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PubMed2002

Respiratory syncytial virus enhances the expression of CD11b molecules and the generation of superoxide anion by human eosinophils primed with platelet-activating factor.

BACKGROUND: Human respiratory syncytial virus (RSV) infection in infancy and early childhood causes acute bronchiolitis and exacerbates bronchial asthma. Eosinophil infiltration may contribute to airway obstruction in RSV infection. OBJECTIVE: We hypothesized that RSV affects eosinophil function. METHODS: Eosinophil activation was evaluated by chemiluminescent detection of superoxide anion (O(2)(-)) generation. Expression of CD11b on eosinophils was determined by flow cytometry. RESULTS: Although RSV did not induce O(2)(-) generation by resting eosinophils, RSV enhanced O(2)(-) generation of eosinophils primed with platelet-activating factor (PAF). Enhancement was significantly inhibited by either continuous agitation to prevent eosinophil adhesion to test tube surfaces or by pretreating cells with anti-CD18 antibody, suggesting that the stimulatory effects of RSV on eosinophils depend on cell adhesion via beta(2)-integrins. In fact, RSV enhanced PAF-induced CD11b expression by eosinophils. CONCLUSIONS These findings suggest that RSV enhances eosinophil CD11b expression and O(2)(-) generation induced by PAF. Thus, RSV infection may exacerbate airway inflammation by enhancing mediator release from eosinophils.

Eosinophils↗
PubMed2002

Effect of extracellular matrix proteins on platelet-activating factor-induced eosinophil chemotaxis.

BACKGROUND: Eosinophils accumulate in tissues during various allergic inflammatory processes. Selective eosinophil recruitment is the result of orchestrated events involving cell adhesion molecules and chemoattractants. However, the exact mechanism of the regulation of integrin avidity during interstitial migration is poorly understood. OBJECTIVE: The purpose of this study was to investigate whether extracellular matrix proteins might activate human eosinophil chemotaxis and, if so, to clarify the mechanism in terms of integrin avidity. METHOD: Eosinophils were purified from the peripheral blood of healthy donors. Eosinophil migration was measured using Boyden chambers with filter membranes coated with fibronectin (Fn), vitronectin (Vn), laminin (Ln), hyaluronic acid, collagen type IV, or bovine serum albumin (BSA) overnight. Platelet-activating factor (PAF) was introduced into the lower chamber of each well. Eosinophils were placed in the upper chamber after incubation with IL-5 for 15 min. The number of eosinophils that transmigrated into the lower chamber was calculated by measuring the eosinophil peroxidase activity. RESULTS: Fn, Ln and BSA enhanced PAF-induced chemotaxis of eosinophils. Inhibition experiments using blocking monoclonal antibodies showed that in the early phase of chemotaxis, Fn and Ln facilitated eosinophil chemotaxis that was mediated by alpha4 and alpha6 integrins, respectively. In the late phase of chemotaxis, BSA, but not other matrix proteins, facilitated both chemokinesis and chemotaxis that was mediated by beta2 integrin. CONCLUSION: Our data strongly suggest that during chemotaxis, matrix proteins might activate eosinophils via binding with integrins to facilitate PAF-induced chemotaxis, and that such a mechanism might participate in allergic inflammatory processes.

Animals↗
PubMed2005

Effect of the cysteinyl leukotriene antagonist pranlukast on transendothelial migration of eosinophils.

BACKGROUND: Evidence shows that leukotriene receptor antagonist (LTRA) can cause a partial reduction of eosinophils in the asthmatic airway. Although cysteinyl leukotrienes (CysLTs) can regulate the development of eosinophilic inflammation, LTRA might modulate the eosinophilic response to other inflammatory molecules involved in allergic inflammation. Montelukast is an LTRA that inhibits eosinophil transendothelial migration (TEM) in response to platelet-activating factor (PAF). The present study evaluates whether pranlukast (an LTRA) modifies eosinophil TEM in response to chemoattractants including PAF and C-C chemokines. METHODS: Eosinophils isolated from the blood of healthy individuals were incubated with or without pranlukast. We then evaluated eosinophil transmigration across human umbilical vein endothelial cells in response to LTD(4), eotaxin, RANTES and PAF. RESULTS: Pranlukast did not modify the spontaneous transmigration of eosinophils (n = 5). As reported, eosinophil TEM was significantly augmented by 0.1 microM LTD(4) and this enhancement was blocked by 1 microM pranlukast (p < 0.001; n = 6). On the other hand, pranlukast did not modify eosinophil transmigration in response to eotaxin, RANTES, or PAF (p > 0.1; n = 5). CONCLUSION: The inhibitory effect of pranlukast on eosinophil transmigration is highly specific for the CysLT1-dependent pathway.

Adult↗
PubMed1994

Role of resident peritoneal cells in eosinophil migration induced by saline.

In the present study, we investigated the role of resident peritoneal cells as well as the mediators involved in the eosinophil migration induced by large volumes of physiological saline. Two consecutive intraperitoneal injections of saline given 48 h apart, induced a selective recruitment of eosinophils into the cavity. This response was not observed with phosphate-buffered saline (PBS). Saline-induced eosinophil migration may be mediated at least in part by LTB4, since the lipoxygenase inhibitors BW A4C and MK 886 prevented the response. In the presence of saline, but not of PBS, mast cells and macrophages incubated in vitro released a factor which induced eosinophil migration when injected into the peritoneal cavity of rats. This release was inhibited by BW A4C and MK 886. These results indicate the importance of mast cells and macrophages in the eosinophil migration induced by saline and suggest the participation of LTB4 in this phenomenon. An abrupt reduction in the extracellular potassium concentration at the membrane of resident cells may be responsible for the saline effects since addition of potassium ions to saline abolished the eosinophil chemotactic activity of the same as well as its ability to stimulate the release of eosinophil chemotactic factor in vitro. Dexamethasone blocked both the saline-induced eosinophil migration and the release of eosinophil chemotactic factor by mast cells and macrophages. Pretreatment of the animals with dexamethasone inhibited the eosinophil migration induced either by the supernatants of saline-stimulated mast cells and macrophages or by LTB4. These results indicate that the release of additional mediators is necessary in order to account for the final eosinophil migration.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2000

Leukotriene receptor antagonists and synthesis inhibitors reverse survival in eosinophils of asthmatic individuals.

Eosinophilia is a feature of airway inflammation associated with asthma. Leukotriene antagonists provide therapeutic benefit in asthma, but their potential antiinflammatory actions have not been fully explored. We have examined the role of eosinophil-derived cysteinyl leukotrienes in the maintenance of eosinophil survival, and the involvement of leukotrienes in the paracrine stimulation of eosinophil survival by mast cells and lymphocytes. We obtained eosinophils and autologous lymphocytes from peripheral blood of asthmatic subjects. Leukotriene (LT)-B(4), LTC(4) and LTD(4), granulocyte-macrophage colony-stimulating factor (GM-CSF), and fibronectin promoted eosinophil survival. LTD(4) (10(-)(6) M) was as effective as GM-CSF (5 ng/ml) and fibronectin (400 ng/ml) in promoting survival. Lymphocytes and conditioned medium from a human mast cell line (HMC-1) induced eosinophil survival. Blockade of cysteinyl leukotriene receptors with SKF 104353 (pobilukast, 3 nM), and inhibition of 5-lipoxygenase (5-LO) with BW A4C (1 microM) and of 5-LO activating protein with MK 886 (1 microM), all increased basal rates of eosinophil apoptosis and reversed GM-CSF-induced eosinophil survival. Fifty percent reversal of GM-CSF- induced survival was achieved with SKF 104353 at 0.3 nM. The potency of SKF 104353 was two orders of magnitude greater than that of the LTB(4) receptor antagonist SB 201146. Mast cell- and lymphocyte-induced eosinophil survival were completely reversed by SB 201146, SKF 104353, BW A4C, and MK 886. These findings provide evidence for the involvement of an autocrine cysteinyl leukotriene pathway that supports eosinophil survival in response to a range of survival stimuli. They also suggest that LTB(4) could act as a paracrine stimulus of eosinophil survival.

Acrylates↗
PubMed1995

The effect of conditioned medium from cultured human bronchial epithelial cells on eosinophil and neutrophil chemotaxis and adherence in vitro.

Several studies have demonstrated that bronchial epithelial cells are capable of synthesizing proinflammatory cytokines that may influence eosinophil and neutrophil activity. We have cultured human bronchial epithelial cells to confluence, as explant cultures, and investigated the effect of conditioned medium from these cells on (1) the chemotaxis of eosinophils and neutrophils and (2) the adherence of these cells to cultured human endothelial cells. Analysis of cytokines, namely interleukin-1 beta (IL-1 beta), interleukin-8 (IL-8), tumor necrosis factor alpha (TNF alpha), granulocyte/macrophage colony-stimulating factor (GM-CSF), and RANTES, which are thought to be involved in these processes, demonstrated that all these cytokines were synthesized and released constitutively from the bronchial epithelial cell cultures. Conditioned medium obtained after 24 h of incubation significantly increased the chemotaxis of eosinophils and neutrophils, from median values of 4.0 cells/per high power field (hpf) (range, 3.0 to 7.0) and 17 cells/hpf (range, 13.0 to 25.0), respectively, for medium 199, to median values of 11.0 cells/hpf (range, 9 to 12; P = 0.005) and 30 cells/hpf (range, 19 to 33; p = 0.01). Whereas anti-GM-CSF and anti-IL-8 neutralizing monoclonal antibodies significantly attenuated the conditioned medium-induced chemotaxis of eosinophils and neutrophils, anti-RANTES neutralizing antibody significantly attenuated the chemotaxis of only eosinophils. Conditioned medium also significantly increased the percentage of eosinophils and neutrophils adhering to endothelial cells in a dose-dependent manner. Both anti-human TNF alpha and anti-human IL-1 beta neutralizing antibodies significantly attenuated the conditioned medium-induced adherence of eosinophils and neutrophils to the endothelial cells and were found to have an additive effect when studied together. Similarly, treatment of endothelial cells with either anti-ICAM-1 or anti-E-selectin, for 1 h before co-culture with eosinophils and neutrophils, significantly attenuated the conditioned medium-induced adherence of both eosinophils and neutrophils to endothelial cells. Treatment of endothelial cells with anti-VCAM-1 attenuated the adherence of eosinophils but not neutrophils. These results suggest that human bronchial epithelial cells, through their ability to generate proinflammatory mediators, are likely to play a role in the pathogenesis of airway disease by influencing chemotaxis and adherence of eosinophils and neutrophils.

Antibodies, Monoclonal↗
PubMed1996

Immunolocalization of interleukin-4 in eosinophils in the bronchial mucosa of atopic asthmatics.

Several studies have shown that human eosinophils can synthesize and release a number of cytokines. The aim of this study was to investigate whether eosinophils contain interleukin (IL)-4 protein. We examined the ultrastructural localization of IL-4 in eosinophils in bronchial mucosal biopsies of 29 nonsmoking atopic asthmatics and 7 controls. No eosinophils were detected in the bronchial mucosa of controls. In the eosinophils (n = 42) of the asthmatics, IL-4 was localized to the electron-dense crystalloid core compartment of 85% of the secondary or specific eosinophil granules (n = 468). Other structures in the eosinophils were unlabeled. Control sections, incubated with an irrelevant primary antibody, were negative. This study demonstrates that pre-formed IL-4 is stored in the secondary eosinophil granules. These results were extended by light microscopic immunodouble-staining for IL-4 protein and eosinophil cationic protein, which showed that a subpopulation of activated eosinophils express IL-4 immunoreactivity in bronchial mucosal biopsies of asthmatics as well as controls. These data indicate that eosinophils may be an important source of IL-4 in allergic inflammation.

Allergens↗
PubMed1997

Migration of eosinophils through basement membrane components in vitro: role of matrix metalloproteinase-9.

In general, inflammatory cells cross basement membranes by producing proteinases. To investigate the role of proteinases in eosinophil basement membrane migration, we studied peripheral blood eosinophils in Matrigel-coated chemotaxis chambers. Electron microscopy showed degradation of the Matrigel layer when eosinophils, added to the upper chamber, transmigrated the membrane in the presence of both platelet-activating factor (PAF) in the lower chamber and interleukin (IL)-5 in both chambers. In the absence of either or both PAF and IL-5, no changes occurred in the Matrigel layer. Matrigel transmigration of eosinophils induced by PAF and IL-5 was inhibited by 1,10-phenanthroline, batimastat, 3,4-dichloroisocoumarin, chymostatin, and a neutralizing antibody for the matrix metalloproteinase (MMP)-9, indicating that serine proteinase(s) and MMP, specifically MMP-9, were involved in the transmigration of eosinophils through Matrigel. In contrast, eosinophil migration through a bare membrane was not affected by batimastat. Using gelatin zymography and immunoblotting, MMP-9 was detected in the migration upper chamber supernatant of the eosinophil transmigration assay and in the conditioned medium of eosinophils. Release of MMP-9 by eosinophils was increased by IL-5, PAF, or both, but the substrate-degrading activity of MMP-9 was increased only in the presence of both IL-5 and PAF, indicating that the releasing and activating mechanisms of MMP-9 are involved in eosinophil basement membrane migration. This study implicates MMP-9 in basement membrane migration of eosinophils and suggests its involvement in inflammatory diseases where tissue eosinophilia plays a role.

Basement Membrane↗
PubMed1998

Ligation of the beta2 integrin triggers activation and degranulation of human eosinophils.

Evidence suggests that cellular adhesion is critical for eosinophil effector functions. Here, we tested the hypothesis that an adhesion molecule, specifically beta2 integrin, participates in intracellular signaling events of eosinophils. Eosinophils stimulated with interleukin (IL)-5 and adherent to protein-coated tissue culture plates via beta2 integrin (CD18) showed tyrosine phosphorylation of a number of proteins. Among these proteins, tyrosine phosphorylation of the 105 kD and 115 kD proteins and the product of the c-cbl protooncogene, Cbl, was specifically inhibited using soluble anti-CD18 monoclonal antibody (mAb) to block eosinophil cell adhesion. Furthermore, phosphoinositide turnover of IL-5-stimulated adherent eosinophils was also inhibited by anti-CD18 mAb, suggesting that cellular adhesion plays important roles in eosinophil signal transduction. alphaM beta2 (Mac-1, CD11b/18) was one of the beta2 integrins involved in eosinophil adhesion to protein-coated plates. We found that direct ligation of eosinophil alphaM beta2 with anti-CD11b mAb coupled to polystyrene microbeads induced tyrosine phosphorylation of a 115 kD protein and Cbl. Furthermore, anti-CD11b mAb microbeads induced increases in both phosphoinositide hydrolysis and the eosinophil degranulation response. Control antibodies, such as mouse myeloma IgG1 and anti-HLA class I antigen mAb, did not induce these cellular responses. These results suggest that engagement of beta2 integrin either by cell adhesion or by anti-CD11b mAb triggers activation of an intracellular signaling cascade, including protein tyrosine phosphorylation and phosphoinositide turnover, and subsequent cellular degranulation in human eosinophils. Tyrosine phosphorylation of a 115 kD protein and Cbl may play important roles in adhesion-dependent cellular functions of eosinophils.

Antibodies, Monoclonal↗
PubMed2000

CD14(+) cells are necessary for increased survival of eosinophils in response to lipopolysaccharide.

There has been considerable interest in the effect that gram-negative bacterial endotoxin (lipopolysaccharide [LPS]) can have in asthma, given that inhalation of LPS has been shown to cause bronchial hyperresponsiveness. Further, there is evidence that the endotoxin-binding protein CD14 may be a marker for asthma. Inhaled LPS has been shown to cause an influx of eosinophils into the nasal airway and to increase the survival of CD16-negatively selected eosinophils in vitro. In this study, we compared survival of eosinophils isolated via CD16-negative selection with eosinophils that were isolated using both CD16- and CD14-negative selection criteria. Survival of CD16-negatively selected eosinophils was enhanced by LPS in a dose-dependent manner and was inhibited by the endotoxin antagonists polymyxin B or lipid X. In contrast, depletion of CD14(+) cells within the eosinophil preparations (CD14/CD16-negatively selected eosinophils) decreased the effect of LPS on survival. Preincubation of CD16-negatively selected eosinophils with antibody 60bd, which blocks LPS binding to CD14, prevented the survival-enhancing effect of LPS. However, CD14 was not detected on eosinophils by flow cytometry, even after incubation with LPS for up to 24 h. These results suggest that the survival-enhancing effect of LPS on eosinophils requires the presence of CD14(+) cells in the population. It is our hypothesis that enhanced eosinophil survival with LPS involves the contribution of another cell type.

Animals↗
PubMed1989

Human lung fibroblast-derived granulocyte-macrophage colony stimulating factor (GM-CSF) mediates eosinophil survival in vitro.

Tissue eosinophilia has been reported to occur in pulmonary fibrosis, a disease characterized by chronic inflammation and lung fibroblast proliferation. We have examined the in vitro interaction of these two cell types by determining the in vitro survival of human peripheral blood eosinophils co-cultured with human lung fibroblasts. Survival of eosinophils cultured alone was 10% at day 3 and less than 1% at day 7. In contrast, survival of eosinophils that had been co-cultured with fibroblasts was 98, 90, 73, and 69% at days 3, 7, 10, and 14, respectively. Fibroblast-conditioned medium (CM) elicited a similar result in a dose-dependent fashion. Survival of eosinophils cultured with CM which had been preincubated with a monoclonal-neutralizing antibody to human GM-CSF was inhibited in a dose-dependent manner. Human recombinant-derived GM-CSF supported eosinophil survival in the dose-dependent fashion. Survival at day 7 of eosinophils treated with one single dose of GM-CSF (10 U/ml) was 64%. The effect of fibroblast-CM on eosinophils likely represents true survival since eosinophil proliferation as determined by [3H]thymidine incorporation did not occur. We also report that freshly isolated eosinophils had normal ultrastructural, scanning and transmission electron microscopy characteristics, and were normodense. In contrast, eosinophils co-cultured for 7 days with fibroblasts acquired irregular shapes and became hypodense and partially degranulated. Thus, our results indicate that human lung fibroblast-derived GM-CSF mediates the in vitro survival of human eosinophils.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies↗
PubMed1993

Platelet-activating factor-induced human eosinophil transendothelial migration: evidence for a dynamic role of the endothelium.

Stimulated migration of eosinophils out of the bloodstream and into the lung is key in the development of tissue eosinophilia and inflammation in asthma. Platelet-activating factor (PAF) has been implicated as an important inflammatory mediator in asthma pathogenesis in part because of its chemotactic capacity. We therefore studied the ability of PAF to induce human peripheral blood eosinophil migration through naked filters and human umbilical vein endothelial cells (HUVECs) cultured on these filters. PAF induced eosinophil migration through both barriers in a time-dependent fashion, with maximal eosinophil migration occurring at 180 min. Significant eosinophil migration was observed at PAF concentration > or = 0.1 microM and was dose dependent up to 10.0 microM. No significant differences in eosinophil chemotactic responses were noted between naked filter and HUVEC barriers. The PAF receptor antagonist, WEB 2086, inhibited (> 85%) eosinophil transendothelial migration when co-incubated with PAF or when used as a pretreatment of either the eosinophils or HUVECs. However, WEB 2086 pretreatment of HUVECs did not inhibit PAF-induced neutrophil transendothelial migration, nor did it affect leukotriene B4-induced neutrophil or eosinophil transendothelial migration. Thus, the data indicate that the endothelial cell plays an important role in PAF-induced eosinophil inflammatory processes. Moreover, these data suggest that PAF's pathogenic role in asthma may in part be due to its ability to stimulate eosinophil migration across endothelial barriers and into the airways.

Azepines↗
PubMed2003

Regulation of bone marrow and airway CD34+ eosinophils by interleukin-5.

The aim of this study was to evaluate the effect of a neutralizing anti-interleukin (IL)-5 monoclonal antibody (TRFK-5) on bone marrow and airway CD34(+) and immature eosinophils. A focus was to determine the effect of the timing of treatment. Balb/c mice were ovalbumin-sensitized and subsequently exposed to ovalbumin for 5-10 d via airway route. Animals were treated intraperitoneally with TRFK-5 or its isotype control (50 microg) once at different time points. Newly produced eosinophils were labeled using 5-bromo-2'-deoxyuridine (BrdU). BrdU(+) and CD34(+) eosinophil numbers were examined by immunocytochemistry. TRFK-5 reduced bone marrow immature eosinophils within 3 d. This effect was closely related to a reduction of BrdU(+) and CD34(+) bone marrow eosinophils, and reduced numbers of blood eosinophils. However, bronchoalveolar lavage (BAL) eosinophilia was not attenuated to the same degree. The effect of TRFK-5 was most prominent in the extended allergen-exposure protocol, where the treatment was given in the middle of the exposure, with strongly reduced bone marrow CD34(+) and immature bone marrow eosinophils, blood eosinophils as well as BAL BrdU(+) eosinophils, and BAL CD34(+) eosinophils. These data argue that anti-IL-5 downregulates eosinophilopoiesis within 3 d by action in the bone marrow, by inhibition of the early stages of eosinophil maturation from CD34(+) cells.

Allergens↗
PubMed2003

Expression of interleukin-5- and granulocyte macrophage-colony-stimulating factor-responsive genes in blood and airway eosinophils.

Because interleukin (IL)-5 family cytokines are critical regulators of eosinophil development, recruitment, and activation, this study was initiated to identify proteins induced by these cytokines in eosinophils. Using oligonucleotide microarrays, numerous transcripts were identified as responsive to both IL-5 and granulocyte macrophage-colony-stimulating factor (GM-CSF), but no transcripts were markedly affected by one cytokine and not the other. Expression of several gene products were seen to be increased following in vitro stimulation of human blood eosinophils, including the IL-3 receptor alpha subunit, lymphotoxin beta, Pim-1, and cyclin D3. Given that eosinophils recovered from the bronchoalveolar lavage fluid of allergic patients after antigen challenge are exposed to IL-5 or GM-CSF in the airway prior to isolation, the hypothesis was tested that selected IL-5- and GM-CSF-responsive genes are upregulated in airway eosinophils relative to the expression in blood cells. Airway eosinophils displayed greater cell surface expression of the IL-3 receptor alpha subunit, CD44, CD25, and CD66e, suggesting that these proteins may be markers of eosinophil activation by IL-5 family cytokines in airway eosinophils. Other genes that were induced by both IL-5 and GM-CSF showed protein expression at similar or decreased levels in airway eosinophils relative to their circulating counterparts (i.e., lymphotoxin beta and CD24). These studies have identified several transcriptional targets of IL-5 and GM-CSF in human eosinophils and suggest that a number of protein products are critical to the responsiveness of airway eosinophils.

Biomarkers↗
PubMed2006

Eosinophil trans-basement membrane migration induced by interleukin-8 and neutrophils.

Neutrophilic inflammation observed with severe asthma is often associated with interleukin-8 (IL-8). Neutrophils can secrete a variety of mediators that may augment the migration of eosinophils. We have reported a positive correlation between the concentrations of neutrophils and eosinophils in sputum from subjects with severe asthma, suggesting a possible role of neutrophils in regulating eosinophilic inflammation. The aim of this study was to investigate whether neutrophils stimulated with IL-8 modify the trans-basement membrane migration (TBM) of eosinophils. Eosinophils and neutrophils were isolated from peripheral blood drawn from healthy donors or subjects with mild asthma. The TBM of eosinophils in response to IL-8 was evaluated in the presence or absence of neutrophils using the chambers with a Matrigel-coated transwell insert. Neither IL-8 alone nor the presence of neutrophils alone induced the TBM of eosinophils. However, when eosinophils were coincubated with neutrophils and stimulated with IL-8, the TBM of eosinophils was significantly augmented. This augmented TBM of eosinophils was inhibited by a matrix metalloproteinase-9 inhibitor, a leukotriene B4 receptor antagonist, platelet-activating factor antagonists, or an anti-TNF-alpha monoclonal antibodies. These results suggest that neutrophils migrated in response to IL-8 may lead eosinophils to accumulate in the airways of asthma and possibly aggravate this disease.

Adult↗
PubMed2006

Dissection of the hyperadhesive phenotype of airway eosinophils in asthma.

Asthma is characterized by appearance of eosinophils in the airway. Eosinophils purified from the airway 48 h after segmental antigen challenge are described as exhibiting greater adhesion to albumin-coated surfaces via an unidentified beta2 integrin and increased expression of alphaMbeta2 (CD11b/18) compared with purified blood eosinophils. We have investigated the determinants of this hyperadhesive phenotype. Airway eosinophils exhibited increased reactivity with the CBRM1/5 anti-alphaM activation-sensitive antibody as well as enhanced adhesion to VCAM-1 (CD106) and diverse ligands, including albumin, ICAM-1 (CD54), fibrinogen, and vitronectin. Purified blood eosinophils did not adhere to the latter diverse ligands. Enhanced adhesion of airway eosinophils was blocked by anti-alphaMbeta2. Podosomes, structures implicated in cell movement and proteolysis of matrix proteins, were larger and more common on airway eosinophils adherent to VCAM-1 when compared with blood eosinophils. Incubation of blood eosinophils with IL-5 replicated the phenotype of airway eosinophils. That is, IL-5 enhanced recognition of alphaM by CBRM1/5; stimulated alphaMbeta2-mediated adhesion to VCAM-1, albumin, ICAM-1, fibrinogen, and vitronectin; and increased podosome formation on VCAM-1. Thus, the hyperadhesion of airway eosinophils after antigen challenge is mediated by upregulated and activated alphaMbeta2.

Antibodies↗
PubMed2002

Human eosinophils produce neurotrophins and secrete nerve growth factor on immunologic stimuli.

Neurotrophins, such as nerve growth factor (NGF) and neurotrophin-3 (NT-3), are essential for development, function, and survival of peripheral sympathetic and sensory neurons. Most eosinophilic leukocytes in the human body are localized in mucosal tissues; however, the roles of eosinophils in human diseases are not fully understood. We found that human eosinophils constitutively express messenger RNA for NGF and NT-3, synthesize and store these proteins intracellularly, and continuously replenish them. Incubation of eosinophils with a transcription inhibitor, actinomycin D, for 8 hours completely depletes intracellular NGF and NT-3. New synthesis of NGF is enhanced by Fc-receptor-mediated stimuli, such as immunoglobulin (Ig)A and IgG immune complexes; in contrast, production of NT-3 is not affected by these stimuli. Notably, supernatants of eosinophils stimulated with IgA immune complex and interleukin 5 promote neurite extension of the PC-12 pheochromocytoma cell line; this effect is abolished by pretreatment of the supernatants with anti-NGF-neutralizing antibody. By enzyme-linked immunosorbent assay, substantial amounts of NGF protein are also detected in the supernatants of stimulated eosinophils. Furthermore, in patients with seasonal allergic rhinitis, the concentrations of NGF in nasal secretions correlate with the magnitudes of eosinophilic inflammation in the airway, suggesting a potential clinical implication of eosinophil NGF. Our observations propose a new pathologic mechanism by which eosinophils may contribute to enhanced neurologic responses in patients with allergic diseases and other eosinophilic disorders. Alternatively, eosinophils may play important roles in maintenance and restoration of homeostatic functions of mucosal tissues through the pleitropic activities of NGF.

Adolescent↗
PubMed2000

Stimulation of eosinophil IgE low-affinity receptor leads to increased adhesion molecule expression and cell migration.

Immunoglobulin binding on eosinophil surface receptors results in activation of these cells. Evaluating blood eosinophils from atopic subjects, it was investigated whether ligation of immunoglobulin E low-affinity receptor (FcepsilonRII/ CD23) with specific monoclonal antibodies (Mabs) resulted in enhanced eosinophil migration and adhesion molecule expression. Eosinophils from 20 subjects with allergic asthma (atopic individuals) and nine nonatopic normal individuals (controls) were purified using Percoll gradients. The effect of antihuman CD23 Mabs on: 1) eosinophil migration through human umbilical vein endothelial cells (HUVECs); and 2) eosinophil expression of the adhesion molecules leukocyte function-associated antigen-1 (LFA-1, CD11a/CD18), macrophage antigen-1 (Mac-1, CD11b/CD18) and very late activation antigen-1 (VLA-4, CD49d/CD29) was evaluated by specific Mab staining and flow cytometric analysis. As compared to controls, freshly isolated eosinophils from atopic individuals showed enhanced migration through HUVECs (p<0.05) and increased LFA-1 expression (p<0.01), but similar Mac-1 and VLA-4 expression (p>0.1 for both). In both controls and atopic individuals, eosinophil incubation with antihuman CD23 Mabs induced a dose-dependent increase in cell migration through HUVECs, significant at antihuman CD23 Mab concentrations of 5 microg x mL(-1) (p>0.05 for all). Similarly, incubation of the cells with antihuman CD23 Mabs induced dose-dependent upregulation of LFA-1 and Mac-1 expression, whereas no changes in VLA-4 expression were observed (p>0.1). Finally, the enhanced eosinophil migration induced by antihuman CD23 Mab stimulation was significantly inhibited by antihuman LFA-1 (84+/-14% (mean+/-SEM); p<0.01) and VLA-4 Mabs (47+/-15%; p<0.05) but not by antihuman Mac-1 Mabs (p>0.1). In both atopic and control subjects, immunoglobulin E, low-affinity receptor stimulation induces functional changes in eosinophils characterized by increased eosinophil migration associated with enhanced late function antigen-1 and Mac-1 expression.

Adolescent↗