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PTX3 is a unique member of the long pentraxins family and plays an indispensable role in regulating the immune system. We previously showed that PTX3 deletion aggravates allergic inflammation via a Th17 -dominant phenotype and enhanced CD4 T cell survival using a murine model of ovalbumin (OVA) induced allergic inflammation. In this study, we identified that upon OVA exposure, increased infiltration of CD11c+CD11b+ dendritic cells (DCs) was observed in the lungs of PTX3-/- mice compared to wild type littermate. Further analysis showed that a short-term OVA exposure led to an increased number of bone marrow common myeloid progenitors (CMP) population concomitantly with increased Ly6Chigh CCR2high monocytes and CD11c+CD11b+ DCs in the lungs. Also, pulmonary CD11c+CD11b+ DCs from OVA-exposed PTX3-/- mice exhibited enhanced expression of maturation markers, chemokines receptors CCR2, and increased OVA uptake and processing compared to wild type controls. Taken together, our data suggest that PTX3 deficiency heightened lung CD11c+CD11b+DC numbers and function, hence exacerbating airway inflammatory response.

Store-operated Ca(2+) entry (SOCE) is the main Ca(2+) entry pathway of non-excitable cells. In the past decade, the activation of this entry has been unveiled, with STIM1, a protein of the endoplasmic reticulum able to sense the intraluminal Ca(2+) content, and Orai1, the pore-forming unit of the Ca(2+) release activated Ca(2+) (CRAC) channels. When Ca(2+) ions are released from the endoplasmic reticulum, STIM1 proteins oligomerize and directly interact with Orai1 proteins, allowing the opening of the CRAC channels and a massive Ca(2+) ion influx known as SOCE. As Ca(2+) is involved in various cellular processes, the discovery of new drugs acting on the SOCE should be of interest to control the cell activity. By testing analogs of 2-aminoethyl diphenylborinate (2-APB), a well known, though not so selective effector of the SOCE, we identified methoxy diethylborinate (MDEB), a molecule able to potentiate the SOCE in three leukocyte and two breast cancer cell lines by increasing the Ca(2+) influx amplitude. Unlike 2-APB, MDEB does not affect the Ca(2+) pumps or the Ca(2+) release from the endoplasmic reticulum. MDEB could therefore represent the first member of a new group of molecules, specifically able to potentiate SOCE. Although not toxic for non-activated Jurkat T cells, it could induce the apoptosis of phytohemagglutinin-stimulated cells. ispartof: Cell Calcium vol:58 issue:2 pages:171-185 ispartof: location:Netherlands status: published

Asthma is characterized by an increase in airway smooth muscle mass and a decreased distance between the smooth muscle layer and the epithelium. Furthermore, there is evidence to indicate that airway smooth muscle cells (ASMC) express a wide variety of receptors involved in the immune response. The aims of this study were to examine the expression of CCR3 on ASMC, to compare this expression between asthmatic and nonasthmatic subjects, and to determine the implications of CCR3 expression in the migration of ASMC. We first demonstrated that ASMC constitutively express CCR3 at both mRNA and protein levels. Interestingly, TNF-α increases ASMC surface expression of CCR3 from 33 to 74%. Furthermore, using FACS analysis, we found that ASMC CCR3 is expressed to a greater degree in asthmatic vs control subjects (95 vs 75%). Functionality of the receptor was demonstrated by calcium assay; the addition of CCR3 ligand eotaxin to ASMC resulted in an increase in intracellular calcium production. Interestingly, ASMC was seen to demonstrate a positive chemotactic response to eotaxin. Indeed, ASMC significantly migrated toward 100 ng/ml eotaxin (2.2-fold increase, compared with control). In conclusion, the expression of CCR3 by ASMC is increased in asthmatics, and our data show that a CCR3 ligand such as eotaxin induces migration of ASMC in vitro. These results may suggest that eotaxin could be involved in the increased smooth muscle mass observed in asthmatics through the activation of CCR3.

The involvement of chemokines in eosinophil recruitment during inflammation and allergic reactions is well established. However, a functional role for chemokines in eosinophil differentiation has not been investigated. Using in situ RT-PCR, immunostaining, and flow cytometric analysis, we report that human CD34+ cord blood progenitor cells contain CCR3 mRNA and protein. Activation of CD34+ progenitor cells under conditions that promote Th2 type differentiation up-regulated surface expression of the CCR3. In contrast, activation with IL-12 and IFN-γ resulted in a significant decrease in the expression of CCR3. Eotaxin induced Ca2+ mobilization in CD34+ progenitor cells, which could explain the in vitro and in vivo chemotactic responsiveness to eotaxin. We also found that eotaxin induced the differentiation of eosinophils from cord blood CD34+ progenitor cells. The largest number of mature eosinophils was found in cultures containing eotaxin and IL-5. The addition of neutralizing anti-IL-3, anti-IL-5, and anti-GM-CSF Abs to culture medium demonstrated that the differentiation of eosinophils in the presence of eotaxin was IL-3-, IL-5-, and GM-CSF-independent. These results could explain how CD34+ progenitor cells accumulate and persist in the airways and peripheral blood of patients with asthma and highlight an alternative mechanism by which blood and tissue eosinophilia might occur in the absence of IL-5.

Chemokines are chemotactic cytokines that play an important role in recruiting leukocytes in allergic inflammation. Monocyte chemoacctractant protein (MCP)-4 is a CC chemokine with potent chemotactic activities for eosinophils, monocytes, T lymphocytes, and basophils and therefore represents a good candidate to participate in allergic reactions. To determine if MCP-4 plays a role in asthma, we have investigated the expression of MCP-4 messenger RNA (mRNA) and protein in the airways of patients with asthma and normal control subjects by in situ hybridization and immunohistochemistry. We found that MCP-4 mRNA and protein was significantly upregulated in the epithelium and submucosa of bronchial biopsies and in the bronchoalveolar lavage (BAL) cells of patients with asthma compared with normal control subjects (p0. 01). In addition, MCP-4 protein was significantly elevated in the BAL fluid of patients with atopic asthma when compared with normal control subjects (p0.01) and there was a significant correlation between MCP-4, eotaxin, and eosinophils. In support of our in situ findings demonstrating MCP-4 expression in epithelial cells and mononuclear cells in vivo, we have found that MCP-4 expression can be induced in these cells in vitro by tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta). Interferon-gamma (IFN-gamma) acted synergistically with TNF-alpha and IL-1beta in the induction of mRNA MCP-4 mRNA expression in A549 cells, whereas the glucocorticoid dexamethasone diminished the cytokine-induced expression of MCP-4. Our findings demonstrate that MCP-4 is upregulated in the airways of patients with asthma and suggest that MCP-4 plays a role in the recruitment of eosinophils into the airways of patients with asthma.

Background The allergen-induced late nasal response is associated with a high local expression of interleukin (IL) -4, a TH2-type cytokine implicated in immunoglobulin (Ig) E production, tissue eosinophilia and other events considered to be relevant to allergic inflammation. Interaction of IL-4 with its receptor activates at least two distinct signalling pathways that culminate in the transcription of specific target genes. One pathway involves the activation of a transcription factor termed signal transducer and activator of transcription factor 6 (STAT6). Objective To investigate the expression of STAT6 in the allergen-induced late nasal response and to examine the effect of local steroid treatment on STAT6 expression. Methods Inferior turbinate biopsies were obtained from subjects with allergic rhinitis out of the allergen season. Subjects were then randomized into topical steroid- (n = 6) and placebo-treated (n = 6) groups in a double-blind fashion. After a 6-week treatment period, a second nasal biopsy was performed 24 h after local challenge with allergen. STAT6 immunoreactivity was examined in biopsy specimens by immunocytochemistry using a specific monoclonal antibody. Numbers of inflammatory cells (CD3+ T cells and MBP+ eosinophils) and IL-4 mRNA+ cells were investigated by immunocytochemistry and in situ hybridization, respectively. Results STAT6 immunoreactivity was detected in all biopsies studied and localized predominantly to inflammatory tissue of the nasal mucosa. After allergen challenge, expression of STAT6 was markedly increased in placebo-treated patients (P

BACKGROUND: Although a large number of studies have documented the interaction of mesenchymal stem cells (MSCs) with cells of both the innate and adaptive immune systems, not much is known about how bacteria interact with MSCs and how this might influence MSCs behavior. In this study, we investigated the impact of Staphylococcus aureus (S. aureus), on viability and cytokines’ production of human Wharton’s jelly-MSCs (WJ-MSCs). OBJECTIVE: To investigate if WJ-MSCs: (1) internalize S. aureus; (2) are able to survive and (3) release immunomodulatory mediators after interaction with S. aureus. METHODS: WJ-MSCs were exposed to S. aureus at a multiplicity of infection (MOI) of 10:1 or 30:1 for different designed times. After interaction, intracellular bacteria were quantified as well as MSCs viability. Expression and cytokine-secretion were assessed using quantitative real-time PCR and ELISA. RESULTS: We found that the challenge of WJ-MSCs with S. aureus resulted in increased internalization of S. aureus in a timedependent manner until six hours post-infection at either MOI of 10:1 and 30:1 and in increased expression of IL-6 mRNA and secretion of TNF-α at six hours and nine hours post-infection ( p< 0.05). CONCLUSIONS: These results indicate that WJ-MSCs are able to internalize S. aureus and reveal a potential important function of these cells in the immune response.

Airway remodeling is not specifically targeted by current asthma medications, partly owing to the lack of understanding of remodeling mechanisms, altogether posing great challenges in asthma treatment. Increased airway smooth muscle (ASM) mass due to hyperplasia/hypertrophy contributes significantly to overall airway remodeling and correlates with decline in lung function. Recent evidence suggests that IgE sensitization can enhance the survival and mediator release in inflammatory cells. Human ASM (HASM) cells express both low affinity (FcεRII/CD23) and high affinity IgE Fc receptors (FcεRI), and IgE can modulate the contractile and synthetic function of HASM cells. IgE was recently shown to induce HASM cell proliferation but the detailed mechanisms remain unknown. We report here that IgE sensitization induces HASM cell proliferation, as measured by 3H-thymidine, EdU incorporation, and manual cell counting. As an upstream signature component of FcεRI signaling, inhibition of spleen tyrosine kinase (Syk) abrogated the IgE-induced HASM proliferation. Further analysis of IgE-induced signaling depicted an IgE-mediated activation of Erk 1/2, p38, JNK MAPK, and Akt kinases. Lastly, lentiviral-shRNA-mediated STAT3 silencing completely abolished the IgE-mediated HASM cell proliferation. Collectively, our data provide mechanisms of a novel function of IgE which may contribute, at least in part, to airway remodeling observed in allergic asthma by directly inducing HASM cell proliferation.

Candida albicans (C. albicans) is the most prevalent fungus in the human oral cavity and has been known as the primary cause of denture-related stomatitis. Candida cells have a high adhering potential to dental material in almost the same manner as to oral tissues, and they are known to form biofilm that leads to C. albicans resistance against antifungal drugs. The aim of this study was to investigate C. albicans adhesion and growth on different restorative dental materials and studied the effect of fluoride on C. albicans growth and morphological transition. To this end, C. albicans (Sc 5314) was cultured on acrylic resins, composite resin, and glass-ionomer materials. Growth was analyzed at various times using scanning electron microscopy analyses and cell proliferation assay. The effect of different concentrations (50 and 100 ppm) of exogenous fluoride on C. albicans growth and yeast-to-hyphae transition was investigated. Scanning electron microscopy showed that C. albicans adhered to all of the tested restorative materials. Adhesion was greater on diamond D and ivocap than on composite resin and glass ionomer. After 1 to 4 days, C. albicans growth on acrylic resins was two folds that of the composite resin and the glass-ionomer. The latter also displayed the lowest adhesion and growth which may be due to the release of antimicrobial molecules such as fluoride present in this material. This hypothesis is supported by our results showing that exogenous fluoride significantly inhibits C. albicans growth and its morphological changes from blastospore to hyphal form. This study clearly demonstrates that restorative materials are conducive to C. albicans adhesion and growth. Exogenous fluoride was also shown to down-regulate C. albicans growth and morphological changes. Overall data suggest the possible integration of fluoride into dental materials which may control oral microbial pathogenesis.

Staphylococcus aureus is frequently isolated from lungs of patients with cystic fibrosis (CF). Upon lung infection with S. aureus, airway epithelial cells (AEC) produce high levels of chemokines that enhance T-cell chemotaxis. Although the number of lymphocytes is increased in the airways and bronchoalveolar lavage fluid of patients with CF, the mechanisms responsible for their accumulation and the role of S. aureus in this process are largely unknown. This study investigated early S. aureus impact on chemokine secretion by CF epithelial cells and chemotaxis of CF T cells. CF and non-CF AEC were grown in a cell culture model and apically stimulated with S. aureus. Supernatants were quantified for chemokine secretions and assayed for T-cell chemotaxis. CF AEC secreted constitutively larger amounts of IL-8, GROalpha, MIG, MIP-3beta, and MCP-1 than non-CF epithelial cells. S. aureus interaction with epithelial cells increased chemokine production by non-CF cells but had no effect on CF cells. Chemotaxis of T cells derived from patients with CF was greater than that of T cells from subjects without CF. Moreover, there were more CF T cells expressing CXCR1 as compared with non-CF T cells. Under our experimental conditions, inhibition of IL-8 or its receptor CXCR1 resulted in a considerable decrease in T-cell chemotaxis (up to 80%). These data suggest that IL-8 and its receptor CXCR1 are key players in the chemotaxis of CF T cells and could be used as targets to develop therapies for CF.

In inflammatory diseases, strong release of elastinolytic proteases results in elastin fiber degradation generating elastin peptides (EPs). Chemotactic activity for inflammatory cells was, among wide range of properties, the former identified biological activity exerted by EPs. Recently, we demonstrated the ability of EPs to favor a Th1 cytokine (IL-2, IFN-γ) cell response in lymphocytes and to regulate IL-1β expression in melanoma cells. We hypothesized that EPs might also influence inflammatory cell properties by regulating cytokine expression by these cells. Therefore, we investigated the influence of EPs on inflammatory cytokine synthesis by human monocytes. We evidenced that EPs down-regulated both at the mRNA and protein levels the proinflammatory TNF-α, IL-1β, and IL-6 expression in LPS-activated monocytes. Such negative feedback loop could be accounted solely for EP-mediated effects on proinflammatory cytokine production because EPs did not affect anti-inflammatory IL-10 or TGF-β secretion by LPS-activated monocytes. Furthermore, we demonstrated that EP effect on proinflammatory cytokine expression by LPS-stimulated monocytes could not be due either to a decrease of LPS receptor expression or to an alteration of LPS binding to its receptor. The inhibitory effects of EPs on cytokine expression were found to be mediated by receptor (spliced galactosidase) occupancy, as being suppressed by lactose, and to be associated with the decrease of NF-κB-DNA complex formation. As a whole, these results demonstrated that EP/spliced galactosidase interaction on human monocytes down-regulated NF-κB-dependent proinflammatory cytokine expression and pointed out the critical role of EPs in the regulation of inflammatory response.

Bullous pemphigoid is an inflammatory disease of the skin associated with eosinophil infiltration and the presence of high levels of Th2 cytokines in the associated blister fluid. Little is known about the contribution of chemokines in this disease. We found that eotaxin and MCP-4 mRNA and immunoreactivity were expressed in all biopsies of BP patients and were mainly localized to the epidermis and eosinophils. The expression of eotaxin and MCP-4 was enhanced in eosinophils following IL-5 treatment. Subsequent stimulation of IL-5-primed eosinophils with Ig-immune complexes, results in increase secretion of eotaxin and MCP-4 in the supernatants. Using immunostaining, these two chemokines were localized to the granules of eosinophils. BF was found to contain chemotactic activity for eosinophils, neutrophils and T cells. The chemotactic effect of BF for eosinophils was more effective when eosinophils were stimulated with IL-5 or IL-4. We also found that the levels of Th(2)-associated chemokines (eotaxin and MCP-4) in BF were significantly higher than the Th(1)-associated chemokines (MIP-1beta and IP-10). This was consistent with the increased chemotaxis of polarized Th(2) cells toward BF, when compared to Th(1)-differentiated T cells. Our results support the involvement of Th(2)-associated chemokines in the pathogenesis of BP disease.

Experimental evidence has suggested that eosinophils bearing receptors for IgE could participate in immune defense against parasites, as well as in allergic manifestations. 1' 2 The first type of IgE-binding molecule to be identified on the eosinophil membrane was the low-affinity IgE receptor FceRII/CD23. 3 However, the low level of membrane expression and CD23 mRNA in eosinophils from patients led us to investigate the existence of other IgE-binding molecules. Recently, Mac2/e-binding protein, a molecule belonging to the S-type lectin family, was detected on human eosinophils, parallel to high amounts of the corresponding mRNA. 4 Inhibition of IgEbinding and IgE-mediated cytotoxicity toward parasites by anti-Mac2 antibodies indicated the functional role of such molecules in IgE-associated diseases. A role for FceRI was first neglected because this receptor was believed to be solely expressed on mast cells and basophils. 5 The demonstration that FceRl is present on Langerhans cells in the skin 6 suggested that it might be expressed by other immune effector cells. In this report, we summarize recent data showing that FceRI is expressed on human eosinophils from patients with hypereosinophilia, 7 is involved in IgE binding to eosinophils, and participates in the selective release of one granule mediator, eosinophil peroxidase (EPO). The participation of FceRI-bearing eosinophils in a protective im

Previous studies have shown that the allergic late airway response (LR) is dependent on the leukotriene (LT) pathway in Brown Norway (BN) rats. In this same model, interleukin-2 (IL-2) has been shown to increase allergic airway responses without increasing LT production. This study examined the relationship between the upregulation of cellular immunity with IL-2 and the LT pathway in ovalbumin-sensitized BN rats. Airway responsiveness to LTD(4) was significantly increased in BN rats pretreated with IL-2 (20,000 U twice a day for 4.5 days). Treatment with montelukast, a cysteinyl LT(1) receptor antagonist, blocked IL-2's induced increase of the LR to ovalbumin challenge. When cytokine expression was assessed either by semiquantitative polymerase chain reaction or in situ hybridization, we found that montelukast decreased the amount of IL-4 mRNA expression in the lungs while increasing the amount of interferon-gamma mRNA expression 8 hours after challenge. These results indicate that upregulation of cellular immunity with IL-2 can increase the sensitivity of the airways to LTD(4) and that inhibition of the LT pathway will block the LR and modulate cytokine expression after antigen challenge.

Polymorphonuclear neutrophils (PMNs) are important effector cells in host defense and the inflammatory response to antigen. The involvement of PMNs in inflammation is mediated mainly by the Fc receptor family, including IgE receptors. Recently, PMNs were shown to express two IgE receptors (CD23/Fc epsilon RII and galectin-3). In allergic diseases, the dominant role of IgE has been mainly ascribed to its high-affinity receptor, Fc epsilon RI. We have examined the expression of Fc epsilon RI by PMNS: mRNA and cell surface expression of Fc epsilon RI alpha chain was identified on PMNs from asthmatic subjects. Furthermore, preincubation with human IgE Fc fragment blocks completely the binding of anti-Fc epsilon RI alpha chain (mAb15--1) to human PMNS: Conversely, preincubation of PMNs with mAb15--1 inhibits significantly the binding of IgE Fc fragment to PMNs, indicating that IgE bound to the cell surface of PMNs mainly via the Fc epsilon RI. Peripheral blood and bronchoalveolar lavage (BAL) PMNs from asthmatic subjects also express intracellular Fc epsilon RI alpha and beta chain immunoreactivity. Engagement of Fc epsilon RI induces the release of IL-8 by PMNS: Collectively, these observations provide new evidence that PMNs express the Fc epsilon RI and suggest that these cells may play a role in allergic inflammation through an IgE-dependent activation mechanism.

There is an emerging body of knowledge defining the role of CD8(+) cells in the pathogenesis of allergic asthma. We have previously demonstrated in sensitized Sprague-Dawley (SD) rats that depletion of CD8(+) cells caused an increase in the late airway response (LAR) and cellular infiltration after antigen challenge. To better delineate the mechanism of CD8(+) cell involvement in the development of the LAR and airway inflammation, we investigated the pattern of chemokine and cytokine production after antigen challenge. SD rats were sensitized to ovalbumin (OA) and subsequently treated with anti-CD8 (OX-8) monoclonal antibody (mAb) for the depletion of CD8(+) cells or with control mouse anti-rat IgG(1) mAb as a control procedure. After OA challenge, CD8- depleted SD rats developed an increased LAR when compared with control rats (area under the curve: 16.65 +/- 6.6 in CD8- depleted rats versus 5.39 +/- 2.0 in control animals; p0.05). Compared with the control animals, the increase in the LAR was accompanied by a significantly increased eosinophilic infiltration of the airways and was associated with increased eotaxin expression (both messenger RNA [mRNA] and protein) in the CD8-depleted group. There were no differences between the groups in RANTES or monocyte chemoattractant protein-1 (MCP-1) expression. In addition, we found a significantly lower interferon gamma (IFN-gamma) mRNA expression in the CD8-depleted rats, without any effects on interleukin (IL)-4 and IL-5 mRNA expression when measured either by semiquantitative reverse transcriptase/polymerase chain reaction (RT-PCR) or by in situ hybridization for the number of cells expressing these cytokines. Taken together, these results suggest that CD8(+) cells from sensitized SD rats exhibit the functional capacity to suppress the LAR, possibly through downregulation of eotaxin expression and increased expression of IFN-gamma mRNA.

Airway eosinophilia is a prominent feature of asthma that is believed to be mediated in part through the expression of specific chemokines such as eotaxin, a potent eosinophil chemoattractant that is highly expressed by epithelial cells and inflammatory cells in asthmatic airways. Airway smooth muscle (ASM) has been identified as a potential source of cytokines and chemokines. The aim of the present study was to examine the capacity of human ASM to express eotaxin. We demonstrate that airway myocytes constitutively express eotaxin mRNA as detected by RT-PCR. Treatment of ASM for 24 h with different concentrations of TNF-alpha and IL-1beta alone or in combination enhanced the accumulation of eotaxin transcripts. Maximal mRNA expression of eotaxin was shown at 12 and 24 h following IL-1beta and TNF-alpha stimulation, respectively. The presence of immunoreactive eotaxin was demonstrated by immunocytochemistry, and constitutive and cytokine-stimulated release of eotaxin was confirmed in ASM culture supernatants by ELISA. Strong signals for eotaxin mRNA and immunoreactivity were observed in vivo in smooth muscle in asthmatic airways. In addition, chemotaxis assays demonstrated the presence of chemoattractant activity for eosinophils and PBMCs in ASM supernatants. The chemotactic responses of eosinophils were partly inhibited with antibodies directed against eotaxin or RANTES, and a combined blockade of both chemokines causes70% inhibition of eosinophil chemotaxis. The results of this study suggest that ASM may contribute to airway inflammation in asthma through the production and release of eotaxin.

Eosinophils are not only the source of cytotoxic and proinflammatory mediators but they can also generate cytokines and growth factors, including their own factors of differentiation, namely IL-3, GM-CSF, and IL-5. Synthesis of IL-5 by eosinophils was demonstrated by in situ hybridization and immunostaining in a variety of diseases, such as coeliac disease, asthma, hypereosinophilic syndrome, or skin diseases. However, IL-5 synthesis by eosinophils was not shown in Crohn's disease, whereas in other diseases, it was restricted to a subpopulation of eosinophils, suggesting some heterogeneity in cytokine-producing eosinophils. Here, we report that human eosinophils, in addition to the synthesis of IL-5, and Th2 cytokine, can synthesize IFN gamma, a Th1 cytokine, as well as IL-10 and IL-4, known to be mainly produced by Th2 cells. Double immunostaining procedures reveal the coexpression of IL-5, IL-4, and IL-10 by the same eosinophil populations, different from IFN gamma-producing eosinophils. We propose that distinct subpopulations of human eosinophils express Th2 or Th1 cytokines. These results point to the importance of cytokines derived from non T cells in the regulation of the immune response.

Recent reports describe the beneficial use of alpha interferon (IFNalpha) for the treatment of idiopathic hypereosinophilic syndrome (HES) unresponsive to conventional therapy. A clinical improvement associated with a rapid decrease of peripheral blood eosinophilia suggested possible direct effects of IFNalpha on eosinophils through the presence of IFNalpha receptors (IFNalphaR). Reverse transcriptase- polymerase chain reaction (RT-PCR) and cytochemistry were used respectively to detect the presence and define the distribution of IFNalphaR on enriched eosinophil preparations purified from blood cells. IFNalphaR was found on eosinophils collected from patients with various eosinophilic disorders. In addition, IFNalpha inhibited the release of eosinophil granule proteins such as eosinophil cationic protein (ECP), neurotoxin (EDN, or interleukin-5 (IL-5). Moreover, antiparasite cytotoxicity was also strongly reduced in a dose-dependent manner by IFNalpha. These results provide the first evidence that human eosinophils express a functional receptor for IFNalpha and represent a potential basis for the beneficial effects of IFNalpha in patients with hypereosinophilic syndromes.

Parasitic infections are often associated with eosinophilia and high levels of immunoglobulin E (IgE). This observation has led to speculation that eosinophils and IgE may act together in the immune response against parasites. In support of this hypothesis, IgE and eosinophils participate in cytotoxic reactions directed against Schistosoma mansoni larvae in vitro. Furthermore, epidemiological studies have shown an inverse correlation between levels of specific IgE and rates of infection with Schistosoma. The low-affinity IgE receptor (Fc epsilon RII/CD23) was first incriminated in eosinophil activation. The fact that the high-affinity IgE receptor (Fc epsilon RI) is not only expressed on mast cells and basophils but also on Langerhans cells led us to investigate the presence of Fc epsilon RI on eosinophils. Here we show that Fc epsilon RI is expressed on eosinophils from hypereosinophilic patients, is involved in eosinophil degranulation, and participates in eosinophil-mediated cytotoxicity against S. mansoni. Our results indicate that Fc epsilon RI may play a major part in immune defence against parasites.

In addition to cytotoxic and proinflammatory mediators, eosinophils can produce a variety of cytokines and growth factors. Besides interleukin (IL)-5, we show in the present work that human eosinophils can synthesize interferon (IFN)-gamma and IL-10, by RT-PCR, in situ hybridization and immunostaining. Double-labelling procedures revealed the coexpression of IL-5 and IL-10 but not IL-5 and IFN-gamma, indicating the existence of subpopulations of eosinophils expressing type 1 or type 2 cytokines. IFN-alpha efficiently used for the treatment of hypereosinophilic syndromes can significantly decrease eosinophil degranulation and IL-5 release by eosinophils, through binding to a receptor for IFN-alpha. Thus, eosinophils can represent major sources of cytokines with regulatory functions, especially in allergic diseases and parasitic infections.

Several reports suggest that activated airway smooth muscle (ASM) cells are capable of generating various proinflammatory mediators, including cytokines and chemokines. However, little is known about the mechanism involved in this process. In this regard, we have examined the expression and the role of the high affinity IgE receptor (Fc epsilonRI) by ASM cells. Human ASM cells were found to constitutively express transcripts coding for alpha, beta, and gamma subunits of Fc epsilonRI. Flow cytometry and Western blot analysis confirmed the expression of Fc epsilonRI alpha-chain protein. Interestingly, Fc epsilonRI alpha-chain immunoreactivity was also demonstrated in smooth muscle within bronchial biopsies of asthmatic subjects. Cross-linking of Fc epsilonRI induced mobilization of free calcium in ASM cells, one of the critical signals to trigger smooth muscle contraction. Furthermore, cultured ASM cells released IL-4, IL-13, IL-5, and eotaxin but not IFN-gamma, when sensitized with IgE followed by anti-IgE Ab cross-linking. The addition of anti-Fc epsilonRI alpha-chain Abs directed against IgE binding site inhibited this release. Taken together, these results suggest a potential new and important mechanism by which ASM cells may participate in airway inflammation and bronchoconstriction associated with allergic asthma.

The existence of a functional receptor for secretory component (SC) on the eosinophil membrane might explain the preferential degranulation induced by secretory IgA (sIgA) when compared to serum IgA. Indeed, flow cytometry analysis revealed that purified human SC could bind to a subpopulation (4-59%) of blood eosinophils purified from 19 patients with eosinophilia. Binding of radiolabeled human SC could be competitively inhibited using unlabeled SC or secretory IgA but not with serum IgA or IgG. Immunoprecipitation and immunosorbent chromatography using human SC revealed the presence of a major component at 15 kDa in eosinophil extracts as well as in culture supernatants but not in neutrophils. The 15-kDa protein eluted from the human SC immunosorbent was able to bind to SC or to sIgA but not to serum IgA. Eosinophils preincubated with human SC or sIgA released eosinophil cationic protein (ECP) and eosinophil peroxidase (EPO) after addition of anti-SC or anti-IgA monoclonal antibody as respective cross-linking reagents. These results indicated that binding of free or complexed SC to human eosinophils could induce eosinophil degranulation. Furthermore, the dose-dependent inhibition by SC of mediator release induced by sIgA but not by serum IgA, suggested that the receptor for SC could be involved in the preferential degranulation mediated by sIgA. These results indicate a novel pathway of eosinophil activation and its potential involvement in mucosal immunity, particularly in inflammatory diseases associated with infiltration of eosinophils and the enhanced production of sIgA.