk and STAT3‐Y705 phosphorylation in the regulatory T cells but promoted NF‐κB and STAT3‐S727 activation. Our study demonstrated that high‐dose wogonin treatments would enhance immune activity by stimulating the effector T cells and by down‐regulating regulatory T cells.     

Antigen-Specific IgG ameliorates allergic airway inflammation via Fcγ receptor IIB on dendritic cells

Background

There have been few reports on the role of Fc receptors (FcRs) and immunoglobulin G (IgG) in asthma. The purpose of this study is to clarify the role of inhibitory FcRs and antigen presenting cells (APCs) in pathogenesis of asthma and to evaluate antigen-transporting and presenting capacity by APCs in the tracheobronchial mucosa.

Methods

In FcγRIIB deficient (KO) and C57BL/6 (WT) mice, the effects of intratracheal instillation of antigen-specific IgG were analysed using the model with sensitization and airborne challenge with ovalbumin (OVA). Thoracic lymph nodes instilled with fluorescein-conjugated OVA were analysed by fluorescence microscopy. Moreover, we analysed the CD11c⁺ MHC class II⁺ cells which intaken fluorescein-conjugated OVA in thoracic lymph nodes by flow cytometry. Also, lung-derived CD11c⁺ APCs were analysed by flow cytometry. Effects of anti-OVA IgG1 on bone marrow dendritic cells (BMDCs) in vitro were also analysed. Moreover, in FcγRIIB KO mice intravenously transplanted dendritic cells (DCs) differentiated from BMDCs of WT mice, the effects of intratracheal instillation of anti-OVA IgG were evaluated by bronchoalveolar lavage (BAL).

Results

In WT mice, total cells and eosinophils in BAL fluid reduced after instillation with anti-OVA IgG1. Anti-OVA IgG1 suppressed airway inflammation in hyperresponsiveness and histology. In addition, the number of the fluorescein-conjugated OVA in CD11c⁺ MHC class II⁺ cells of thoracic lymph nodes with anti-OVA IgG1 instillation decreased compared with PBS. Also, MHC class II expression on lung-derived CD11c⁺ APCs with anti-OVA IgG1 instillation reduced. Moreover, in vitro, we showed that BMDCs with anti-OVA IgG1 significantly decreased the T cell proliferation. Finally, we demonstrated that the lacking effects of anti-OVA IgG1 on airway inflammation on FcγRIIB KO mice were restored with WT-derived BMDCs transplanted intravenously.

Conclusion

Antigen-specific IgG ameliorates allergic airway inflammation via FcγRIIB on DCs.     

Interleukin‐38 protects against sepsis by augmenting immunosuppressive activity of CD4+CD25+ regulatory T cells

Naturally occurring CD4⁺CD25⁺ regulatory T cells (Tregs) are required to limit immune‐induced pathology and to maintain homeostasis during the early‐phase of sepsis. This study aimed to investigate the role of interleukin (IL)‐38, a newly described member of the IL‐1 cytokine family, in mediated immune response of CD4⁺CD25⁺ Tregs in sepsis. Here, we provide evidence that expressions of IL‐38 and its receptor were detected in murine CD4⁺CD25⁺ Tregs. Stimulation of CD4⁺CD25⁺ Tregs with LPS markedly up‐regulated the expression of IL‐38. Treatment with rmIL‐38 dramatically enhanced the immunosuppressive activity of CD4⁺CD25⁺ Tregs after LPS stimulation and in septic mice induced by CLP, resulting in amplification of helper T cell (Th) 2 response and reduction in the proliferation of effector T cells. These effects were robustly abrogated when anti–IL‐38 antibody was administered. Administration of rmIL‐38 improved the survival rate of CLP mice. In addition, CD4⁺CD25⁺ Tregs depletion before the onset of sepsis obviously abolished IL‐38–mediated protective response. These findings suggest that IL‐38 enhances the immunosuppressive activity of CD4⁺CD25⁺ Tregs, which might contribute to the improvement of host immune function and prognosis in the setting of sepsis.     

Clonorchis sinensis-derived total protein attenuates airway inflammation in murine asthma model by inducing regulatory T cells and modulating dendritic cell functions

Asthma is characterized by Th2-mediated inflammation, resulting in airway hyperresponsiveness (AHR) through airway remodeling. Recent epidemiological and experimental reports have suggested an inverse relationship between the development of allergy and helminth infections. Infection by Clonorchis sinensis, a liver fluke that resides in the bile duct of humans, is endemic predominantly in Asia including Korea and China. Using a murine model for asthma, we investigated the effects of C. sinensis-derived total protein (Cs-TP) on allergen-induced airway inflammation and the mechanism underlying the protective effects of Cs-TP administration on asthma. Treatment with Cs-TP attenuated OVA-induced airway inflammation and methacholine-induced AHR, as well as eosinophilia development, lymphocyte infiltration into the lung, and goblet cell metaplasia. This protective effect of Cs-TP is associated with markedly reduced OVA-specific IgE and Th1/Th2 cytokine production. Moreover, Cs-TP increased the number of CD4⁺CD25⁺Foxp3⁺ regulatory T (Treg) cells as well as their suppressive activity. In fact, proliferation of OVA-restimulated splenocytes was suppressed significantly. Cs-TP also inhibited the expression of such co-stimulatory molecules as CD80, CD86, and CD40 in LPS- or OVA-stimulated dendritic cells (DCs), suggesting that Cs-TP could interfere with the capacity of airway DCs to prime naïve T cells. These data demonstrate the capacity of C. sinensis to ameliorate allergic asthma and broaden our understanding of the paradoxical relationship between the allergic immune response and helminth infection.     

Human herpesvirus-6-specific interleukin 10-producing CD4+ T cells suppress the CD4+ T-cell response in infected individuals

Human herpesvirus‐6 (HHV‐6) infection normally persists for the lifetime of the host and may reactivate with immunosuppression. The mechanism behind HHV‐6 latent infection is still not fully understood. In this study, we observed that decreased proliferation of CD4⁺ T cells and PBMCs but not CD8⁺ T cells from HHV‐6‐infected individuals was stimulated with HHV‐6‐infected cell lysates. Moreover, HHV‐6‐stimulated CD4⁺ T cells from HHV‐6‐infected individuals have suppressive activity on naïve CD4⁺ T and CD8⁺ T cells from HHV‐6‐uninfected individuals. However, no increased proportion of CD4⁺ CD25⁺ Treg cells from HHV‐6‐infected individuals contributed to the suppressive activity of the HHV‐6‐stimulated CD4⁺ T cells from HHV‐6‐infected individuals. Transwell experiments, ELISA and anti‐IL‐10 antibody blocking experiment demonstrated that IL‐10 may be the suppressive cytokine required for suppressive activity of CD4⁺ T cells from HHV‐6‐infected individuals. Results of intracellular interleukin (IL)‐10 and IL‐4 further implicated the HHV‐6‐speciflc IL‐10‐producing CD4⁺ T cells in the suppressive activity of CD4⁺ T cells from HHV‐6‐infected individuals. Results of intracellular interferon (IFN)‐γ demonstrated a decreased frequency of HHV‐6‐speciflc IFN‐γ‐producing CD4⁺ T, but not CD8⁺ T cells in HHV‐6‐infected individuals, indicating that it was the CD4⁺ Th1 responses in HHV‐6‐infected individuals that were selectively impaired. Our findings indicated that HHV‐6‐specific IL‐10‐producing CD4⁺ T cells from HHV‐6‐infected individuals possess T regulatory type 1 cell activity: immunosuppression, high levels of IL‐10 production, with a few cells expressing IFN‐γ, but none expressing IL‐4. These cells may play an important role in latent HHV‐6 infection.     

Induction of regulatory T cells by physiological level estrogen

Naturally occurring CD4⁺CD25⁺ regulatory T cells (Treg) exert an important role in mediating maternal tolerance to the fetus during pregnancy, and this effect might be regulated via maternal estrogen secretion. Although estrogen concentration in the pharmaceutical range has been shown to drive expansion of CD4⁺CD25⁺ Treg cells, little is known about how and through what mechanisms E2 within the physiological concentration range of pregnancy affects this expansion. Using in vivo and in vitro mouse models in these experiments, we observed that E2 at physiological doses not only expanded Treg cell in different tissues but also increased expression of the Foxp3 gene, a hallmark for CD4⁺CD25⁺ Treg cell function, and the IL‐10 gene as well. Importantly, our results demonstrate that E2, at physiological doses, stimulated the conversion of CD4⁺CD25^? T cells into CD4⁺CD25⁺ T cells which exhibited enhanced Foxp3 and IL‐10 expression in vitro. Such converted CD4⁺CD25⁺ T cells had similar regulatory function as naturally occurring Treg cells, as demonstrated by their ability to suppress naïve T cell proliferation in a mixed lymphocyte reaction. We also found that the estrogen receptor (ER) exist in the CD4⁺CD25^? T cells and the conversion of CD4⁺CD25^? T cells into CD4⁺CD25⁺ T cells stimulated by E2 could be inhibited by ICI182,780, a specific inhibitor of ER(s). This supports that E2 may directly act on CD4⁺CD25^? T cells via ER(s). We conclude that E2 is a potential physiological regulatory factor for the peripheral development of CD4⁺CD25⁺ Treg cells during the implantation period in mice. J. Cell. Physiol. 214: 456–464, 2008. © 2007 Wiley‐Liss, Inc.     

Parasitic Nematode-Induced CD4+Foxp3+T Cells Can Ameliorate Allergic Airway Inflammation

Background

The recruitment of CD4⁺CD25⁺Foxp3⁺T (T_reg) cells is one of the most important mechanisms by which parasites down-regulate the immune system.

Methodology/Principal Findings

We compared the effects of T_reg cells from Trichinella spiralis-infected mice and uninfected mice on experimental allergic airway inflammation in order to understand the functions of parasite-induced T_reg cells. After four weeks of T. spiralis infection, we isolated Foxp3-GFP-expressing cells from transgenic mice using a cell sorter. We injected CD4⁺Foxp3⁺ cells from T. spiralis-infected [Inf(+)Foxp3⁺] or uninfected [Inf(-)Foxp3⁺] mice into the tail veins of C57BL/6 mice before the induction of inflammation or during inflammation. Inflammation was induced by ovalbumin (OVA)-alum sensitization and OVA challenge. The concentrations of the Th2-related cytokines IL-4, IL-5, and IL-13 in the bronchial alveolar lavage fluid and the levels of OVA-specific IgE and IgG1 in the serum were lower in mice that received intravenous application of Inf(+)Foxp3⁺ cells [IV(inf):+(+) group] than in control mice. Some features of allergic airway inflammation were ameliorated by the intravenous application of Inf(-)Foxp3⁺ cells [IV(inf):+(-) group], but the effects were less distinct than those observed in the IV(inf):+(+) group. We found that Inf(+)Foxp3⁺ cells migrated to inflammation sites in the lung and expressed higher levels of T_reg-cell homing receptors (CCR5 and CCR9) and activation markers (Klrg1, Capg, GARP, Gzmb, OX40) than did Inf(-)Foxp3⁺ cells.

Conclusion/Significance

T. spiralis infection promotes the proliferation and functional activation of T_reg cells. Parasite-induced T_reg cells migrate to the inflammation site and suppress immune responses more effectively than non-parasite-induced T_reg cells. The adoptive transfer of Inf(+)Foxp3⁺ cells is an effective method for the treatment and prevention of allergic airway diseases in mice and is a promising therapeutic approach for the treatment of allergic airway diseases.     

Trichinella spiralis: infection reduces airway allergic inflammation in mice

In an effort to define the mechanism underlying the host immune downregulation inherent to Trichinella spiralis infection, we compared the levels of Th1, Th2, and regulatory cytokines and CD4⁺CD25⁺ forkhead box P3 (FoxP3)⁺ T (T_reg) cell recruitment, as well as cellular pathology in the airway between T. spiralis infected and uninfected asthma-induced mice. After the induction of allergic airway inflammation, we noted influxes of inflammatory cells into the peribronchial tree. However, in the T. spiralis infection groups, cellular infiltration was minimal around the bronchial tree, with only a smattering of inflammatory cells. In the OVA-challenged group after T. spiralis infection, the numbers of macrophages and eosinophils in the bronchial alveolar lavage fluid were reduced by 23% and 52%, respectively, as compared to those of the OVA-challenged group. Airway hyperresponsiveness of OVA-challenged mice after T. spiralis infection was significantly suppressed as compared to the OVA-only challenged mice. The T. spiralis-infected mice exhibited a significant reduction in IL-5 concentrations relative to that noted in the OVA-challenged group (p < 0.01). Nevertheless, the regulatory cytokines IL-10 and TGF-β levels were increased significantly as the result of T. spiralis infection, and we verified the recruitment of T_reg cells in lung draining lymph nodes via T. spiralis infection. Therefore, T_reg cells, which were recruited by T. spiralis infection, might ameliorate lung function and reduce allergic airway inflammation.     

Heme oxygenase-1-mediated CD4+CD25high regulatory T cells suppress allergic airway inflammation

Heme oxygenase-1 (HO-1) has anti-inflammatory effects in asthma. CD4+CD25(high) regulatory T cells (Treg) are a potent immunoregulator that suppresses the immune response. We studied the effects of HO-1-mediated CD4+CD25(high) Treg on suppression of allergic airway inflammation by comparing mice treated with hemin, OVA, Sn-protoporphyrin (SnPP), and hemin plus SnPP. Airway responsiveness, airway eosinophil infiltration, the level of OVA-specific IgE, and the numbers of cells in general and eosinophils in particular in bronchial alveolar lavage fluid were lower in the hemin group than in the OVA, SnPP, and hemin plus SnPP groups. The expressions of HO-1 mRNA and protein in the lung were increased by repeated administrations of hemin and SnPP. However, the activity of HO-1 was highest in hemin mice. The percentage and suppressive function of CD4+CD25(high) Treg and the expression of Foxp3 mRNA were obviously enhanced after treatment with hemin. This increase was diminished by the administration of SnPP. The concentration of serum IL-10 was higher in the hemin group than in the other groups, whereas the level of serum TGF-beta did not significantly differ across groups. Furthermore, the ratio of IFN-gamma/IL-4 mRNA in the lung was higher in hemin-treated mice than in OVA and SnPP mice. The suppressive capacity of CD4+CD25(high) Treg was not enhanced in the IL-10-deficient mice treated with hemin. In conclusion, our experiments in the animal model demonstrated that HO-1 has anti-inflammatory effects, probably via enhancement of the secretion of IL-10 and promotion of the percentage of CD4+CD25(high) Treg.     

Attenuation of antigen-induced airway hyperresponsiveness and inflammation in CXCR3 knockout mice

Background

CD8+ T cells participate in airway hyperresponsiveness (AHR) and allergic pulmonary inflammation that are characteristics of asthma. CXCL10 by binding to CXCR3 expressed preferentially on activated CD8+ T cells, attracts T cells homing to the lung. We studied the contribution and limitation of CXCR3 to AHR and airway inflammation induced by ovalbumin (OVA) using CXCR3 knockout (KO) mice.

Methods

Mice were sensitized and challenged with OVA. Lung histopathological changes, AHR, cellular composition and levels of inflammatory mediators in bronchoalveolar lavage (BAL) fluid, and lungs at mRNA and protein levels, were compared between CXCR3 KO mice and wild type (WT) mice.

Results

Compared with the WT controls, CXCR3 KO mice showed less OVA-induced infiltration of inflammatory cells around airways and vessels, and less mucus production. CXCR3 KO mice failed to develop significant AHR. They also demonstrated significantly fewer CD8+ T and CD4+ T cells in BAL fluid, lower levels of TNFα and IL-4 in lung tissue measured by real-time RT-PCR and in BAL fluid by ELISA, with significant elevation of IFNγ mRNA and protein expression levels.

Conclusions

We conclude that CXCR3 is crucial for AHR and airway inflammation by promoting recruitment of more CD8+ T cells, as well as CD4+ T cells, and initiating release of proinflammatory mediators following OVA sensitization and challenge. CXCR3 may represent a novel therapeutic target for asthma.     

Preventative Effect of an Herbal Preparation (HemoHIM) on Development of Airway Inflammation in Mice via Modulation of Th1/2 Cells Differentiation

HemoHIM, an herbal preparation of three edible herbs (Angelica gigas Nakai, Cnidium officinale Makino, Paeonia japonica Miyabe) is known to increase the Th1 immune response as well as reduce the allergic response in human mast cells. Here, our goal was to determine whether or not HemoHIM could induce Th1 cell differentiation as well as inhibit the development of airway inflammation. To study Th1/Th2 cell differentiation, naive CD4⁺ T cells isolated from C57BL/6 mouse spleens were cultured with or without HemoHIM. To examine airway inflammation, C57BL/6 mice were fed HemoHIM for 4 weeks before sensitization and provocation with ovalbumin (OVA). In an in vitro experiment, naive CD4⁺ T cells displayed increased Th1 (IFN-γ⁺ cell) as well as decreased Th2 (IL-4⁺ cell) differentiation in a HemoHIM concentration-dependent manner. Furthermore, in an airway inflammation mice model, eosinophil numbers in BALF, serum levels of OVA-specific IgE and IgG1, and cytokine (IL-4, IL-5, and IL-13) levels in BALF and the supernatant of splenocytes all decreased upon HemoHIM (100 mg/kg body weight) pretreatment (4 weeks). These results show that HemoHIM attenuated allergic airway inflammation in the mouse model through regulation of the Th1/Th2 balance.     

Protection against the allergic airway inflammation depends on the modulation of spleen dendritic cell function and induction of regulatory T cells in mice

Background

Allergen-induced imbalance of specific T regulatory (Treg) cells and T helper 2 cells plays a decisive role in the development of immune response against allergens.

Objective

To evaluate effects and potential mechanisms of DNA vaccine containing ovalbumin (OVA) and Fc fusion on allergic airway inflammation.

Methods

Bronchoalveolar lavage (BAL) levels of inflammatory mediators and leukocyte infiltration, expression of CD_11c ⁺CD₈₀ ⁺ and CD_11c ⁺CD₈₆ ⁺ co-stimulatory molecules in spleen dendritic cells (DCs), circulating CD₄ ⁺ and CD₈ ⁺ T cells, Foxp3⁺ in spleen CD₄ ⁺ T cells and spleen CD₄ ⁺ T cells were measured in OVA-sensitized and challenged animals pretreated with pcDNA, OVA-pcDNA, Fc-pcDNA, and OVA-Fc-pcDNA.

Results

OVA-Sensitized and challenged mice developed airway inflammation and Th2 responses, and decreased the proliferation of peripheral CD₄ ⁺and CD₈ ⁺ T cells and the number of spleen Foxp₃ ⁺ Treg. Those changes with increased INF-γ production and reduced OVA-specific IgE production were protected by the pretreatment with OVA-Fc-pcDNA.

Conclusion

DNA vaccine encoding both Fc and OVA showed more effective than DNA vaccine encoding Fc or OVA alone, through the balance of DCs and Treg.