Ding Chuan Tang Attenuates Airway Inflammation and Eosinophil Infiltration in Ovalbumin-Sensitized Asthmatic Mice.

Asthma is a T helper 2 (Th2) cell-associated chronic inflammatory diseases characterized with airway obstruction, increased mucus production, and eosinophil infiltration. Conventional medications for asthma treatment cannot fully control the symptoms, and potential side effects are also the concerns. Thus, complement or alternative medicine (CAM) became a new option for asthma management. Ding Chuan Tang (DCT) is a traditional Chinese herbal decoction applied mainly for patients with coughing, wheezing, chest tightness, and asthma. Previously, DCT has been proved to improve children airway hyperresponsiveness (AHR) in a randomized and double-blind clinical trial. However, the mechanisms of how DCT alleviates AHR remain unclear. Since asthmatic features such as eosinophil infiltration, IgE production, and mucus accumulation are relative with Th2 responses, we hypothesized that DCT may attenuate asthma symptoms through regulating Th2 cells. Ovalbumin (OVA) was used as a stimulant to sensitize BALB/c mice to establish an asthmatic model. AHR was detected one day before sacrifice. BALF and serum were collected for immune cell counting and antibody analysis. Splenocytes were cultured with OVA in order to determine Th2 cytokine production. Lung tissues were collected for histological and gene expression analyses. Our data reveal that DCT can attenuate AHR and eosinophil accumulation in the 30-day sensitization asthmatic model. Histological results demonstrated that DCT can reduce cell infiltration and mucus production in peribronchial and perivascular site. In OVA-stimulated splenocyte cultures, a significant reduction of IL-5 and IL-13 in DCT-treated mice suggests that DCT may alleviate Th2 responses. In conclusion, the current study demonstrates that DCT has the potential to suppress allergic responses through the reduction of mucus production, eosinophil infiltration, and Th2 activity in asthma.

Anti-allergic effects of sesquiterpene lactones from Saussurea costus (Falc.) Lipsch. determined using in vivo and in vitro experiments.

ETHNOPHARMACOLOGICAL RELEVANCE: Saussurea costus (Falc.) Lipsch. root has been used in Asian traditional medicine for the treatment of asthma, rheumatism, and other conditions. S. costus extracts were shown to alleviate house dust mite-induced atopic-like dermatitis in Nc/Nga mice; besides, sesquiterpene lactones were isolated from S. costus extracts. AIMS OF THE STUDY: We aimed to investigate the effects of sesquiterpene lactones (alantolactone, costunolide, and dehydrocostuslactone) in allergic asthma using female Balb/c mice and rat RBL-2H3 mast cells. MATERIALS AND METHODS: Antigen-induced degranulation was assessed by measuring ß-hexosaminidase activity in vitro. In addition, a murine ovalbumin-induced allergic asthma model was used to test the in vivo efficacy of sesquiterpene lactones. RESULTS: Sesquiterpene lactones inhibited antigen-induced degranulation, wherein dehydrocostuslactone > costunolide > alantolactone in potency. Administration of sesquiterpene lactones decreased the number of immune cells, particularly eosinophils, and reduced the expression and secretion of Th2 cytokines (IL-4 and IL-13) in the bronchoalveolar lavage fluid and lung tissues of mice with ovalbumin-induced allergic asthma. Histological studies showed that sesquiterpene lactones reduced inflammation and mucin production in the lungs. Similar to the in vitro study, dehydrocostuslactone showed the highest potency, followed by costunolide and alantolactone. CONCLUSION: These findings provide evidence that sesquiterpene lactones might be potential anti-allergic therapeutics.

IL-13 Stimulates Proliferation and Expression of Mucin and Immunomodulatory Genes in Cultured Conjunctival Goblet Cells.

PURPOSE: To investigate the effects of IL-13 on goblet cell proliferation, differentiation, and expression of mucin and immunomodulatory genes. METHODS: Explants were excised from the conjunctiva of young C57BL/6 mice. Cultures received 200 µL per week of either Keratinocyte media (KSFM) or KSFM supplemented with 10 ng/mL IL-13 and were incubated for 3 (D3), 7 (D7), or 14 (D14) days. Subsequently, cell proliferation was assessed or cultures were immunostained, collected for dot blot, or for reverse transcription (RT) and quantitative real-time PCR (qPCR) or for RT-PCR gene array. RESULTS: The cultured conjunctival epithelium expressed goblet cell associated keratin 7 and mucins MUC5AC and MUC2 and when stimulated with IL-13 showed increased proliferation at D3 and D7 (P < 0.05) compared with control. MUC5AC expression was increased in the IL-13-treated group at D3 and D14 (P < 0.05). IL-13-treated cultures showed increased chemokine ligand 26 (CCL26), chloride channel calcium activated channel 3 (CLCA3), fas ligand (FasL), and Relm-ß at D7. All conjunctival cultures expressed MUC2, and its expression was decreased at D3 (P < 0.05) and increased at D14 (P < 0.05) with IL-13 treatment. CONCLUSIONS: This study demonstrated that conjunctival goblet cells are IL-13 responsive cells that produce factors known to maintain epithelial barrier, stimulate mucin production, and modulate immune response in nonocular mucosa when treated with IL-13. The functional significance of IL-13-stimulated factors remains to be determined.

Direct contact between dendritic cells and bronchial epithelial cells inhibits T cell recall responses towards mite and pollen allergen extracts in vitro.

Airway epithelial cells (AECs) form a polarized barrier along the respiratory tract. They are the first point of contact with airborne antigens and are able to instruct resident immune cells to mount appropriate immune responses by either soluble or contact-dependent mechanisms. We hypothesize that a healthy, polarized epithelial cell layer inhibits inflammatory responses towards allergens to uphold homeostasis. Using an in-vitro co-culture model of the airway epithelium, where a polarized cell layer of bronchial epithelial cells can interact with dendritic cells (DCs), we have investigated recall T cell responses in allergic patients sensitized to house dust mite, grass and birch pollen. Using allergen extract-loaded DCs to stimulate autologous allergen-specific T cell lines, we show that AEC-imprinted DCs inhibit T cell proliferation significantly of Bet v 1-specific T cell lines as well as decrease interleukin (IL)-5 and IL-13 production, whereas inhibition of Phl p 5-specific T cells varied between different donors. Stimulating autologous CD4(+) T cells from allergic patients with AEC-imprinted DCs also inhibited proliferation significantly and decreased production of both T helper type 1 (Th1) and Th2 cytokines upon rechallenge. The inhibitory effects of AECs' contact with DCs were absent when allergen extract-loaded DCs had been exposed only to AECs supernatants, but present after direct contact with AECs. We conclude that direct contact between DCs and AECs inhibits T cell recall responses towards birch, grass and house dust mite allergens in vitro, suggesting that AECs-DC contact in vivo constitute a key element in mucosal homeostasis in relation to allergic sensitisation.

Cutaneous or respiratory exposures to peanut allergens in mice and their impacts on subsequent oral exposure.

BACKGROUND: Recent data suggested that non-gastrointestinal exposure can lead to sensitisation to food allergens. We thus assessed the immune impact of respiratory or cutaneous exposure to peanut proteins on non-altered epithelium and investigated the effect of such pre-exposure on subsequent oral administration of peanut. METHODS: BALB/cJ mice were exposed to purified Ara h 1 or to a non-defatted roasted peanut extract (PE) by simple deposit of allergens solutions on non-altered skin or in the nostrils. Exposures were performed 6 times at weekly intervals. Pre-exposed mice then received intra-gastric administrations of PE alone or in the presence of the Th2 mucosal adjuvant cholera toxin (CT). The specific humoral and cellular immune response was assessed throughout the protocol. RESULTS: Both cutaneous and respiratory exposures led to the production of specific IgG1. Local and systemic IL-5 and IL-13 production were also evidenced, demonstrating activation of specific Th2 cells. This effect was dose-dependent and most efficient via the respiratory route. Moreover, these pre-exposures led to the production of specific IgE antibodies after gavage with PE, whatever the presence of CT. CONCLUSIONS: Cutaneous or respiratory exposures to peanut induce Th2 priming in mice. Moreover, pre-exposures promote further sensitisation via the oral route without the use of CT; this proposes a new adjuvant-free experimental model of sensitisation to food that may reflect a realistic exposure pattern in infants. These results also suggest that non-gastrointestinal peanut exposure should be minimised in high-risk infants, even those with non-altered skin, to potentially reduce allergic sensitisation to this major food allergen.

In vitro action of sho-seiryu-to on allergen-exposed mononuclear cells.

Although Sho-seiryu-to (SST), used as a traditional herbal (Kampo) medicine mainly in China and Korea, is shown to have immunomodulating potential, such as an anti-allergic one, its underlying mechanism has not been completely clarified. To partially address the issue, we explored its effects on allergen-exposed mononuclear cells. Male balb/c mice were intraperitoneally administered ovalbumin (OVA: 20 μg) plus alum or vehicle twice (Day 0 and Day 14). At Day 21, mice were sacrificed and splenocytes (mononuclear cells) were isolated and cultured in the presence or absence of OVA with or without SST. Thereafter, helper T-related cytokines in the culture supernatants were evaluated by means of ELISA. Protein level of interferon-γ was lower than 5.0 pg/mL in the supernatants from OVA– non-exposed or -exposed mononuclear cells in the presence or absence of OVA stimulation. On the other hand, SST induced the cytokine from both types of mononuclear cells in the presence (P < 0.05) or absence of OVA stimulation as compared to corresponding control. By contrast, interleukin (IL)-4 level tended to be decreased by SST in OVA-non-exposed mononuclear cells as did IL-13 in both non-exposed and exposed mononuclear cells as compared to vehicle. In conclusion, immunoregulating efficacy by SST on allergy-prone subjects may include, at least in part, restoring helper T balance mainly through hyperproduction of IFN-γ against mononuclear cells such as lymphocytes.

Omega-3 fatty acids suppress Th2-associated cytokine gene expressions and GATA transcription factors in mast cells.

Because the interaction between omega-3 fatty acids and mast cells has remained largely unknown in allergies, we investigated whether omega-3 fatty acids affect the activation of mast cells by examining Th2-associated cytokine production and possible molecular mechanisms. Alpha-linolenic acid and its metabolites including eicosapentaenoic acid and decosahexaenoic acid induced a dramatic decrease in the production of interleukin (IL)-4, IL-5 and IL-13 in a dose-dependent manner, as well as mRNA expression of their genes, in activated MC/9 mast cells and bone marrow-derived mast cells. The effects were comparable to those of cyclosporin A (1 µM), a well-known immunosuppressive agent. Nuclear expression of GATA binding protein-1 (GATA-1) and GATA binding protein-2 (GATA-2), essential transcription factors for mast cell activation, was also greatly suppressed. However, their mRNA expressions were not affected. In P815 mast cells, which do not express GATA-1, the suppressive effects on cytokines were abolished. On the contrary, omega-3 fatty acids had less significant effects on IL-4 and IL-5 and resulted in a slight decrease in IL-13 production in EL-4 T cells. Finally, oral administration of fish oil containing high level of omega-3 fatty acids significantly reduced the severity of dermatitis and the thickening of epidermis/dermis in a NC/Nga murine atopic model. The number of cells expressing CD117(+) and FcεRIα(+) was greatly decreased and GATA-1 expression in the cells was also diminished. Taken together, omega-3 fatty acids might target mast cells to a greater extent than T cells to suppress Th2 cytokine expression by inhibiting GATAs for alleviation of allergic disease.

n-3 Fatty acids inhibit transcription of human IL-13: implications for development of T helper type 2 immune responses.

Fish oil supplementation during pregnancy has been associated with lower levels of cord blood IL-13, suggesting that the administration of n-3 fatty acids may attenuate the development of allergic disease. The present study aimed to investigate the mechanism by which n-3 fatty acid administration influences the production of IL-13. Pregnant BALB/c mice were fed nutritionally complete high-fat diets (15 %, w/w) with an n-3 fatty acid-enriched (DHA 1 %, w/w) or control diet (0 % DHA) immediately following delivery. Pups were exposed during suckling and weaned to the maternal diet for the remainder of the study. The production of IL-13, IL-4, IL-10 and interferon-γ from the splenocytes of ovalbumin (ova)-sensitised animals was assessed following in vitro ova stimulation or unstimulated conditions. Human T helper type 2 (Th2) cells were mitogen-stimulated in the presence or absence of DHA (10 µM) and assessed for IL-13 and IL-4 expression using intracellular flow cytometry. The influence on transcriptional activation was studied using a human IL-13 promoter reporter construct and electromobility shift assay. Ova-activated splenocytes from DHA-fed mice produced less IL-13 (57.2 (se 21.7) pg/ml) and IL-4 (7.33 (SE 3.4) pg/ml) compared with cells from the animals fed the control diet (161.5 (SE 45.0), P< 0.05; 33.2 (SE 11.8), P< 0.05). In vitro, DHA inhibited the expression of IL-13 protein from human Th2 cells as well as transcriptional activation and binding of the transcription factors cyclic AMP response element binding and activating transcription factor 2 to the human IL-13 promoter. These data indicate the potential of n-3 fatty acids to attenuate IL-13 expression, and suggest that they may subsequently reduce allergic sensitisation and the development of allergic disease.

Magnesium affects the cytokine secretion of CD4⁺ T lymphocytes in acute asthma.

INTRODUCTION: Magnesium (Mg) administration has been shown to promote bronchodilation and to improve lung function in asthma. It also plays an additional role in modulating the immune responses. This study was initiated to explore if Mg supplementation could affect the secretion of cytokines in acute asthmatic CD4⁺ T cells. METHODS: Total serum Mg concentrations of the acute asthmatic patients and healthy controls were determined. CD4⁺ T cells were isolated from the blood of the acute asthmatic patients. They were cultured in various concentrations of Mg-supplemented (0.8, 5, 10, 15, and 20 mmol/l) medium. Cytokine (IL-5, IL-13, and IFN-γ) levels were determined by Enzyme-Linked Immunosorbnent Assay (ELISA). RESULTS: Serum Mg concentration was lower in the acute asthmatic patients than that in the healthy controls (p < .05). The secretion of IL-5 and IL-13 was decreased, while the acute asthmatic CD4⁺ T cells were cultured in 10 and 15 mmol/l Mg-supplemented medium, respectively, as compared to the 0.8 mmol/l Mg group (p < .05). The secretion of IFN-γ increased in the 10 mmol/l Mg group (p < .05). CONCLUSION: Mg supplementation was able to modulate the immune responses of acute asthmatic CD4⁺ T cells and decrease the secretion of type 2 CD4⁺ T lymphocytes cytokines.

Effectiveness of Cissampelos sympodialis and its isolated alkaloid warifteine in airway hyperreactivity and lung remodeling in a mouse model of asthma.

BACKGROUND: Cissampelos sympodialis Eichl. (Menispermaceae) is a plant found in Northeastern and Southeast of Brazil and hot water infusion of C. sympodialis root bark is largely used in the indigenous and folk medicine to treat several inflammatory disorders, including asthma. Asthma is a chronic inflammatory allergic disease characterized by airway hyperreactivity (AHR), eosinophil tissue infiltration and lung remodeling. The aim of this study was to evaluate the therapeutic effect of C. sympodialis and its isolated alkaloid warifteine on allergen triggered airway hyperreactivity (AHR) and lung remodeling in murine model of asthma. METHODOLOGY/PRINCIPAL FINDINGS: The oral pre-treatment with C. sympodialis or warifteine inhibited allergen-induced AHR to inhaled methacholine and IL-13 levels in the bronchoalveolar lavage (BAL). In order to investigate the therapeutic potential of C. sympodialis and warifteine, animals were treated 1h after the last ovalbumin (OVA) challenge in sensitized animals. Similarly to the pre-treatment, post-treatment with warifteine was effective to inhibit significantly AHR to inhaled methacholine and to reduce IL-13 levels in the BAL. In addition, oral pre- or post-treatments with C. sympodialis or warifteine reduced OVA-induced eosinophil tissue infiltration, mucus production and subepithelial fibrosis to values similar to nonallergic controls. CONCLUSIONS: Our data show the anti-allergic and immunoregulatory properties of C. sympodialis, acting mostly through the active compound warifteine, to inhibit the airway hyperreactivity and lung remodeling through a mechanism at least partially dependent of IL-13 and eosinophil inhibition. Therefore placing warifteine as an interesting therapeutic candidate in allergic inflammation and corroborating the folk medicine use of C. sympodialis as anti-allergic plant.

Ethanol extract of Elaeocarpus petiolatus inhibits lipopolysaccharide-induced inflammation in macrophage cells.

Elaeocarpus petiolatus is known to exert active oxygen scavenging, anti-aging, and whitening actions. However, the biological effects of E. petiolatus on inflammation and the underlying mechanisms are yet to be established. In the present study, we investigated the anti-inflammatory effects of the ethanol extract from E. petiolatus (EPE) bark in murine Raw264.7 macrophages stimulated with lipopolysaccharide (LPS). EPE inhibited the production of PGE(2), TNF-α, and IL-1ß in a dose-dependent manner in Raw264.7 cells stimulated with LPS. The decrease in PGE(2) production was correlated with reduced COX-2 expression. Furthermore, EPE suppressed the phosphorylation of extracellular signal-related kinases (ERK), c-Jun N-terminal kinase (JNK), and p38 as well as translocation of the NF-κB p65 subunit from the cytosol to nucleus. Our results suggest that EPE exerts anti-inflammatory activity through inhibition of inflammatory mediators, such as PGE(2), TNF-α, and IL-1ß, and downregulation of COX-2 via suppression of NF-κB translocation and phosphorylation of ERK, JNK, and p38 in LPS-stimulated Raw264.7 cells.

A randomized placebo-controlled trial of rush preseasonal depigmented polymerized grass pollen immunotherapy.

BACKGROUND: Specific subcutaneous immunotherapy (SCIT) for seasonal rhinoconjunctivitis with unmodified allergen extracts is effective, but limited by risk of side-effects and involves treatment over 3 years. We examined a depigmented polymerized grass pollen extract for immunogenicity and for clinical efficacy in a rush preseasonal regimen. METHODS: Depigmented polymerized grass pollen extract was tested for proliferation and cytokine production by peripheral blood mononuclear cells. A prospective, double-blind, placebo-controlled trial of 195 grass pollen allergic patients treated with preseasonal rush immunotherapy using depigmented polymerized allergenic extract of mixed grass pollen was performed over 2 years. Primary outcome was combined symptom and medication score (SMS) during the peak of the second grass pollen season. Secondary outcomes included combined score over the whole season, during the first grass pollen season, individual symptom and medication scores, quality of life, well days/hell days and responder analysis. Adverse events were classified using the EAACI scale. Grass pollen-specific IgE and IgG4 were measured before and during treatment. RESULTS: Depigmented polymerized extract stimulated dose-dependent T-cell proliferation and cytokine production. Patients treated with preseasonal SCIT showed improved combined scores during peak season at year 2 (median 3.93, interquartile range 0.77-6.27 vs median 5.86 for placebo, 3.11-8.36, P < 0.01). Most secondary outcomes were significantly better for active treatment. Side-effects were minimal, with no grade 3 or 4 reactions. CONCLUSIONS: Depigmented polymerized grass pollen extract is immunogenic and clinically effective in rush preseasonal SCIT. This form of immunotherapy may be an attractive option for some patients.

The soy isoflavone equol enhances antigen-specific IgE production in ovalbumin-immunized BALB/c mice.

Although an immunomodulatory role of the soy isoflavone genistein has been demonstrated, the effects of other soy isoflavones on induction of antigen (Ag)-specific immune responses are not known. In this study, we therefore investigated the effects of daidzein and equol on ovalbumin (OVA)-specific T cell and B cell responses in BALB/c mice. Mice that had been treated with 20 mg/kg equol showed a significantly higher level of OVA-specific IgE than control mice. Levels of interferon (IFN)-gamma and interleukin (IL)-4 production were not different between the control and equol groups. However, IL-13 production level in mice administered 20 mg/kg equol was significantly higher than that in control mice. Strong induction of OVA-specific IgE production by equol was also observed in ovariectomized BALB/c mice, suggesting that the immunomodulatory effect of equol is not affected by endogenous estrogen.

Differential T-cell responses and allergen uptake after exposure of dendritic cells to the birch pollen allergens Bet v 1.0101, Bet v 1.0401 and Bet v 1.1001.

The major birch pollen allergen Bet v 1 is present in pollen as a mixture of at least 14 isoforms that share high sequence and structural identities. These isoforms possess either a high or a low IgE-binding capacity which defines them as allergenic or hypoallergenic. Recently, we could demonstrate that only the allergenic isoform Bet v 1.0101 was able to induce an IgE response in birch pollen allergic individuals. The hypoallergenic isoforms Bet v 1.0401 and Bet v 1.1001 were unable to induce IgE synthesis. T-helper cell responses against allergens are characterised by increased levels of Th2 cytokines. Therefore, we examined extent and polarisation of the Th cell response and the kinetics of the allergen uptake after exposure of dendritic cells (DCs) to these isoforms. Monocyte-derived DCs (MDDCs) from birch pollen allergic and non-atopic individuals stimulated with Bet v 1.0101, Bet v 1.0401 or Bet v 1.1001 in combination with the maturation factors TNF-α and IL-1ß resulted in a mature DC phenotype as measured by costimulatory molecule up-regulation. Only Bet v 1.0101-stimulated MDDCs from allergic subjects enhanced proliferation of autologous Th cells and the expression of the Th2 cytokines IL-5 and IL-13. Immature MDDCs of allergic individuals internalised equivalent amounts of the allergenic Bet v 1.0101 and the hypoallergenic Bet v 1.0401. In contrast, the uptake of the hypoallergenic Bet v 1.0401 by immature MDDCs of non-atopic individuals was significantly higher. These results provide evidence that DCs discriminate between allergens and highly related hypoallergens. This process may have an impact on the early phase of sensitisation.

Aqueous extracts from dietary supplements influence the production of inflammatory cytokines in immortalized and primary T lymphocytes.

BACKGROUND: Congaplex and Immuplex are dietary supplements that have been traditionally used to support immune system function. The purpose of these experiments was to determine whether Congaplex and Immuplex affect immune function using primary and immortalized T lymphocytes. METHODS: Immortalized CEM and Jurkat T lymphocytes and primary peripheral mononuclear blood cells (PBMCs) were treated with the aqueous extracts from Congaplex and Immuplex to determine the effects of these products on cytokine production in activated T lymphocytes. RESULTS: Congaplex enhanced phytohemagglutinin/phorbol 12-myristate 13-acetate (PHA/PMA) stimulation of both CEM and Jurkat cells as measured by the production of cytokines, while Immuplex suppressed PHA/PMA-induced production of cytokines, with the exception of interleukin (IL)-8 which was enhanced by Immuplex. In vitro treatment of PBMCs from 10 healthy donors with Congaplex or Immuplex decreased PHA-stimulated production of interferon (IFN)-gamma but increased the production of IL-13. CONCLUSIONS: While the effects of Congaplex and Immuplex differed in these two models, these data demonstrate that the aqueous extracts from these two dietary supplements can affect the inflammatory response of T lymphocytes.

Development of an in vitro test system measuring transcriptional downregulatory activities on IL-13.

Interleukin-13 (IL-13) has been proposed as a therapeutic target for bronchial asthma as it plays crucial roles in the pathogenesis of the disease. We developed an in vitro test system measuring transcriptional downregulatory activities on IL-13 as a primary screening method to select drug candidates from natural products. The promoter region of IL-13 (-2,048 to +1) was cloned into the upstream of a luciferase gene in the plasmid pGL4.14 containing the hygromycin resistance gene as a selection marker, generating pGL4.14-IL-13. The EL-4 thymoma and RBL-2H3 mast cells transiently expressing this plasmid highly produced the luciferase activities by responding to PI (PMA and ionomycin) stimulation up to 8-fold and 13-fold compared with the control, respectively, whereas cyclosporin A, a wellknown antiasthmatic agent, significantly downregulated the activities. The BF1 clone of RBL-2H3 cells constitutively expressing pGL4.14-IL-13 was established by selecting surviving cells under a constant lethal dose of hygromycin treatment. The feasibility of this system was evaluated by measuring the downregulatory activities of 354 natural products on the IL-13 promoter using the BF1 clone. An extract from Morus bombycis (named TBRC 156) significantly inhibited PI-induced luciferase activities and IL-13 mRNA expression, but not the protein expression. Fisetin (named TBRC 353) inhibited not only PI-induced luciferase activities and mRNA expression, but also the IL-13 protein secretion, whereas myricetin (named TBRC 354) could not suppress the IL-13 expression at all. Our data indicated that this in vitro test system is able to discriminate the effects on IL-13 expression, and furthermore, that it might be suitable as a simple and time-saving primary screening system to select antiasthmatic agents by measuring transcriptional activities of the IL-13 promoter.

The transcription factor NFATc2 controls IL-6-dependent T cell activation in experimental colitis.

The nuclear factor of activated T cells (NFAT) family of transcription factors controls calcium signaling in T lymphocytes. In this study, we have identified a crucial regulatory role of the transcription factor NFATc2 in T cell-dependent experimental colitis. Similar to ulcerative colitis in humans, the expression of NFATc2 was up-regulated in oxazolone-induced chronic intestinal inflammation. Furthermore, NFATc2 deficiency suppressed colitis induced by oxazolone administration. This finding was associated with enhanced T cell apoptosis in the lamina propria and strikingly reduced production of IL-6, -13, and -17 by mucosal T lymphocytes. Further studies using knockout mice showed that IL-6, rather than IL-23 and -17, are essential for oxazolone colitis induction. Administration of hyper-IL-6 blocked the protective effects of NFATc2 deficiency in experimental colitis, suggesting that IL-6 signal transduction plays a major pathogenic role in vivo. Finally, adoptive transfer of IL-6 and wild-type T cells demonstrated that oxazolone colitis is critically dependent on IL-6 production by T cells. Collectively, these results define a unique regulatory role for NFATc2 in colitis by controlling mucosal T cell activation in an IL-6-dependent manner. NFATc2 in T cells thus emerges as a potentially new therapeutic target for inflammatory bowel diseases.

[Inhibitory effects of Salvia miltiorrhiza injection coordinated with dexamethasone on interleukin-13 and eotaxin expression in lung of asthmatic rats].

OBJECTIVE: To investigate the molecular mechanism of inhibitory effect of Salvia miltiorrhiza Injection (SMI) coordinated with dexamethasone (DXM) on allergic airway inflammation in asthmatic rats. METHODS: Forty SD rats were randomly divided into 5 groups equally: the normal group, the asthma model group, the DXM group, the SMI group and the DXM + SMI group, they were treated with correspondant herbal medicines. Pathologic changes of lung tissue were obseved with HE stain, count of WBC and eosinophil (Eos) in bronchoalveolar lavage fluid (BALF) were estimated and the expressions of interleukin-13 (IL-13) and Eotaxin in lung tissue were measured by RT-PCR and SP method of immunohistochemistry assay. RESULTS: There was moderate inflammation in lung tissue in the SMI group, and mild inflammation in the DXM + SMI and the DXM group, which was similar to that in the normal group. Compared with the asthma model group, Eos and WBC count in BALF and the expression of IL-13 and Eotaxin in the lung tissue were significantly lower in the three treated groups (P < 0.05), particularly in the DXM + SMI group, showing a significant difference as compared with the other two groups (P < 0.05 or P < 0.01). Additionally, IL-13 expression was positively correlated with Eotaxin expression (r = 0.92, P < 0.01). CONCLUSION: SMI could inhibit the expression of IL-13 and Eotaxin in the lung of asthmatic rats, showing inhibitory effects synergistic with DXM on airway inflammation.

Interleukin-16 inhibits interleukin-13 production by allergen-stimulated blood mononuclear cells.

Expression of interleukin (IL)-16 is increased in bronchial mucosal biopsies of atopic asthmatics compared to normal controls. The functional significance of increased expression of IL-16 at sites of allergic inflammation is not yet clear. We have previously shown that IL-16 inhibits IL-5 secretion by allergen-stimulated peripheral blood mononuclear cells (PBMC). We investigated whether IL-16 inhibits the production of other T helper 2 cytokines, namely IL-13 and IL-4, by allergen-specific T cells. PBMC from ragweed-sensitive atopic subjects were stimulated with allergen extract for cytokine production in the presence or absence of rhIL-16. Production of cytokines was assessed by enzyme-linked immunosorbent assay and reverse transcription-polymerase chain reaction. To evaluate whether the modulatory effect of IL-16 on cytokine synthesis was mediated by interferon-gamma (IFN-gamma), IL-10, IL-12 or IL-18, allergen-stimulated PBMC were cultured in presence of IL-16 and neutralizing concentrations of relevant antibodies. Allergen-stimulated PBMC produced significantly elevated levels of IL-13 (90-740 pg/ml) as compared to unstimulated PBMC (0-375 pg/ml, P < 0.01). Addition of rhIL-16 resulted in down-regulation of IL-13 mRNA expression as well as significantly reduced amounts of IL-13 released by allergen-stimulated PBMC (0-457 pg/ml, P < 0.001), as observed for IL-5. No effect of IL-16 was observed on IL-4 mRNA expression. Treatment with IL-16 resulted in increased levels of IL-10 and IL-18 in allergen-stimulated cell culture. Neutralization of IFN-gamma, IL-12, IL-10 or IL-18 did not alter the inhibitory effects of IL-16 on IL-13 and IL-5 secretion by allergen-stimulated PBMC. IL-16 did not modify IL-13 synthesis by anti-CD3-stimulated CD4(+) T cells, but it significantly reduced the production of IL-5. These data suggest that IL-16 may play an important immunoregulatory role in allergic states in response to allergen.

Variation of T-cell activation in allergic subjects during natural pollen exposure.

BACKGROUND: Allergic inflammation is characterized by a Th2 activation. However, little is known about dynamics of T-cell cytokine production during natural allergen exposure. The aim of this study was to assess the Th1/Th2 balance in cypress allergic patients compared with controls, and variations of this balance over the pollen season. METHODS: Twenty cypress allergic patients and 10 controls were studied, distributed during two consecutive pollen seasons. Cytokine production was assessed by flow cytometry and ELISA. The variation of cytokine production during the pollen season was analyzed among patients in four occasions, and the preseason values were compared with controls. IL-13 and IFN-gamma-containing T cells were assessed among whole blood cells and PBMC. In addition the effect of specific stimulation by Juniperus ashei pollen extract was studied. RESULTS: Compared with controls, IL-13-producing T cells were increased in allergics in any case. By contrast, compared with controls, allergic IFN-gamma-producing T cells were decreased in whole blood, but not in PBMC, and were increased after specific stimulation. During the season, an increase in IFN-gamma- and a decrease in IL-13-producing T cells occurred in patients, whatever the culture conditions. CONCLUSION: These results show that the allergic T-cell activation is not limited to a Th2 profile: allergen-stimulated T cells are able to produce IFN-gamma at baseline, and the Th1/Th2 ratio increases during the pollen season.