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PubMed1993

Human eosinophils can express the cytokines tumor necrosis factor-alpha and macrophage inflammatory protein-1 alpha.

By in situ hybridization, 44-100% of the blood eosinophils from five patients with hypereosinophilia and four normal subjects exhibited intense hybridization signals for TNF-alpha mRNA. TNF-alpha protein was detectable by immunohistochemistry in blood eosinophils of hypereosinophilic subjects, and purified blood eosinophils from three atopic donors exhibited cycloheximide-inhibitable spontaneous release of TNF-alpha in vitro. Many blood eosinophils (39-91%) from hypereosinophilic donors exhibited intense labeling for macrophage inflammatory protein-1 alpha (MIP-1 alpha) mRNA, whereas eosinophils of normal donors demonstrated only weak or undetectable hybridization signals for MIP-1 alpha mRNA. Most tissue eosinophils infiltrating nasal polyps were strongly positive for both TNF-alpha and MIP-1 alpha mRNA. By Northern blot analysis, highly enriched blood eosinophils from a patient with the idiopathic hypereosinophilic syndrome exhibited differential expression of TNF-alpha and MIP-1 alpha mRNA. These findings indicate that human eosinophils represent a potential source of TNF-alpha and MIP-1 alpha, that levels of expression of mRNA for both cytokines are high in the blood eosinophils of hypereosinophilic donors and in eosinophils infiltrating nasal polyps, that the eosinophils of normal subjects express higher levels of TNF-alpha than MIP-1 alpha mRNA, and that eosinophils purified from the blood of atopic donors can release TNF-alpha in vitro.

Cells, Cultured↗
PubMed2004

Characteristics of eosinophils migrating around fungal hyphae in nasal discharge.

It is known that eosinophil granular proteins cause tissue damage. To explore how eosinophils degranulate, we studied the degranulation of eosinophils that had migrated around fungal hyphae. In electron microscopic observations of allergic mucin from patients with allergic fungal sinusitis, fungal hyphae were detected, surrounded by numerous eosinophils. A number of eosinophils, including many disintegrated eosinophils, adhered to the cuticular layer of the hyphal surface. Although the fungal hyphae were detected in allergic mucins in all 5 patients, fungal hyphae surrounded by eosinophils were observed in only 1 patient. In the eosinophil cytoplasm, the cell membrane was invaginated, deep, and sheetlike, and the space formed by its infolding was filled with a highly electron-dense substance. This substance appeared to be a mixture of the cuticular substance of the hyphal surface and granular proteins. Thus, the eosinophil phagocytosed the cuticular substance of the hyphae into a sheetlike invaginated space, and released granular proteins into that space. The structure invaginated in the cytoplasm retained its form even after disintegration of the eosinophil, and adhered to the cuticular layer. This structure detected in eosinophils has not been reported previously, and is considered to be an interesting finding from the viewpoint of the function of eosinophils.

Cell Adhesion↗
PubMed1993

Bronchoalveolar eosinophilic cells in a canine model of asthma: two distinctive populations.

In a canine model of asthma, the identity and relationships of two types of cells with eosinophilic granules found in bronchoalveolar lavage (BAL) were investigated. Five female and three male newborn mongrel dogs were sensitized by intraperitoneal injection of 500 micrograms ragweed in 30 mg aluminum hydroxide repeated weekly for 8 weeks and then biweekly until 16 weeks of age. Three female and two male littermate controls received 30 mg aluminum hydroxide. From 4 months of age, dogs received six breaths of wet-nebulized ragweed solution through an endotracheal tube bimonthly. Bronchoalveolar lavage was performed 2 months or more after the last exposure to ragweed and was repeated 5 hours after ragweed inhalation. Dogs studied were 15 to 38 months of age. Eosinophilic cell populations were determined in BAL and peripheral blood; cells were examined by light and electron microscopy. Two eosinophilic cell types were in BAL: an eosinophil and an atypical cell with globule leukocyte characteristics. Specific microgranules, a constant feature of eosinophils, were prominent in the atypical cell and suggest a relationship to the eosinophil granulocyte series for the atypical cell and cells identified by others as globule leukocytes. In ragweed sensitized animals, there were more eosinophilic cells in BAL fluid and more eosinophils in peripheral blood. There was a proportional increase in BAL eosinophils 5 hours after ragweed inhalation and a corresponding decline in peripheral blood eosinophils. There was no increase in numbers of eosinophils in the sensitized animals; eosinophil and atypical cell numbers in littermate controls were unchanged.

Allergens↗
PubMed1995

Relationships between eosinophil-associated parameters and disease severity in atopic dermatitis.

This study demonstrated not only the presence of house dust mite antigen (HDM)-specific IgE and IgG antibodies in the sera of 27 patients with atopic dermatitis (AD), but also considerably increased levels of serum eosinophil cationic protein (ECP) and uniformly nadir levels of plasma ECP, which provides evidence for ECP release from eosinophils in vitro. Besides blood eosinophilia, AD patients had an increased number of CD32+ eosinophils and EG2+ "activated" eosinophils; both of which, especially the latter, were significantly correlated with elevated levels of serum ECP. AD-source eosinophils had enhanced sensitivity for IgG-immune complexes corresponding to their increased expression of CD32, whereas neither HDM alone nor IgE/HDM-immune complex induced eosinophil activation. There were no gamma globulin-bearing cells in AD-source eosinophils, indicating that serum ECP levels may be dependent on EG2+ eosinophil numbers rather than the state of CD32-mediated eosinophil degranulation, as shown in vitro. Since HDM can cause an eczematous skin reaction accompanying the eosinophil accumulation and degranulation in sensitized AD, increased numbers of both CD32+ and EG2+ eosinophils according to the extent of AD severity indicates that these eosinophil in vivo generations and their subsequent functioning in the lesional skin may be regulated through HDM-initiated dermatitis events, and then, may form a vicious circle as the AD lesions spread.

Adult↗
PubMed2002

TNF-alpha potentiates C5a-stimulated eosinophil adhesion to human bronchial epithelial cells: a role for alpha 5 beta 1 integrin.

Cooperative action of inflammatory mediators and adhesion molecules orchestrates eosinophil recruitment during allergic inflammation in the airways. This study investigated the mechanisms involved in increasing eosinophil adhesion to human bronchial epithelial cells (HBEC) following priming and activation of eosinophils with TNF-alpha and complement protein C5a, respectively. Under primed conditions, eosinophil adhesion increased 3-fold from basal (16%), and the effect was significantly greater (p < 0.05) than the increase following stimulation with C5a alone (2-fold). Eosinophil contact with HBEC was essential for priming. In contrast to C5a, adhesion of eotaxin-stimulated eosinophils to HBEC was not primed with TNF-alpha nor IL-5, a known eosinophil-priming agent. Priming caused activation of alpha(M)beta(2) integrin; mAb against either the common beta(2) integrin subunit or its ICAM-1 ligand reduced the primed component of adhesion. Using mAbs against beta(1) or alpha(5), but not alpha(4) integrin subunit, together with anti-beta(2) integrin mAb, reduced stimulated adhesion to basal levels. Cross-linking alpha(5)beta(1) integrin increased alpha(M)beta(2) integrin-dependent adhesion of eosinophils. There are no known adhesion molecule ligands of alpha(5)beta(1) integrin expressed on HBEC; however, fibronectin, the major matrix protein ligand for alpha(5)beta(1) integrin, was detected in association with HBEC monolayers. A mAb against fibronectin, in combination with anti-beta(2) integrin mAb, reduced adhesion to basal levels. In conclusion, alpha(5)beta(1) integrin may provide a contact-dependent costimulus for eosinophil priming that, together with TNF-alpha, potentiated C5a activation of alpha(M)beta(2) integrin and increased eosinophil adhesion to ICAM-1. Fibronectin, associated with HBEC, may act as a ligand for alpha(5)beta(1) integrin. Dual regulation of eosinophil priming may prevent inappropriate activation of eosinophils in the circulation.

Adjuvants, Immunologic↗
PubMed2002

Biological activities of ecalectin: a novel eosinophil-activating factor.

Ecalectin, produced by Ag-stimulated T lymphocytes, is a potent eosinophil-specific chemoattractant in vitro as well as in vivo and thus is implicated in allergic responses. Ecalectin differs structurally from other known eosinophil chemoattractants (ECAs); ecalectin belongs to the galectin family defined by their affinity for beta-galactosides and by their conserved carbohydrate recognition domains. These characteristic features suggest that ecalectin has unique activities associated with allergic inflammation besides ECA activity. Conversely, ecalectin may mediate ECA activity by binding to a receptor of a known ECA via affinity for the beta-galactosides present on this receptor. In this study, we have tested whether ecalectin mediates ECA activity by binding to a receptor of a known ECA, and we have assessed its effects on eosinophils. Ecalectin did not mediate ECA activity by binding to the IL-5R or to CCR3. Also, the ECA activity of ecalectin was mainly chemokinetic. In addition, ecalectin induced concentration-dependent eosinophil aggregation, a marker for eosinophil activation. Ecalectin induced concentration-dependent superoxide production from eosinophils but did not induce degranulation; usually these two events are coupled in eosinophil activation. Moreover, ecalectin directly prolonged eosinophil survival in vitro and did not trigger eosinophils to secrete cytokines that prolong eosinophil survival. These results demonstrate that ecalectin has several unique effects on eosinophils. Therefore, we conclude that ecalectin is a novel eosinophil-activating factor. Presumably, these effects allow ecalectin to play a distinctive role in allergic inflammation.

Antibodies↗
PubMed2003

Expression and production of the CXC chemokine growth-related oncogene-alpha by human eosinophils.

Eosinophils are seen together with neutrophils at sites of inflammation. However, their roles are not clear. In addition, eosinophils infiltrate tumor tissue in some neoplastic diseases. In this study, we show that large amounts of the neutrophil-activating CXC chemokine growth-related oncogene (GRO)-alpha can be produced by human eosinophils. Eosinophils showed presence of preformed GRO-alpha in the crystalloid-containing specific granules (190 pg/2 x 10(6) cells). During incubation, a strong increase in GRO-alpha gene expression was seen. At a low cell density, addition of TNF-alpha or IL-1 beta increased the production of GRO-alpha in eosinophils, which was not the case at a higher cell density. Eosinophils can produce TNF-alpha themselves, and neutralizing Abs against TNF-alpha significantly inhibited GRO-alpha production. This suggests that autocrine and paracrine effects from TNF-alpha can be important when up-regulating GRO-alpha gene expression. In contrast, IFN-gamma, a prototypic Th1-cytokine, down-regulated expression of GRO-alpha. This may be important during resolution of inflammation but also suggests different roles for eosinophils depending on the inflammatory context. Tumor-infiltrating eosinophils in Hodgkin's disease of the nodular sclerosing type are associated with a poor prognosis. Eosinophils from such tumor tissue showed an abundant expression of GRO-alpha. The GRO-alpha receptor CXCR2 was also detected in tumor tissue, proposing interactions between eosinophils and the tumor. Our findings suggest that eosinophils can promote inflammation through recruitment of CXCR2-bearing cells. In addition, this feature of the eosinophils indicates a role for these cells in the biology of certain tumors.

Autocrine Communication↗
PubMed2005

Nonpathogenic, environmental fungi induce activation and degranulation of human eosinophils.

Eosinophils and their products are probably important in the pathophysiology of allergic diseases, such as bronchial asthma, and in host immunity to certain organisms. An association between environmental fungal exposure and asthma has been long recognized clinically. Although products of microorganisms (e.g., lipopolysaccharides) directly activate certain inflammatory cells (e.g., macrophages), the mechanism(s) that triggers eosinophil degranulation is unknown. In this study we investigated whether human eosinophils have an innate immune response to certain fungal organisms. We incubated human eosinophils with extracts from seven environmental airborne fungi (Alternaria alternata, Aspergillus versicolor, Bipolaris sorokiniana, Candida albicans, Cladosporium herbarum, Curvularia spicifera, and Penicillium notatum). Alternaria and Penicillium induced calcium-dependent exocytosis (e.g., eosinophil-derived neurotoxin release) in eosinophils from normal individuals. Alternaria also strongly induced other activation events in eosinophils, including increases in intracellular calcium concentration, cell surface expression of CD63 and CD11b, and production of IL-8. Other fungi did not induce eosinophil degranulation, and Alternaria did not induce neutrophil activation, suggesting specificity for fungal species and cell type. The Alternaria-induced eosinophil degranulation was pertussis toxin sensitive and desensitized by preincubating cells with G protein-coupled receptor agonists, platelet-activating factor, or FMLP. The eosinophil-stimulating activity in Alternaria extract was highly heat labile and had an M(r) of approximately 60 kDa. Thus, eosinophils, but not neutrophils, possess G protein-dependent cellular activation machinery that directly responds to an Alternaria protein product(s). This innate response by eosinophils to certain environmental fungi may be important in host defense and in the exacerbation of inflammation in asthma and allergic diseases.

Alternaria↗
PubMed1996

Corticosteroid-induced apoptosis of eosinophils in atopic dermatitis patients.

We examined the mechanism by which steroid administration significantly decreases the high eosinophil cell count in peripheral blood in patients with atopic dermatitis (AD). Eosinophils were isolated from the peripheral blood of patients with moderate or severe adult AD, and cultured. After steroid was added to the culture medium, we examined the changes in eosinophils, i.e., 1) the survival rates, 2) morphological changes and 3) fragmentation of DNA with respect to 2 factors, the steroid concentration and culture time. The steroid or interleukin-5 (IL-5) were added to eosinophils from patients with AD and those from non-AD patients with eosinophilia to compare serial changes in the survival rate. In eosinophils from patients with AD, the survival rate significantly decreased time-dependently and steroid concentration-dependently. Steroid administration significantly inhibited the survival rate of eosinophils from patients with AD compared to the survival rates of monocytes and neutrophils. The nuclei of eosinophils were serially reduced, and disappeared 72 hours after steroid administration. Simultaneously, cell size decreased, although the cell membrane remained intact. Granules developed in the cell membrane. In the steroid-treated group, apoptotic cells appeared earlier than in the untreated group. The number of cells showing apoptosis was increased steroid concentration-dependently. The number of DNA ladders was increased time-dependently and steroid concentration-dependently. In eosinophils derived from patients with AD and those derived from non-AD patients with eosinophilia, treatment with recombinant human (rh) IL-5 prolonged the life-span of cells. However, there were differences in the survival rates. In the presence of rhIL-5, the eosinophils from non-AD patients survived 1.4 times longer than those from AD patients at 24 hours (P < 0.05). In the presence of steroid, the eosinophils from non-AD patients survived twice as long as those from AD patients at 24 hours (P < 0.01). These findings suggest that apoptosis induced by steroids decreases the eosinophil count in vivo in patients with AD. There may be a difference in the incidence of steroid-induced apoptosis between eosinophil cells from patients with AD and those from patients with eosinophilia due to other underlying diseases.

Adolescent↗
PubMed1999

Endogenous platelet-activating factor is critically involved in effector functions of eosinophils stimulated with IL-5 or IgG.

Eosinophil activation and subsequent release of inflammatory mediators are implicated in the pathophysiology of allergic diseases. Eosinophils are activated by various classes of secretagogues, such as cytokines (e.g., IL-5), lipid mediators (e.g., platelet-activating factor (PAF)), and Ig (e.g., immobilized IgG). However, do these agonists act directly on eosinophils or indirectly through the generation of intermediate active metabolites? We now report that endogenous PAF produced by activated eosinophils plays a critical role in eosinophil functions. Human eosinophils produced superoxide when stimulated with immobilized IgG, soluble IL-5, or PAF. Pretreating eosinophils with pertussis toxin abolished their responses to these stimuli, suggesting involvement of a metabolite(s) that acts on G proteins. Indeed, PAF was detected in supernatants from eosinophils stimulated with IgG or IL-5. Furthermore, structurally distinct PAF antagonists, including CV6209, hexanolamine PAF, and Y-24180 (israpafant), inhibited IgG- or IL-5-induced superoxide production and degranulation. Previous reports indicated that exogenous PAF stimulates eosinophil eicosanoid production through formation of lipid bodies. We found in this study that IgG or IL-5 also induces lipid body formation and subsequent leukotriene C4 production mediated by endogenous PAF. Finally, inhibition of cytosolic phospholipase A2, one of the key enzymes involved in PAF synthesis, attenuated both PAF production and effector functions of eosinophils. These findings suggest that endogenous PAF plays important roles in eosinophil functional responses to various exogenous stimuli, such as cytokines and Igs. Therefore, inhibition of PAF synthesis or action may be beneficial for the treatment of eosinophilic inflammation.

Eicosanoids↗
PubMed1999

Ligation of Fc gamma RII (CD32) pivotally regulates survival of human eosinophils.

The low-affinity IgG Fc receptor, FcgammaRII (CD32), mediates various effector functions of lymphoid and myeloid cells and is the major IgG Fc receptor expressed by human eosinophils. We investigated whether FcgammaRII regulates both cell survival and death of human eosinophils. When cultured in vitro without growth factors, most eosinophils undergo apoptosis within 96 h. Ligation of FcgammaRII by anti-CD32 mAb in solution inhibited eosinophil apoptosis and prolonged survival in the absence of growth factors. Cross-linking of human IgG bound to FcgammaRII by anti-human IgG Ab or of unoccupied FcgammaRII by aggregated human IgG also prolonged eosinophil survival. The enhanced survival with anti-CD32 mAb was inhibited by anti-granulocyte-macrophage-CSF (GM-CSF) mAb, suggesting that autocrine production of GM-CSF by eosinophils mediated survival. In fact, mRNA for GM-CSF was detected in eosinophils cultured with anti-CD32 mAb. In contrast to mAb or ligands in solution, anti-CD32 mAb or human IgG, when immobilized onto tissue culture plates, facilitated eosinophil cell death even in the presence of IL-5. Cell death induced by these immobilized ligands was accompanied by DNA fragmentation and was inhibited when eosinophil beta2 integrin was blocked by anti-CD18 mAb, suggesting that beta2 integrins play a key role in initiating eosinophil apoptosis. Thus, FcgammaRII may pivotally regulate both survival and death of eosinophils, depending on the manner of receptor ligation and beta2 integrin involvement. Moreover, the FcgammaRII could provide a novel mechanism to control the number of eosinophils at inflammation sites in human diseases.

Antibodies, Monoclonal↗
PubMed2000

Peyer's patch eosinophils: identification, characterization, and regulation by mucosal allergen exposure, interleukin-5, and eotaxin.

The gastrointestinal immune system is traditionally thought to be composed of lymphocytes located within Peyer's patches and the lamina propria. We have recently reported that eosinophils also reside in the gastrointestinal tract during healthy states, in particular, within the lamina propria, and that these cells substantially increase after oral allergen exposure. We now demonstrate the presence of eosinophils in Peyer's patches and characterize the signals that regulate the accumulation of eosinophils in Peyer's patches. In contrast to the lamina propria, intestinal Peyer's patches have very low levels of eosinophils under healthy states. However, elevated levels of interleukin-5 (IL-5), generated by transgenic or pharmacologic approaches, result in a dramatic increase in eosinophil levels in Peyer's patches. Most eosinophils are located in the outer cortex and interfollicular regions of the Peyer's patches. To dissect the mechanism of eosinophil trafficking to Peyer's patches, the role of eotaxin was examined. Mice transgenic for IL-5 and genetically deficient in eotaxin were found to have reduced levels of eosinophils in Peyer's patches compared with IL-5-transgenic mice. To prove that eosinophils also traffic to Peyer's patches in wild-type mice, allergic hypersensitivity was induced and Peyer's patches were examined. Exposure to mucosal allergen promoted marked accumulation of eosinophils in Peyer's patches and this process was attenuated in eotaxin-deficient mice. In summary, these data demonstrate that elevated levels of IL-5 and mucosal allergen exposure promote eotaxin-dependent eosinophil trafficking to Peyer's patches. These studies suggest that eosinophils may cooperate with lymphocytes in the development of mucosal immune responses in the gastrointestinal tract. (Blood. 2000;96:1538-1544)

Animals↗
PubMed1992

IFN-gamma induces expression of Fc gamma RIII (CD16) on human eosinophils.

The extent to which eosinophils constitutively express FcRIII (CD16) is controversial. We were unable to detect this receptor on freshly isolated, peripheral blood eosinophils. The capacity of eosinophils to change their Fc gamma R expression in vitro has not been previously demonstrated. Culture with IFN-gamma for 1 to 2 days induced FcRIII expression on eosinophils. This effect was dose-dependent and significant at concentrations of 100 U/ml IFN-gamma and above. Expression of FcRI (CD64) and FcRII (CDw32) was also upregulated. These increases were inhibited by cycloheximide (10(-6) M), suggesting a requirement for protein synthesis, and dexamethasone (10(-6) M). Northern blot analysis demonstrated the presence of FcRIII mRNA in eosinophils cultured with IFN-gamma for 2 days but not in unstimulated eosinophils. By contrast, culture with IL-3 caused an up-regulation of eosinophil FcRII expression but did not induce expression of FcRI or FcRIII. The FcRIII expressed by eosinophils after IFN-gamma stimulation was functionally active, as shown by the triggering of eosinophil membrane depolarization and LTC4 generation by an anti-CD16 mAb. Treatment of IFN-gamma-stimulated eosinophils with phosphatidylinositol-specific phospholipase C reduced FcRIII expression, suggesting that, like neutrophils, eosinophils express the phosphatidylinositol glycan-linked form of this receptor. Therefore, this study demonstrates that IFN-gamma-treated eosinophils express a functionally active, phosphatidylinositol glycan-anchored form of FcRIII.

Antigens, Differentiation↗
PubMed1992

Mature eosinophils stimulated to develop in human-cord blood mononuclear cell cultures supplemented with recombinant human interleukin-5. II. Vesicular transport of specific granule matrix peroxidase, a mechanism for effecting piecemeal degranulation.

The mechanism of piecemeal degranulation by human eosinophils was investigated. Mature eosinophils that developed in rhIL-5-containing conditioned media from cultured human cord blood mononuclear cells were prepared for ultrastructural studies using a combined technique to image eosinophil peroxidase by cytochemistry in the same sections on which postembedding immunogold was used to demonstrate Charcot-Leyden crystal protein. Vesicular transport of eosinophil peroxidase from the specific granule matrix compartment to the cell surface was associated with piecemeal degranulation. This process involved budding of eosinophil peroxidase-loaded vesicles and tubules from specific granules. Some eosinophil peroxidase that was released from eosinophils remained bound to the cell surface; some was free among the cultured cells. Macrophages and basophils bound the released eosinophil peroxidase to their plasma membranes, internalized it in endocytotic vesicles, and stored it in their respective phagolysosomes and secretory granules. Charcot-Leyden crystal protein was diffusely present in the nucleus and cytoplasm of IL-5-stimulated mature eosinophils. Extensive amounts were generally present in granule-poor and subplasma membrane areas of the cytoplasm in contrast to eosinophil peroxidase, which was secreted and bound to the external surface of eosinophil plasma membranes. These studies establish vesicular transport as a mechanism for emptying the specific eosinophil granule matrix compartment during IL-5-associated piecemeal degranulation.

Blood Cells↗
PubMed1992

Mast cell mediators prostaglandin-D2 and histamine activate human eosinophils.

Airway damage secondary to eosinophil activation is thought to contribute to the development of asthma. Using the fluorescent dye FURA-2 to measure the concentration of cytosolic calcium, we found that supernatants from anti-IgE-stimulated human lung mast cells increased cytosolic calcium in human eosinophils. We then examined the major mast cell mediators (histamine, PGD2, platelet-activating factor (PAF), eosinophil chemotactic factor of anaphylaxis (ECF-A), leukotriene (LT)C4 and LTB4) for their ability to increase cytosolic calcium in eosinophils. We found that both PAF (5 x 10(-9) to 5 x 10(-6) M) and PGD2 (two of five donors responsive at 1 x 10(-9) M) were potent stimuli for calcium mobilization. LTB4 (10(-8), 10(-7) M) and histamine were also active, although higher concentrations of histamine were required to see a response (3 x 10(-7) to 10(-5) M). LTC4, val-ECF-A, and ala-ECF-A were inactive. The effects of PGD2 and histamine were specific for eosinophils, although LTB4 and PAF increased calcium in both neutrophils and eosinophils. The histamine-induced increase in intracellular calcium was not blocked by the H1 or H2 antagonists pyrilamine or cimetidine (10(-4) M), respectively; however, the response to 10(-6) M histamine was completely blocked by the specific H3 antagonist thioperamide (10(-6) M). To evaluate the relative contribution of these stimulatory mast cell mediators on the calcium mobilizing activity in supernatants from anti-IgE-stimulated human lung mast cell (HLMC), we examined the effect of supernatants from HLMC pretreated with indomethacin and/or the 5-lipoxygenase pathway inhibitor MK886. These supernatants were added to FURA-2-loaded eosinophils that had been preincubated with thioperamide and/or the PAF antagonist WEB-2086. We found that the increase in eosinophil calcium in response to supernatants from anti-IgE-stimulated-HLMC was totally inhibited only when the mast cells were challenged in the presence of indomethacin and MK886, and the eosinophils were preincubated with thioperamide. WEB-2086 had little effect. When we examined the effect of these mediators on eosinophil secretory function, we found that PGD2 (not histamine) primed eosinophils for enhanced release of LTC4 in response to the calcium ionophore A23187. We conclude that the activation of eosinophils by PGD2 and other mast cell products may contribute to airways inflammation that is characteristic of asthma.

Azepines↗
PubMed2005

The involvement of adhesion molecules and lipid mediators in the adhesion of human platelets to eosinophils.

Platelet-leukocyte interactions represent an important determinant of the inflammatory response. Although mechanisms of platelet-neutrophil adhesion were studied extensively, little is known on the mechanisms of platelet-eosinophil interactions. The aim of the present study was to analyze the involvement of adhesion molecules and lipid mediators in platelet-eosinophil adhesion as compared to platelet-neutrophil adhesion. For that purpose human platelets, eosinophils and neutrophils were isolated and platelet-eosinophil and platelet-neutrophil adhesion induced by thrombin (30 mU/ml), LPS (0.01 microg/ml) and fMLP (1 microM) was quantified using the "rosettes" assay. The involvement of adhesion molecules such as selectin P, glycoprotein IIb/IIIa (GPIIb/IIIa) and lipid mediators such as of thromboxane A2 (TXA2), platelet activating factor (PAF) and cysteinyl leukotrienes (cysLTs) were studied using monoclonal antibodies and pharmacological inhibitors, respectively. Thrombin (30 mU/ml), LPS (0.01 microg/ml) and fMLP (1 microM) each of them induced platelet-eosinophil adhesion that was even more pronounced as compared with platelet-neutrophil adhesion induced by the same stimulus. Anti-CD62P antibody (1 microg/ml) and anti-GP IIb/IIIa antibody (abciximab-3 microg/ml) strongly inhibited platelet-eosinophil as well as platelet-neutrophil adhesion. Aspirin inhibited platelet-eosinophil adhesion, while MK 886-a FLAP inhibitor (10 microM), or WEB 2170-a PAF receptor antagonist (100 microM) were less active. On the other hand aspirin, MK 886 and WEB 2170 all three of them inhibited platelet-neutrophil adhesion. In summary, platelets adhered avidly to eosinophils both after activation of platelets by thrombin, eosinophils by fMLP or simultaneous activation of platelets and eosinophils by LPS. Similarly to platelet-neutrophil interaction adhesion of platelets to eosinophils involved not only adhesion molecules (selectin P, GPIIb/IIIa), but also lipid mediators such as TXA2. The involvement of PAF and cysteinyl leukotrienes in platelet-eosinophil adhesion was less pronounced as compared to platelet-neutrophil adhesion.

Abciximab↗
PubMed1977

Histamine modulation of eosinophil migration.

Preincubation of eosinophils with 10(-5) M or higher concentrations of histamine inhibited the eosinophil chemotactic response to endotoxin-activated serum whether by using the nucleopore filter assay and counting the cells migrating through the filter, or by using the Zigmond-Hirsch assay and counting the cells at each 10-mum interval. When the H2-receptor sites on the eosinophils were blocked by metiamide, the inhibitory capacity of histamine was prevented. Preincubation of eosinophils with 10(-6) M histamine increased the number of responding eosinophils to endotoxin-activated serum and this enhancement was blocked by an H1-receptor antagonist. Isoproteronol and aminophylline inhibited eosinophil movement and increasing concentrations of dibutryl cyclic AMP inhibited eosinophil migration. Concentrations of histamine that consistently resulted in inhibition of eosinophil movement stimulated an increase in cyclic AMP that was prevented by blocking the H2-receptor but not the H1-receptor. Thus, histamine-dependent inhibition of the eosinophil chemotactic response to other agents is mediated through the H2-receptor and is associated with an increase in the intracellular level of cyclic AMP whereas histamine dependent enhancement of eosinophil migration to other agents appears to be mediated through the H1-receptor. Eosinophils behave as a heterogeneous population as assessed by the ability of histamine to augment or inhibit cell migration. This may reflect differences in H1 to H2 receptor density or cell responsiveness to receptor stimulation. The chemoattractant activity of histamine itself is not influenced by H1 or H2 receptor antagonists, thus it is possible that an eosinophil has a third type of histamine receptor.

Bucladesine↗
PubMed1990

IL-5 enhances the in vitro adhesion of human eosinophils, but not neutrophils, in a leucocyte integrin (CD11/18)-dependent manner.

In an attempt to explain the preferential accumulation of eosinophils at sites of allergic tissue reactions, we have studied the effects of interleukin-5 (IL-5) on the adherence of human eosinophils and neutrophils to plasma-coated glass (PCG) or human microvascular endothelial cells (HMVEC). IL-5 was compared with IL-3, granulocyte-macrophage colony-stimulating factor (GM-CSF) and platelet-activating factor (PAF), since all these agents have biological properties associated with eosinophil activation and/or survival in vitro. IL-5, IL-3 and GM-CSF induced a time-dependent increase in adherence of normal density eosinophils to PCG optimal at 60 min, whereas the effect of PAF was greater at 15 min. Similar results were obtained with neutrophils, with the exception that IL-5 had minimal and non-significant effects on this cell type. Unstimulated eosinophils and neutrophils also adhered to PCG or HMVEC, but in low numbers. Preincubation of eosinophils with IL-5, GM-CSF or PAF resulted in dose-dependent increases in the numbers of adherent cells to PCG. IL-3 had a smaller but significant effect on enhanced eosinophil adhesion to PCG, while IL-2 and lyso-PAF were ineffective. Neutrophils gave similar levels of baseline and stimulated adhesion to PCG as eosinophils, IL-5 again had no significant stimulatory effect. IL-5 also increased eosinophil, but not neutrophil, adherence to HMVEC in a concentration-dependent manner. Preincubation with the protein synthesis inhibitor cycloheximide had no effect on IL-5-, GM-CSF- or PAF-stimulated eosinophil adhesion. The contribution of the CD11/18 leucocyte integrins to IL-5- and PAF-induced eosinophil hyperadherence was investigated by inhibition experiments utilizing monoclonal antibodies (mAb). Enhanced adhesion to PCG (by PAF) or HMVEC (by IL-5) was inhibited by (ranked in order of potency) anti-CR3 alpha = common beta-chain greater than LFA-1 alpha. Anti-p150,95 alpha had no measurable effect. Baseline adhesion by unstimulated eosinophils was not significantly influenced by prior incubation with these mAb. Using flow cytometry, IL-5 and IL-3 were found to up-regulate cosinophil but not neutrophil CR3 expression. These findings demonstrate that IL-5 enhances cosinophil, but not neutrophil, adherence reactions, by a mechanism dependent, at least in part, on the CD11/18 family of adhesion glycoproteins.

Cell Adhesion↗