Migration through basement membrane modulates eosinophil expression of CD44

BACKGROUND: Tissue eosinophils express more membrane receptors and release more mediators than blood eosinophils, suggesting that migration from blood to tissue modulates eosinophil phenotype and functions. OBJECTIVE: We postulated that eosinophil passage through endothelial basement membrane, an important step of eosinophil migration into tissue, may be responsible for some of these changes. METHOD: We previously showed that 5-oxo-6, 8, 11, 14-eicosatetraenoic acid (5-oxo-ETE) in combination with IL-5 promotes eosinophil migration through Matrigel, a mouse tumour cell-derived basement membrane. Using this model, we evaluated the effect of trans-Matrigel migration on purified human blood eosinophil expressions of CD44, CD69 and HLA-DR that either increase or appear on activated eosinophils, and releases of peroxidase (EPO), leukotriene (LT) C(4) and granulocyte-monocyte colony stimulating factor (GM-CSF). RESULTS: IL-5, but not 5-oxo-ETE, increased eosinophil expression of CD44 and CD69. Migration of eosinophils through Matrigel significantly increased CD44 expression level over the one induced by IL-5 (P = 0.0001). Migration through Matrigel did not modify CD69 expression compared with the one obtained in the presence of IL-5 alone; however, incubation of eosinophils on Matrigel decreased IL-5-induced CD69 (P = 0.0001). Trans-Matrigel migration did not modify HLA-DR expression, nor EPO, LTC(4) and GM-CSF releases. CONCLUSION: These data show that in vitro trans-Matrigel migration and Matrigel contact modulate eosinophil membrane receptor expression. Consequently, they suggest that migration through basement membrane mediates changes in cell-surface phenotype observed on activated eosinophils and probably prepares them for interactions with tissue components and cells.     

CC chemokines and transmigration of eosinophils in the presence of vascular cell adhesion molecule 1.

BACKGROUND: Interaction between eosinophil alpha4 integrin and vascular cell adhesion molecule 1 (VCAM-1) expressed on activated endothelial cells may be a key step in the selective recruitment of eosinophils from the circulation to sites of inflammation. OBJECTIVE: To investigate the factor(s) that induces transmigration of eosinophils after firm adhesion via the alpha4 integrin/VCAM-1 pathway. METHODS: We examined the effects of a variety of inflammatory mediators on the migration of eosinophils across recombinant human (rh) intracellular adhesion molecule 1- or rhVCAM-1-coated Transwell filters or VCAM-1-expressing human pulmonary microvascular endothelial cells (HPMECs) that had been stimulated with interleukin 4 (IL-4) and tumor necrosis factor alpha. The number of eosinophils that had transmigrated was evaluated by measuring eosinophil peroxidase activity. RESULTS: The CC chemokines RANTES (regulated on activation, normal T-cell expressed, and secreted), eotaxin, eotaxin 2, monocyte chemotactic protein 3 (MCP-3), and MCP-4 each increased eosinophil transmigration across rhVCAM-1-coated filters compared with fetal calf serum-blocked or rh intracellular adhesion molecule 1-coated filters (P < .01). On the other hand, platelet-activating factor, C5a, formyl-methionyl-leucil-phenylalanine, granulocyte-macrophage colony-stimulating factor, IL-5, and IL-8 did not enhance migration across rhVCAM-1. The enhancement of migration by RANTES in the presence of rhVCAM-1 was blocked by an anti-alpha4 integrin monoclonal antibody. CC chemokines augmented eosinophil transmigration across VCAM-1-expressing HPMECs compared with resting HPMECs (P < .01). Conversely, the transmigration induced by platelet-activating factor, C5a, formyl-methionyl-leucil-phenylalanine, or IL-8 was not modified by the expression of VCAM-1 on HPMECs. CONCLUSIONS: CC chemokines induce transendothelial migration of eosinophils after interaction between eosinophil alpha4 integrin and endothelial VCAM-1.     

Inhibition of HLA-DR Expression on Activated Human Blood Eosinophils by Transforming Growth Factor-β1

Peripheral blood, bronchoalveolar lavage and sputum eosinophils of patients with asthma but not peripheral blood eosinophils from normal controls have been shown to express human leucocyte antigen (HLA)-DR on their cell surface. Cytokines implicated in the activation of eosinophils, such as interleukin (IL)-3 and granulocyte–macrophage colony-stimulating factor (GM-CSF), can up-regulate HLA-DR expression. However, little is known about antagonistic factors that might down-regulate HLA-DR expression on eosinophils. In this study we investigated whether transforming growth factor-β (TGF-β), which has been shown to reduce survival of activated eosinophils, can also modulate HLA-DR expression on eosinophils. For this purpose, isolated peripheral blood eosinophils were stimulated with IL-3 and GM-CSF for 24 h and HLA-DR expression was measured by flow cytometry. We found that while isolated eosinophils expressed low levels of surface HLA-DR, incubation with GM-CSF and IL-3 increased HLA-DR expression on eosinophils. TGF-β alone did not change HLA-DR expression on isolated eosinophils. However, co-incubation of eosinophils with TGF-β and either GM-CSF or IL-3 significantly decreased HLA-DR expression compared to eosinophils incubated with either GM-CSF or IL-3 alone and this was not reversed by addition of IL-5. This effect of TGF-β on IL-3-induced HLA-DR expression was attenuated dose-dependently in the presence of monoclonal anti-TGF-β antibodies. Our results suggest that TGF-β can reduce cytokine-induced HLA-DR expression on eosinophils and could thus influence eosinophil activation.     

CD69 is expressed by human eosinophils activated in vivo in asthma and in vitro by cytokines.

CD69 is an early activation marker for T cells and cross-linking of CD69 on platelets triggers aggregation and mediator release. Expression of a number of membrane receptors is induced on eosinophils after culture with certain cytokines. Therefore, we investigated whether cytokine-activated eosinophils expressed CD69. Unstimulated, peripheral blood eosinophils did not express CD69, as determined by immunofluorescence and flow cytometry (n = 15). CD69 expression was induced on eosinophils by granulocyte-macrophage colony-stimulating factor (GM-CSF) in a time- and dose-dependent manner. After 1 day in culture, expression was significant at concentrations of 10(-11) M and above. CD69 expression could be detected after stimulation with GM-CSF for only 1 hr, was significant after 2 hr and was sustained over 1-2 days in culture. CD69 expression was also induced by interleukin-3 (IL-3), IL-5 and interferon-gamma (IFN-gamma), but stimulation of eosinophils with platelet-activating factor (PAF) (10(-6) M) for up to 2 hr did not induce CD69 expression. Cycloheximide (10(-6) M) significantly inhibited GM-CSF-induced CD69 expression, suggesting a requirement for protein synthesis. However, unlike up-regulation of CR3 expression, GM-CSF-induced CD69 expression was not inhibited by dexamethasone. CD69 was present on eosinophils from the bronchoalveolar lavage (BAL) fluid of patients with mild asthma (5/5), suggesting that the in vitro findings may have biological relevance in vivo. Therefore, CD69 can be used as a marker of eosinophil activation by cytokines and is a candidate receptor for triggering eosinophil mediator release in the airways in asthma.     

Lower apoptosis rate and higher CD69 expression in neutrophils from atopic individuals

OBJECTIVE: In this study we aimed to examine the kinetics of CD69 expression and the susceptibility to apoptosis, in eosinophils and neutrophils, in the presence or absence of GM-CSF. We also addressed the question whether differences between atopic patients and healthy individuals exist in this respect. MATERIALS AND METHODS: Freshly isolated eosinophils and neutrophils from non-atopic and atopic donors were analysed by flow cytometry for Annexin/PI staining, caspase 3 activation and CD69 expression. RESULTS: We found a higher CD69 expression when atopic neutrophils were incubated with GM-CSF compared to non-atopic neutrophils, and that the kinetics of CD69-expression in neutrophils, but not in eosinophils, differed between non-atopic and atopic individuals (p < 0.004). We also found a higher viability in GM-CSF-stimulated neutrophils from non-atopic individuals as compared to neutrophils from atopic individuals (p < 0.05). CONCLUSION: These data suggest a potential role for neutrophils in the allergic inflammatory reaction through differences in apoptosis rates and CD69 expression between atopic and non-atopic individuals.     

Role of monocytes and eosinophils in human respiratory syncytial virus infection in vitro

RSV infection in airway epithelial cells (EC) results in production of the chemokines RANTES and MIP1alpha and the leukocyte differentiation factor GM-CSF. The chemokines attract monocytes and eosinophils to the site of infection, where GM-CSF may influence their function and differentiation. In turn, these inflammatory cells may limit the progression of RSV infection, as well as initiate immune responses. In the present study, the effect of monocytes and eosinophils on viral replication and infection-dependent release of EC-derived cytokines was investigated. The modulation of immune cell costimulatory molecules, CD80, CD86, CD40, and HLA-DR, and the release of the CD4(+) T cell chemoattractant IL-16 were also investigated. Employing immunofluorescence techniques, monocytes and eosinophils in cocultures with infected EC were found to inhibit the spread of RSV to uninfected cells. Monocytes also had a significant effect on replication of RSV. Monocytes phagocytized the virus, while eosinophils inhibited reinfection mainly by extracellular means. The release of G-CSF and GM-CSF in the infected cultures was not significantly affected by either monocytes or eosinophils, while RANTES release was significantly decreased. The expression of CD40, CD80, CD86, and HLA-DR on monocytes, but not on eosinophils, increased in an RSV-dose-dependent manner. IL-16 release was not induced in RSV-infected EC, but was significantly increased in coculture with monocytes. These results suggest that both monocytes and eosinophils attracted to the site of RSV infection play an important role in confining infection, while RSV-exposed monocytes may be involved in promoting/polarizing immune responses to RSV.     

Changes in Phenotype of Monocyte-Like THP-1 Cells Associated with Transendothelial Migration

The level of expression of CD11b and HLA-DR surface molecules on monocyte-like THP-1 cells increased significantly as a result of transmigration of these cells through a monolayer of endothelial cells. The expression of all studied markers (CD11b, HLADR, and CD-14) increased significantly after transendothelial migration in the presence of TNF-α and IFN-γ. The changes in surface phenotype of THP-1 cells after transendothelial migration in the presence of TNF-α were more pronounced than in the presence of IFN-γ. Transendothelial migration of THP-1 cells in the presence of IL-4 caused less pronounced changes in the surface phenotype, which did not differ from changes in transmigration without cytokines. Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 146, No. 11, pp. 531–534, November, 2008     

Expression of activation, adhesion molecules and intracellular cytokines in acute pancreatitis.

Adhesion and activation molecules as well as cytokines play an important role in an immune scenario. In acute pancreatitis, we have studied some of these in order to evaluate dysregulation. For this we took peripheral blood mononuclear cells and pancreatitis tissue cells. We analysed activation markers like CD69, CD25 and HLA-DR and found a marked elevation of CD69 as well as CD25 in both peripheral blood cells and tissue mononuclear cells when compared to controls. In PBMC-CD69: P<0.01 and CD25: P<0.01; in tissue-CD69: P<0.001 and CD25: P<0.001. The HLA-DR levels, however, were reduced in the disease state (in acute pancreatitis patient blood (P<0.01) and tissue cells (P<0.001)). The adhesion molecules showed unanimous rise in the blood and the tissue samples. In blood samples, CD11a: P<0.05 and CD11b: P<0.05 and tissue samples CD11a: P<0.01 and CD11b: P<0.01and CD54 in peripheral blood (P<0.05) and tissue (P<0.01) of AP was high as compared to controls. By simultaneous flowcytometric analysis, we determined the co-expression of a surface marker (CD4/CD8/CD14) and intracellular cytokine (TNF-alpha and IFN-gamma) in individual cells. The IFN-gamma producing CD8+T cells were elevated in pancreatic tissue (P<0.01). TNF-alpha producing cell numbers were significantly higher in tissue cells than in blood and also in CD8+ T cells (P<0.001). We conclude that monocyte function is affected in AP as shown by reduced HLA-DR numbers and lowered TNF-alpha producing cells. Moreover, the CD8+T cells appear to play an important role in cytokine synthesis at the effector site.     

MHC Class II and CD9 in human eosinophils localize to detergent-resistant membrane microdomains

Eosinophils function in murine allergic airways inflammation as professional antigen-presenting cells (APCs). In murine professional APC cell types, optimal functioning of MHC Class II depends on its lateral association in plasma membranes and colocalization with the tetraspanin CD9 into detergent-resistant membrane microdomains (DRMs). With human eosinophils, we evaluated the localization of MHC Class II (HLA-DR) to DRMs and the functional significance of such localization. In granulocyte-macrophage colony-stimulating factor-stimulated human eosinophils, antibody cross-linked HLA-DR colocalized by immunofluorescence microscopy focally on plasma membranes with CD9 and the DRM marker ganglioside GM1. In addition, HLA-DR coimmunoprecipitates with CD9 after chemical cross-linking of CD9. HLA-DR and CD9 were localized by Western blotting in eosinophil DRM subcellular fractions. DRM disruption with the cholesterol-depleting agent methyl-β-cyclodextrin decreased eosinophil surface expression of HLA-DR and CD9. We show that CD9 is abundant on the surface of eosinophils, presenting the first electron microscopy data of the ultrastructural immunolocalization of CD9 in human eosinophils. Disruption of HLA-DR-containing DRMs decreased the ability of superantigen-loaded human eosinophils to stimulate CD4(+) T-cell activation (CD69 expression), proliferation, and cytokine production. Our results, which demonstrate that eosinophil MHC Class II localizes to DRMs in association with CD9 in a functionally significant manner, represent a novel insight into the organization of the antigen presentation complex of human eosinophils.     

Bcl-2-mediated regulation of CD69-induced apoptosis of human eosinophils: identification and characterization of a novel receptor-induced mechanism and relationship to CD95-transduced signalling

Elimination of the eosinophils from the airways by selective induction of apoptosis represents a therapeutic approach for asthma. Here we report on a possible target molecule, the surface receptor CD69. To simulate an asthmatic response, segmental allergen challenge in mild asthmatics was performed. Eosinophil numbers increased in bronchoalveolar lavage (BAL) at 18 h. In contrast to blood cells, BAL eosinophils expressed the activation marker CD69. Purified blood eosinophils stimulated with granulocyte/macrophage colony-stimulating factor (GM-CSF) or interferon-gamma (IFN-gamma) expressed CD69 and showed prolonged viability. Only IFN-gamma enhanced constitutive CD95 expression. Coincubation with anti-CD69 or anti-CD95 monoclonal antibody (MoAb) induced apoptosis, as revealed by propidium iodide incorporation, membrane blebbing and nuclear fragmentation. Additionally, both anti-CD69 and anti-CD95 MoAb reduced cytokine-enhanced Bcl-2 expression. In conclusion, CD69 transduces a Bcl-2-dependent death signal when ligated by a specific antibody. As, in contrast to the ubiquitous death-inducer CD95, the function of CD69 appears to be restricted to activated eosinophils, it represents an ideal target for therapeutic intervention in asthma.     

Immunohistochemical detection of GM-CSF, IL-4 and IL-5 in a murine model of allergic bronchopulmonary aspergillosis

Background Granulocyte macrophage-colony stimulating factor (GM-CSF), interleukin-4 (IL-4) and IL-5 are important in tissue eosinophil accumulation and high IgE production in allergic inflammatory reaction. Objective We examine lung GM-CSF, IL-4 and IL-5 expression in a murine model of allergic bronchopulmonary aspergillosis (ABPA) characterized by eosinophil and lymphocyte lung infiltration and elevated serum IgE level. Methods C57BL/6 mice were intranasally treated three times a week for 1, 2 or 3 week(s) with Aspergillus fumigatus (Af) antigen or saline and were sacrificed on days 7, 14 and 21. Immunohistochemical analyses for GM-CSF, IL-4 and IL-5 were performed on lung sections. Results Af treatment induced a remarkable pulmonary eosinophil influx. Increased numbers of lung T lymphocytes and GM-CSF positive cells were observed on days 14 and 21. IL-4 and IL-5 positive cells were increased significantly only on day 14. Immunostained serial sections showed that most (≥98%) cytokine positive cells were CD3 positive. Few eosinophils (<2% of cytokine positive cells) were immunoreactive for GM-CSF and IL-5. Significant correlations were found between the number of GM-CSF and IL-5 positive cells, and the number of eosinophils in Af-treated lung (r = 0.62, P < 0.05 and r = 0.52, P < 0.05, respectively), and between the number of IL-4 positive cells and the serum total IgE level (r = 0.64, P<0.01). Conclusions Our data suggest a role for T lymphocyte GM-CSF, IL-4 and IL-5 in Af-induced mouse pulmonary eosinophilia and increased serum IgE production and further support the importance of T helper (TH) cells in the pathogenesis of ABPA.     

IL-34 对类风湿关节炎髓系树突状细胞表型的影响

目的:探讨IL-34 对类风湿关节炎(RA)患者单核细胞向髓系树突状细胞(DC)分化的诱导作用,及其在分化过程中对细胞表型的影响。方法:采集RA 患者外周血,Ficoll 密度梯度离心法获得PBMC,培养4 h 后将贴壁细胞分别用GM-CSF+IL-4、IL-4、IL-4+IL-34 刺激3 d,流式细胞术检测CD83、CD86 和CD14 的表达水平;再将GM-CSF+IL-4 刺激3 d 后的细胞加入TNF-α和(或)IL-34 培养4 d,流式细胞术检测CD83、CD86 和(或)CD14 的表达水平。结果:(1)IL-34+IL-4 诱导后CD83、CD86 表达水平较IL鄄4 单独作用明显上调(P<0.01),CD14 表达无差异(P>0.05);IL-34+IL-4 诱导CD86、CD14 表达水平较GM-CSF+IL-4 刺激组下降(P<0.05),CD83 表达无差异(P>0.05)(2)GM-CSF+IL-4+IL-34 诱导CD83、CD86 表达水平较GM-CSF+IL-4+TNF-α组低(P<0.05),但与GM-CSF+IL-4 刺激组对比CD83、CD86 表达水平差异无统计学意义(P>0.05)。(3)GM-CSF+IL-4+TNF-α+IL-34 诱导DC CD83 表达水平较GM-CSF+IL-4+TNF-α刺激组低(P<0.05),但两组CD86、CD14 表达差异无统计学意义(P>0.05)。结论:IL-34 在不成熟DC 诱导过程中发挥一定作用,效应略弱于GM-CSF;IL-34 对成熟DC诱导作用不显著,但参与了不成熟DC 的维持。     

The Effect of Age on the Transendothelial Migration of Human T Lymphocytes

Since the migration of immunologically active cells into perivascular tissue is an important step in acute and chronic inflammation, the authors studied the possible influence of age on the transendothelial migration of T cells in an in vitro model. After incubation on collagen gels coated with human endothelial cells, three populations of T cells of elderly and young subjects were harvested: non-adherent (NAD), bound (BND) and migrated (MIG) cells. The percentages of the BND and MIG fractions were similar in the young and elderly groups. Phenotypic analysis showed that in the starting population of the elderly subjects T-cell subpopulations that have an increased capacity of transendothelial migration (e.g. CD4⁺ memory cells, CD8⁺/CD57⁺ cells) were enriched: nevertheless ageing was not associated with an increase in the transendothelial migration of T cells.     

Enhancement of monocyte transendothelial migration by granulocyte-macrophage colony-stimulating factor: requirement for chemoattractant and CD11a/CD18 mechanisms

Granulocyte-macrophage colony-stimulating factor (GM-CSF) enhances and primes monocyte functions, but its role in monocyte migration is poorly understood. We examined monocyte migration across human umbilical vein endothelial cells (HUVEC) grown on filters. GM-CSF had no chemotactic or chemokinetic effect. However, GM-CSF enhanced monocyte transendothelial migration (TEM) through unstimulated and IL-1-activated (5 h) HUVEC in response to C5a or monocyte chemoattractant protein-1 in a dose-dependent fashion, increasing the migration from 28.7 +/- 5.3% to 41.8 +/- 6.2% (n = 8, p < 0.05) and from 34.8 +/- 6% to 50.3 +/- 3.1%, p < 0.05), respectively. The enhanced TEM was inhibited by monoclonal antibodies (mAb) to LFA-1, but not by mAb to Mac-1 or to VLA-4. Furthermore, GM-CSF up-regulated and activated LFA-1, as assessed by NKI-L16 neoepitope expression. The results indicate that: (1) GM-CSF can prime monocytes for increased TEM, (2) GM-CSF enhances LFA-1-mediated monocyte TEM and (3) this effect is in part mediated by increasing LFA-1 expression and activation. Thus, increased GM-CSF production may promote monocyte accumulation in inflammation not only by inducing monocytosis, but also enhancing migration.     

棕榈酸通过上调脂肪酸转位酶诱导THP-1细胞的炎症反应

目的:探讨脂肪酸转位酶(fatty acid translocase,FAT/CD36)在棕榈酸诱导的人源单核巨噬细胞THP-1炎症反应中的作用。方法:给予不同浓度棕榈酸(0 mmol/L、0.1 mmol/L和0.2 mmol/L)处理THP-1细胞24h。Transwell迁移实验检测细胞迁移能力;real-time PCR检测CD36、肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)及单核细胞趋化蛋白1(MCP-1)的m RNA表达水平;ELISA和Western blot法检测靶蛋白的蛋白含量。利用RNA干扰技术构建低表达CD36(si CD36)的THP-1细胞模型,观察抑制CD36表达对细胞迁移、炎症及趋化因子表达的影响。结果:棕榈酸促进了THP-1细胞CD36的m RNA和蛋白表达,且增强了THP-1细胞炎症/趋化因子的m RNA和蛋白表达,差异有统计学意义(P0.05)。棕榈酸处理组THP-1细胞的迁移能力明显强于对照组。与阴性对照组细胞相比,si CD36组炎症因子的m RNA和蛋白表达明显降低(P0.05),THP-1细胞迁移水平也明显降低(P0.05)。结论:棕榈酸通过上调THP-1巨噬细胞中CD36表达,促进了巨噬细胞的迁移,促使细胞产生大量炎症/趋化因子,导致巨噬细胞炎症反应。     

Characterization of the Priming Effect by Pituitary Canine Growth Hormone on Canine Polymorphonuclear Neutrophil Granulocyte Function

In this report, we demonstrate that canine growth hormone (cGH) is capable of priming canine polymorphonuclear neutrophil granulocytes (PMN) in a manner resembling that of human PMN. The cGH influences important functions that are involved in the process of recruitment of PMN, i.e., shape change, chemotaxis, CD11b/CD18 expression, adhesion, and subsequent transendothelial migration. Also, intracellular O₂⁻ production was evaluated. We investigated the priming effect by incubating PMN with purified pituitary cGH at various concentrations (10 to 800 μg/liter). The capacity for shape change was significantly (P < 0.05) enhanced, whereas the chemotactic response under agarose was significantly (P < 0.05) reduced. The chemotactic migration in Boyden chambers (10-μm-thick polycarbonate filter; lower surface count technique) was significantly (P < 0.05) enhanced, presumably due to cGH-induced hyperadhesiveness to the lower surface of the filters. The adhesion in albumin-coated microtiter plates and adherence to canine pulmonary fibroblasts were significantly (P < 0.05) increased, and the increased adhesion resulted in a significant (P < 0.01) increase in transendothelial migration using canine jugular vein endothelial cells. The increase in adhesion was associated with a significant increase in CD11b/CD18 expression. Furthermore, intracellular O₂⁻ production was significantly enhanced in response to both phorbol myristate acetate (P < 0.01) and opsonized zymosan (P < 0.05). In the absence of a PMN-stimulating agent, cGH did not influence the effector functions investigated except for an increased expression of CD11b/CD18.     

Spontaneous B-cell IgE production in a patient with remarkable eosinophilia and hyper IgE.

BACKGROUND: The pathophysiology of eosinophilia and hyper-IgE is not fully elucidated yet. OBJECTIVE: To clarify the pathophysiology of a patient with remarkable eosinophilia and hyper IgE, we examined cytokine levels in serum, surface antigens of peripheral blood eosinophils and IgE production in vitro. RESULTS: Concentrations of tumor necrosis factor-alpha (TNF-alpha), interleukin-3 (IL-3), interleukin-4 (IL-4), interleukin-5 (IL-5), and granulocyte/macrophage-colony stimulating factor (GM-CSF) in the serum were 21 pg/mL, <15 pg/mL, <15 pg/mL, 8 pg/mL, and <5 pg/mL pg/mL, respectively. Newly expressed surface antigens CD4, CD25, CD69, and HLA-DR, but not CD54, were observed on peripheral blood eosinophils. Extremely high levels of IgE secretion was found in the patient's mononuclear cells without stimuli; this was not enhanced by IL-4 or IL-4 plus anti-CD40 monoclonal antibody stimulation. Furthermore, highly purified B cells spontaneously produced large amounts of IgE and the production was not enhanced in addition of his T cells. CONCLUSION: The eosinophils were activated, and the B cells spontaneously produced IgE independently of T cells or cytokines, suggesting that intrinsic abnormality of B cells leading to dysregulated production of IgE in this disease.     

IL-4-induced migration of eosinophils in allergic inflammation

T lymphocytes present in allergically inflamed tissue synthesize and secrete the cytokines IL-3, IL-4, IL-5 and GM-CSF which may act as chemotaxins on eosinophils. In contrast to the former cytokines, IL-4 is chemotactic only for eosinophils from peripheral blood of patients with atopic dermatitis and not for eosinophils from normal individuals. IL-4 has the same chemotactic potency as the other cytokines. The optimal chemotactic potency is reached at a concentration of 10 nM. In contrast, neutrophils do not respond chemotactically to IL-4. Checkerboard analysis, inhibition studies with monoclonal anti-IL-4. Abs and desensitization experiments indicated specific interaction of IL-4 with eosinophils. In eosinophils from normal individuals, IL-4 responsiveness could be induced by pretreatment of the cells with IL-5 and GM-CSF. In addition to the fact that IL-4 may be responsible for selective eosinophil transendothelial migration, IL-4 may exert an important modulatory mode of action on eosinophil migration and function within allergically inflamed tissue. Our findings suggest the presence of a functional IL-4R on eosinophils from atopic dermatitis patients.