Interleukin-2 inhibits eosinophil migration but is counteracted by IL-5 priming

Conflicting data on the role of interleukin-2 in the recruitment of eosinophil granulocytes (EOS) to sites of inflammation have been presented. The objective of the present study was to investigate the effect of recombinant human IL-2 and anti-IL-2 on the migration of purified blood EOS. Neutralizing antibodies to IL-2 were added to a cytokine mixture with significant eosinophil chemotactic activity (ECA), and afterwards the ECA was tested on EOS from both normal and allergic donors. EOS migration was measured by a modification of the Boyden technique, using a 48-well microchemotaxis chamber. Recombinant human IL-2 was either added to the lower compartment of the chemotaxis chamber, or to the EOS for a pre-incubation period of 20 min, before migration assays towards the chemotaxins were performed. Anti-IL-2 caused a significant increase of EOS migration towards the cytokine mixture. Pre-incubation of the EOS with rhIL-2 inhibited the chemotaxis towards RANTES, PAF, IL-8 and eotaxin, and EOS migration towards IL-2 was lower than that towards buffer. These effects were more pronounced on EOS from normal than from allergic donors. Priming of the EOS with IL-5 prevented the inhibitory effect of IL-2. We hypothesize that IL-2 acts as an autocrine regulator of EOS migration, and that this inhibitory effect may be downregulated in allergy, allowing an increased migration of EOS towards chemotactic factors.     

Galectin-10 mRNA is overexpressed in peripheral blood of aspirin-induced asthma

Background:  In sensitive patients, aspirin is associated with nasal and bronchial inflammation, eliciting local symptoms. Although the disease is clinically well characterized, its physiopathology is incompletely understood and noninvasive procedures, allowing an effective distinction between aspirin-induced asthma (AIA) and aspirin-tolerant asthma (ATA) are missing.
Objectives:  The aims of the study were to compare AIA and ATA cohorts for clinical characteristics and to screen peripheral blood for differential mRNA expression.
Methods:  Patients experiencing symptoms following aspirin ingestion were considered as aspirin sensitive. Peripheral blood was collected to quantify mRNA expression, using microarray technology and quantitative RT-PCR.
Results:  Data indicated that AIA and ATA share large number of similarities for clinical phenotype. Screening of mRNA expression using microarray showed an overexpression of galectin-10 mRNA in AIA (AIA/ATA ratio = 1.9, P  < 0.05). Results were confirmed using qRT-PCR. A positive correlation was established between microarray and qRT-PCR results for galectin-10 mRNA expression ( r  = 0.92, P  < 0.0001). Finally, qRT-PCR results were validated on a subset of asthmatics and controls, showing an increased expression of galectin-10 mRNA in AIA vs ATA ( P  < 0.001) and vs controls ( P  < 0.01).
Conclusions:  Our results demonstrate that AIA and ATA remain difficult to distinguish using clinical criteria. Employing two molecular biological methods, we demonstrate that galectin-10 mRNA is overexpressed in AIA, suggesting a novel candidate gene and a potentially innovative pathway for mucosal inflammation in aspirin intolerance.     

Interleukin-12 inhibits eosinophil degranulation and migration but does not promote eosinophil apoptosis

BACKGROUND: Animal and human studies demonstrated that interleukin (IL)-12, a Th1 cytokine, reduces blood and bronchial eosinophilia, and airway hyperreactivity. According to current concepts, these effects are mediated through the release of cytokines promoting eosinophil recruitment and activation. However, the presence of IL-12 receptors on eosinophils suggests that IL-12 also acts directly on eosinophils. We postulated that IL-12 directly modulates eosinophil functions and has the capacity to regulate eosinophil degranulation, migration and survival, in vitro. METHOD: Effects of IL- 12 on purified human blood eosinophils were evaluated for peroxidase (EPO) release, eotaxin-induced migration through a model of basement membrane (Matrigel), and survival. RESULTS: IL-12 inhibited 50% of PAF and secretory IgA-induced EPO release (n = 8, p < 0.001). IL-12 also reduced eotaxin-induced migration through Matrigel by 54 +/-6% (n = 6, p < 0.01). These effects were not explained by an IL-12-induced impaired viability or apoptosis. CONCLUSION: Our results demonstrate that IL-12 directly modulates eosinophil functions without promoting apoptosis and explain, at least in part, the effects of IL-12 on eosinophils observed in in vivo studies.     

Cyclooxygenase 1 and cyclooxygenase 2 expression is abnormally regulated in human nasal polyps

BACKGROUND: There is evidence that impairment of prostanoid metabolism might be involved in the pathogenesis of nasal polyps (NPs). Prostanoids are synthesized by 2 cyclooxygenase (Cox) enzymes, one constitutive (Cox-1) and another inducible (Cox-2). OBJECTIVE: The aim of these studies was to investigate Cox-1 and Cox-2 regulation in NPs of aspirin-tolerant human patients compared with that seen in nasal mucosa (NM). METHODS: Cultured explants from human NPs and healthy mucosa from patients undergoing polypectomy and corrective nasal surgery, respectively, were examined for Cox-1 and Cox-2 expression by means of semiquantitative competitive PCR and Western blotting. RESULTS: Cox-1 mRNA was spontaneously upregulated in cultured NM but not in NPs. A spontaneous but delayed upregulation of Cox-2 mRNA was found in NPs (24 hours) compared with that seen in NM (6 hours). After cytokine stimulation (IFN-gamma, IL-1beta, and TNF-alpha), the induction of Cox-2 mRNA and protein was also faster in NM (1 hour) than in NPs (4 hours). CONCLUSION: These data showing an abnormal regulation of Cox-1 and Cox-2 in NPs from aspirin-tolerant patients reinforce the concept that prostanoid metabolism might be important in the pathogenesis of inflammatory nasal diseases and suggest a potential role for this alteration in the formation of NPs.     

Immunotherapy decreases antigen-induced eosinophil cell migration into the nasal cavity

We investigated the effect of immunotherapy (IT) on eosinophil (EOS) migration into the nasal cavity after nasal provocation with ragweed antigen and during seasonal exposure. In the first study, three groups of subjects participated: one group with no treatment (N = 19), one group with 10 months of IT, reaching maintenance at 2 micrograms of Amb a I (antigen E) (N = 15), and one group with 22 months of IT, reaching maintenance at 24 micrograms of Amb a I (N = 10). The percent of EOSs in nasal lavages performed during December before and 24 hours after nasal challenge with ragweed extract was determined. No significant difference between groups existed before challenge. The no treatment group demonstrated a significant increase in the percent of EOSs from 26% to 69.5% (p less than 0.008), whereas the treated groups demonstrated no significant change. In the second study, 45 patients were divided into four groups based on maintenance dose in micrograms of Amb a I and duration of treatment: (1) no treatment (N = 15), (2) 1 year at 2 micrograms (N = 13), (3) 2 years at 2 micrograms (N = 11), and (4) 3 years at 24 micrograms (N = 9). Nasal mucosal brushings were done during the ragweed season. A significantly smaller percentage of EOSs in 3-year IT-treated individuals was obtained compared to the control group (18 versus 8.4; p less than 0.04). The smaller dose of IT, regardless of duration, did not reveal a reduction compared to that in the no-treatment group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Allergen-induced increase of eosinophil cationic protein in nasal lavage fluid: effect of the glucocorticoid budesonide

It was our aim to study the effect of nasal allergen provocation on the concentration of eosinophil cationic protein (ECP) in nasal lavage fluid, with and without glucocorticoid pretreatment. Twenty grass-pollen sensitive volunteers were provoked outside the pollen season on 2 consecutive days after pretreatment for 2 weeks with the glucocorticoid, budesonide, as a nasal spray (400 micrograms/day) and with placebo with a double-blind, crossover design. Nasal lavage fluid was repeatedly collected during a 10-hour period to study both early and late-phase responses. 99mTechnetium-albumin was added to the lavage fluid, making it possible to calculate the amount of secretion and the degree of dilution. The results were as follows: (1) There was no correlation between ECP concentration and dilution factor in the individual samples. (2) The mean concentration of ECP in lavage fluid from untreated, prechallenge noses was 400 micrograms/L. (3) The ECP level did not increase during the early phase response. (4) There was a late occurring increase in the ECP concentration (6 to 24 hours). (5) This increase was completely inhibited by budesonide pretreatment. (6) The glucocorticoid therapy also reduced the prechallenge ECP concentration. In conclusion, allergen provocation in the nose results in a late occurring increase of ECP in nasal lavage fluid, and one of the therapeutic effects of topical glucocorticoid therapy may be an inhibition of the allergen-induced increase of this cytotoxic molecule.     

Thrombin affects eosinophil migration via protease-activated receptor-1

BACKGROUND: Protease-activated receptors (PARs) are a unique class of G-protein-coupled receptors, which are activated by proteolytic cleavage of the amino terminus of the receptor itself. Although expression of the PAR1, which is typically activated by thrombin, on human eosinophils has been demonstrated, no effect of thrombin on eosinophil function has been shown yet. Thus we investigated whether thrombin affects eosinophil migration in vitro. METHODS: Eosinophils were obtained from venous blood of healthy donors. Cell migration was studied by micropore filter assays. Involvement of PARs in thrombin-dependent migration was tested functionally using selective agonist peptides for PARs and a cleavage blocking PAR1 antibody. RESULTS: Thrombin significantly stimulated eosinophil chemotaxis in a dose-dependent manner. This effect was mimicked by the PAR1 but not the PAR2 agonist and was reversed by the cleavage blocking PAR1 antibody. Checkerboard experiments indicated that eosinophil migration depends on the presence of thrombin in a concentration gradient. CONCLUSIONS: Data suggest that activation of PAR1 by thrombin stimulates directed migration of human eosinophils and thereby may affect eosinophils in tissue and allergic inflammation.     

人鼻息肉组织中MMP-9及TIMP-1表达的变化

鼻息肉临床常见,发病率高.它的发生是一个涉及多因素、多步骤的过程.本研究应用免疫组织化学方法对38例鼻息肉及10例正常鼻黏膜组织中MMP-9及TIMP-1的表达进行检测,探讨它们与鼻息肉的临床分期分型、患者的年龄及性别的相互关系,从而为鼻息肉的临床诊断及治疗提供帮助.     

Epithelial damage in nasal polyps

Bronchial epithelial damage occurs regularly in bronchial asthma, but it is not known whether such damage occurs in the mucosa of nasal polyps. We obtained nasal polyp tissues from thirty patients and we examined these tissues for evidence of epithelial damage. Immediately after resection, polyp tissue was fixed in Karnovsky's fixative, embedded in methacrylate and stained with Giemsa pH 6-5. Normal nasal tissue from eight patients undergoing nasal septal reconstruction was similarly processed. As a disease control, we examined tissue from eight patients with nasal polyps associated with cystic fibrosis. Tissues were viewed by microscopy and epithelial damage was expressed as the percent of surface involved. Twenty-eight of the thirty patients with idiopathic nasal polps (93%) showed complete loss of nasal mucosa in varying degrees (range 3-81% of surface; mean, 29%). All patients showed evidence of some epithelial damage, either complete loss or marked desquamation (range 9-99% of surface; mean 54%). In contrast, six of eight biopsies from patients undergoing septal reconstruction and five of eight nasal polyps from patients with cystic fibrosis showed little or no evidence of epithelial damage. The results indicate that nasal polyps regularly show evidence of epithelial damage similar to that seen in bronchial asthma, and this abnormality may partly explain the rhinorrhea which is prominently associated with nasal polyps.     

Angiogenesis in chronic rhinosinusitis with nasal polyps and in antrochoanal polyps

Objectives and study design  

Angiogenesis may be related to the pathogenesis and prognosis of chronic rhinosinusitis with nasal polyposis. This cross-sectional study, in a tertiary university hospital, evaluates angiogenesis parameters in nasal polyps, antrochoanal polyps and middle turbinates.     

Topical glucocorticoids downregulate COX-1 positive cells in nasal polyps

Background:  Influx of inflammatory cells is one of the hallmarks of nasal polyposis. As glucocorticoids (GC) are known to exhibit strong anti-inflammatory effects, these drugs are frequently used in the treatment of the disease. Part of the anti-inflammatory effects of GC is attributed to their interference with prostanoid synthesis. As cyclooxygenases (COX) are key enzymes in the synthesis of both pro- (COX-1, COX-2) and anti-inflammatory prostanoids (COX-2), we investigated the role of topical GC on COX-1, COX-2 and inflammatory markers in nasal polyps (NP).
Methods:  Immunohistochemical analysis of inflammatory markers (CD68, CD117, MBP, elastase, IgE, BB-1, IL-4, IL-5 and IL-6), COX-1 and COX-2 was performed on normal nasal mucosa (NM) ( n  = 18), non-GC treated NP ( n  = 27) and topical GC treated NP ( n  = 12). NP groups were matched for allergy, asthma and ASA intolerance.
Results:  Increased numbers of eosinophils, IL-5+ cells and IgE+ cells and decreased numbers of mastcells are striking features of NP inflammation ( P  < 0.05). In addition, increased numbers of COX-1+ cells are observed in NP epithelium compared to NM ( P  < 0.05).
Conclusion:  Topical GC significantly reduce the number of COX-1+ NP cells ( P  < 0.05), but have no significant effect on COX-2+ NP cells. No significant reduction in the number of eosinophils is observed for GC treated NP. The number of IL-5+ cells is however increased significantly upon GC treatment ( P  < 0.05).     

Theophylline inhibits TNF-alpha-induced CD4 expression on human eosinophils and CD4+ eosinophil migration

BACKGROUND: Increasing evidence regarding asthma suggests that CD4+ cells are preferentially recruited to sites of bronchial inflammation. Interleukin (IL)-16 has been reported as playing an important role in the accumulation of CD4+ cells. We have shown that the CD4 molecule is expressed on normal human eosinophils by tumor necrosis factor (TNF)-alpha stimulation. METHODS: We evaluated the effects of theophylline, KF19514 [a selective phosphodiesterase (PDE) IV inhibitor] and dexamethasone on CD4 expression on eosinophils and eosinophil migration in response to IL-16, a natural soluble ligand of the CD4 molecule. RESULTS: The maximum eosinophil migration was observed when eosinophils were cultured with TNF-alpha at 10 ng/ml for 18 h and the concentration of IL-16 was 10 pg/ml. CD4+ eosinophil migration in response to IL-16 was mostly, if not fully, chemokinetic and this migration was significantly inhibited by Fab of anti-CD4 monoclonal antibody. Theophylline (10(-4)-10(-3) M), KF19514 (10(-7)-10(-6) M) and dexamethasone (10(-8)- 10(-6) M) significantly inhibited CD4 expression on eosinophils induced by TNF-alpha. Theophylline (10(-3) M) and KF19514 (10(-6) M) inhibited CD4+ eosinophil migratory responses induced by IL-16, but 10(-6) M dexamethasone did not. Theophylline and KF19514 augmented the intracellular adenosine-3',5'-cyclic monophosphate (cAMP) concentration in eosinophils, suggesting modulation by cAMP of CD4 expression and eosinophil migration. CONCLUSIONS: These data suggest that TNF-alpha-induced CD4+ eosinophils may contribute to eosinophil migratory responses induced by IL-16. Theophylline and selective PDE IV inhibitor may prevent airway inflammation by downregulating CD4 expression on eosinophils and inhibiting eosinophil migration through CD4 and IL-16 interaction.     

抗白介素5单克隆抗体抑制哮喘小鼠嗜酸性粒细胞迁移

目的 观察抗白介素5(IL-5)单克隆抗体对哮喘小鼠嗜酸性粒细胞(Eos)从骨髓到气道迁移的影响.方法 15只雄性C57BL/6小鼠.常规法复制哮喘,并在每次卵白蛋白(Ova)激发前2 h鼻腔内滴入抗IL-5单克隆抗体,同时设立阴性对照组,即以生理盐水代替Ova 和抗IL-5抗体.在末次抗原激发后12 h测定支气管肺泡灌洗液(BALF)、外周血(PB)和骨髓(BM)中炎症细胞数以及肺组织内Eos数.结果 Ova抗原激发后12 h小鼠的BALF、PB和BM及细支气管和肺组织中的Eos数显著增加(P＜0.01, P＜0.05).抗IL-5单克隆抗体则使上述变化显著减轻(P＜0.05,P＜0.01).结论 抗IL-5单克隆抗体能显著抑制哮喘小鼠嗜酸性粒细胞的迁移.     

A new antihistamine levocetirizine inhibits eosinophil adhesion to vascular cell adhesion molecule-1 under flow conditions

BACKGROUND: We previously demonstrated that low concentrations of a new antihistamine levocetirizine inhibited eosinophil transmigration through human microvascular endothelial cells. OBJECTIVE: Here, the inhibitory effect of levocetirizine on eosinophil adhesion to recombinant human vascular cell adhesion molecule-1 (rhVCAM)-1 was examined under conditions of shear stress using an in vitro model of the post-capillary venules. METHODS: Eosinophils isolated from normal subjects were pre-incubated with a concentration range of levocetirizine (10(-6)-10(-10) m) or negative dilution control. Resting or granulocyte macrophage-colony stimulating factor (GM-CSF)-stimulated cells were pumped through rhVCAM-1 (10 microg/mL) coated capillary tubes using a microfluidic syringe pump at a precise and constant flow rate (1 dyn/cm(2)). Images of rolling and firmly adherent eosinophils were captured using real-time video microscopy. RESULTS: Levocetirizine significantly inhibited resting eosinophil adhesion to rhVCAM-1 with maximal effect at 10(-8) M with an EC(50) of 10(-9) m. Levocetirizine almost abolished resting eosinophil adhesion by the 15 min time-point. GM-CSF significantly enhanced eosinophil adhesion and their ability to flatten on rhVCAM-1. Both phenomena were inhibited by levocetirizine in a dose-dependent manner, at both 5 and 15 min (optimal concentration of 10(-8) m with an EC(50) of 10(-9) m). Real-time imaging revealed that the effect of levocetirizine on post-adhesion behaviour (detachment, flatness) contributed to its inhibitory action on eosinophil adhesion to rhVCAM-1. In contrast, very late antigen (VLA)-4 mAb inhibited eosinophil adhesion to rhVCAM-1 from the earliest time-points. CONCLUSION: Physiologically relevant concentrations of levocetirizine inhibit resting and GM-CSF-stimulated firm eosinophil adhesion to rhVCAM-1 under flow conditions.