The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

The Journal of Immunology, 2008, 180, 7622 -7635
Copyright © 2008 by The American Association of Immunologists, Inc.
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Johansson, M. W.
Right arrow Articles by Mosher, D. F.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Johansson, M. W.
Right arrow Articles by Mosher, D. F.
Right arrowPubmed/NCBI databases
*Compound via MeSH
*Substance via MeSH
Medline Plus Health Information
*Asthma

Up-Regulation and Activation of Eosinophil Integrins in Blood and Airway after Segmental Lung Antigen Challenge1

Mats W. Johansson2,*, Elizabeth A. B. Kelly{dagger}, William W. Busse{dagger}, Nizar N. Jarjour{dagger} and Deane F. Mosher*,{dagger}

* Department of Biomolecular Chemistry and {dagger} Department of Medicine, University of Wisconsin, Madison, WI 53706


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
We hypothesized that there are clinically relevant differences in eosinophil integrin expression and activation in patients with asthma. To evaluate this, surface densities and activation states of integrins on eosinophils in blood and bronchoalveolar lavage (BAL) of 19 asthmatic subjects were studied before and 48 h after segmental Ag challenge. At 48 h, there was increased expression of {alpha}D and the N29 epitope of activated β1 integrins on blood eosinophils and of {alpha}M, β2, and the mAb24 epitope of activated β2 integrins on airway eosinophils. Changes correlated with the late-phase fall in forced expiratory volume in 1 s (FEV1) after whole-lung inhalation of the Ag that was subsequently used in segmental challenge and were greater in subjects defined as dual responders. Increased surface densities of {alpha}M and β2 and activation of β2 on airway eosinophils correlated with the concentration of IL-5 in BAL fluid. Activation of β1 and β2 on airway eosinophils correlated with eosinophil percentage in BAL. Thus, eosinophils respond to an allergic stimulus by activation of integrins in a sequence that likely promotes eosinophilic inflammation of the airway. Before challenge, β1 and β2 integrins of circulating eosinophils are in low-activation conformations and {alpha}Dβ2 surface expression is low. After Ag challenge, circulating eosinophils adopt a phenotype with activated β1 integrins and up-regulated {alpha}Dβ2, changes that are predicted to facilitate eosinophil arrest on VCAM-1 in bronchial vessels. Finally, eosinophils present in IL-5-rich airway fluid have a hyperadhesive phenotype associated with increased surface expression of {alpha}Mβ2 and activation of β2 integrins.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
The contribution of eosinophils to certain aspects of asthma, such as airway hyperresponsiveness, remains controversial; nevertheless, there is evidence that eosinophil recruitment to the airway contributes to asthma exacerbations and the chronic character of asthma by regulating airway inflammation and remodeling (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15). Thus, the study of how eosinophils traffic from blood to airway is of considerable importance. Integrins, which are versatile cellular adhesion receptors (16, 17), are likely determinants of how eosinophils roll and arrest on lung endothelium, extravasate and migrate through the endothelium, underlying basement membrane, and tissue to the airway wall, and traverse the bronchial epithelium to the airway lumen (18, 19, 20).

Human eosinophils express seven integrin heterodimers: {alpha}4β1 (CD49d/CD29), {alpha}6β1 (CD49f/CD29), {alpha}Lβ2 (CD11a/CD18), {alpha}Mβ2 (CD11b/CD18), {alpha}Xβ2 (CD11c/CD18), {alpha}Dβ2 (CD11d/CD18), and {alpha}4β7 (CD49d/β7) (13, 21, 22). Each integrin heterodimer interacts with its own set of ligands, which are counterreceptors on other cells or extracellular matrix components (17, 23). The functions of an integrin on a given cell is regulated by expression level and activation state (24, 25, 26).

We hypothesized that there are clinically relevant differences in eosinophil integrin expression levels and activation states in patients with asthma. The purpose of the present study was to define changes in the expression levels and activation states of blood and airway eosinophil integrins 48 h following segmental allergen challenge in asthmatic subjects, thus reflecting the development of allergic inflammation. Segmental Ag challenge induces a strong local recruitment of eosinophils (27). We found characteristic integrin changes on blood and airway eosinophils, differences between single and dual responder asthmatics, and associations among integrin changes, magnitude of eosinophil recruitment to airway, and IL-5 concentration in bronchoalveolar lavage (BAL)3 fluid. We propose a scenario in which the following occur: 1) Ag challenge leads to activation of β1 integrins and increased surface expression of {alpha}Dβ2 on blood eosinophils; 2) such activated eosinophils are more prone to arrest on VCAM-1 (CD106)-bearing endothelium in challenged segments; and 3) IL-5 and other cytokines trigger activation of β2 integrins, contributing to the hyperadhesive phenotype of airway eosinophils.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Subjects and screening

Nineteen subjects with mild asthma as diagnosed by an allergist were studied (Table I). These subjects had a history of asthma exacerbation to aeroallergen, a provocative concentration of methacholine producing a 20% fall in forced expiratory volume in 1 s (FEV1) of <8 mg/ml and/or reversibility to β-agonist of >12%. Subjects were screened as described previously with allergen skin prick tests, determination of airway hyperresponsiveness, and spirometry (28). All subjects had a positive skin prick test to one or more aeroallergens, were nonsmokers, did not have a respiratory infection within 30 days of study, and had not received antihistamines within seven days or corticosteroids within 30 days of study enrollment. The studies were reviewed and approved by the University of Wisconsin Health Sciences Human Subjects Committee (Madison, WI). Informed written consent was obtained from each subject before participation.


View this table:
[in this window]
[in a new window]

  		Table I. Subject characteristics

 
Determination of provocative allergen dose and early- and late-phase response to whole-lung allergen challenge

At least 4 wk before bronchoscopy, a graded whole-lung inhaled Ag challenge was performed as described (28) to determine the provocative dose of Ag producing a 20% fall in FEV1 (AgPD20) and the magnitude of early- and late-phase responses. Briefly, baseline spirometry was performed and repeated after five breaths of saline diluent. If FEV1 remained within 10% of baseline, five breaths of allergen were inhaled and spirometry was repeated 10 min later. Consecutively greater concentrations of allergen were given until FEV1 fell by ≥20% from baseline. The maximum immediate- or early-phase (within 15–30 min) fall in FEV1 was determined and subjects were then monitored every 15 min until FEV1 returned to within 10% of baseline. Thereafter, the subjects were monitored at 1-h intervals for 8 h to determine whether a late-phase response was present (28, 29). Subjects having a FEV1 fall of ≥15% 3–8 h after the whole-lung Ag challenge were considered to have a dual response phenotype; the other subjects were considered single responders (28).

Segmental bronchoprovocation with allergen and BAL

Bronchoscopy, segmental bronchoprovocation with allergen, and subsequent BAL were performed in two different bronchopulmonary segments as described (27, 28, 30). Baseline BAL at 0 h immediately before segmental bronchoprovocation was performed in two segments. Then, a total dose of 30% of the subject’s AgPD20 was administered incrementally to enhance subject safety as follows: 10% of the AgPD20 in the first segment and, when this dose was well tolerated, 20% in the second segment. Forty-eight hours later, a second bronchoscopy was performed by instilling 160 ml of sterile 0.9% NaCl warmed to 37°C in each segment. BAL fluid recovered from the two segments was pooled for analysis, and the volume of recovered fluid was measured.

Abs for flow cytometry and ELISA and recombinant protein standards

Anti-{alpha}D integrin mAb 240I (31) was obtained as a gift from ICOS. Activation-sensitive β2 integrin mAb24 (32, 33) was a gift from N. Hogg (Cancer Research U.K. London Research Institute, London, U.K.). Anti-β1 mAb MAR4, anti-β2 L130, anti-β7 Fib504, anti-{alpha}4 9F10, anti-{alpha}6 GoH3, anti-{alpha}L AI111, anti-{alpha}X Bly6, PE-conjugated goat anti-mouse and anti-rat IgG, FITC-conjugated anti-CD14 and anti-CD16, the isotype controls mouse IgG1 {kappa} (clone A112-2) and rat IgG2a {kappa} (A110-2), and the unlabeled and biotinylated anti-IL-3, anti-IL-5, and anti-IFN-{gamma} mAbs and corresponding recombinant protein standards for ELISA were from BD Biosciences. Anti-{alpha}M LM11 and activation-sensitive anti-β1 N29 (34) were from Chemicon. Unlabeled and biotinylated anti-GM-CSF mAb and recombinant protein standard for ELISA were from R&D Systems.

Flow cytometry of blood and BAL cell samples

Because purification of blood eosinophils has been found to cause the activation of β1 (35), flow cytometry was done on unfractionated blood and BAL cells. Surface expressions of {alpha}M, {alpha}L, {alpha}X, {alpha}D, {alpha}4, {alpha}6, β1, β2, β7, activated β1, and activated β2 were determined. Blood was drawn routinely into lavender top standard tubes (giving a final EDTA concentration of 1.8 mg/ml) (BD Vacutainer Systems). For determination of mAb24 reactivity, blood was drawn into green top tubes (giving a final heparin concentration of 14 U.S. Pharmacopeia (USP) units/ml). Control experiments revealed that the anticoagulant had no effect on the results except, as reported, for mAb24, whose epitope is not exposed in the presence of EDTA (32). Not all samples were subjected to complete analysis due to changes in the mAb panel that were made based on ongoing analysis of results from this study and other studies. Originally, we focused on differences between blood and BAL fluid 48 h after segmental challenge. Blood from before segmental challenge was included as a routine later, when we had indications of interesting differences between blood after vs before challenge. Similarly, we originally focused on subunits of the integrins known to be able to bind VCAM-1, {alpha}4β1, {alpha}4β7, and {alpha}Dβ2 (13, 21) and added {alpha}M later when we had evidence that {alpha}Mβ2 is involved in the adhesion of purified airway eosinophils to VCAM-1 and other ligands (22, 36). BAL fluid cells were recovered, cytospun, and stained for differential counts as described (29, 30).

EDTA-treated blood (100 µl) was incubated with 0.5 µg of primary Ab or isotype control in 100 µl of FACS buffer (PBS with 2% BSA and 0.2% NaN3) for 30 min. For mAb24 (and its isotype control), heparin-treated blood was used because the mAb24 epitope is not exposed in EDTA (32) and was incubated with primary Ab in RPMI 1640 with 10% FBS at 37°C following the protocol especially designed for mAb24 (37). After primary Ab incubation, samples were washed with 1 ml PBS, washed with 250 µl of FACS buffer, and resuspended in 250 µl of FACS buffer containing PE-conjugated goat anti-mouse or anti-rat IgG (2 µg/ml). After incubation for 30 min, samples were washed again with PBS, resuspended in 100 µl FACS buffer with a mixture of FITC-conjugated anti-CD14 (0.125 µg) and anti-CD16 (0.625 µg) and incubated for 30 min. RBC were lysed by incubation with 2 ml of FACS lysing solution (BD Biosciences) for 10 min, followed by centrifugation. Incubations were at room temperature until after RBC lysis and then at 4°C. Samples were washed with 500 µl of FACS buffer, resuspended in 250 µl of FACS fixative (1% paraformaldehyde, 67.5 mM sodium cacodylate, and 113 mM NaCl (pH 7.2)), stored at 4°C in the dark, and washed with 1 ml of PBS and resuspended in 250 µl of FACS buffer just before data collection. Fixation did not decrease signals (data not shown). Data were collected from 30,000 - 170,000 events, using a FACSCalibur (BD Biosciences; available through the Flow Cytometry Facility, Comprehensive Cancer Center, University of Wisconsin, Madison, WI) and CellQuest software (BD Biosciences). Rainbow calibration fluorescent beads (Spherotech) were run each day to calibrate and check the performance of the instrument and channels. Data were analyzed using CellQuest and Flow Jo (Tree Star). Eosinophils were gated based both on scattering (high side scatter) and lack of staining with anti-CD14 and anti-CD16 (30, 35). Thus, the cells that were analyzed for the PE signal fit two criteria for eosinophils by being gated inside both characteristic regions in a plot of side scatter vs FITC staining and a plot of side vs forward scatter. When cells gated on side vs forward scatter were collected by cell sorting followed by cytospin, ≥96% stained for the eosinophil marker eosinophil major basic protein. To analyze circulating neutrophils and monocytes in the same data sets, these leukocyte types were gated inside characteristic regions in the two plots, neutrophils having intermediate side scatter and FITC positivity and monocytes having relatively low side scatter and FITC positivity. Data are expressed as specific geometric mean channel fluorescence (gMCF; specific gMCF = gMCF with a specific integrin mAb – gMCF with isotype control) and as percentage of positive cells (isotype control set with a marker to 2% positive cells) as before (35). For unfractionated BAL cells, flow cytometry was performed with 2 x 105 cells (in 100 µl) and the protocol was as described for blood samples, except that all incubations were at 4°C and the erythrocyte lysis step was omitted and replaced by a wash with 1 ml of PBS.

ELISA for cytokines in BAL fluid

To measure cytokine concentrations, BAL fluid was concentrated 10-fold at 4°C using a low protein-binding Centriprep centrifugal filter unit (Millipore) with a molecular mass cutoff limit of 3 kDa. A sensitive two-step sandwich ELISA was used as described (38). The assay sensitivities were below 3 pg/ml for IL-5 and GM-CSF, 12 pg/ml for IL-3, and 25 pg/ml for IFN-{gamma}. Values are presented as the concentration in recovered BAL fluid before the concentration step.

Statistics

The Mann-Whitney U test was used to compare data between groups. The Spearman rank correlation test was used to analyze correlations. A level of p ≤ 0.05 was considered significant. Analyses were performed using Prism 3.0 (GraphPad).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
Response to whole-lung allergen challenge

All subjects responded to whole-lung inhaled Ag challenge with a drop in FEV1 within 15–30 min to a median value of 31% early FEV1 fall (Table I). Nine of the 19 subjects had a late-phase response (39) as defined by a FEV1 fall ≥15% from baseline 3–8 h after challenge (Table I). Using this criterion (28), the nine subjects were classified as having a dual-response phenotype. The other 10 subjects were considered single responders (Table I). There were no significant differences between the single and dual responders in regards to age, sex, airway responsiveness to methacholine, FEV1 at the screening visit, or early-phase fall in response to whole-lung Ag challenge (Table I). FEV1 48 h after segmental Ag challenge was decreased minimally compared with before segmental Ag challenge and was not significantly different between single and dual responders (medians 98 and 96% of the value before segmental challenge in single and dual responders, respectively).

Median number of BAL eosinophils after segmental challenge was >10-fold higher in the dual responders than in the single responders (Table I). Median numbers of BAL neutrophils and macrophages, in contrast, were ~2-fold higher and not significantly different in dual vs single responders (medians in single and dual responders were 3.1 x 106 and 7.5 x 106 neutrophils and 43 x 106 and 93 x 106 macrophages; respectively). The neutrophil percentage was not significantly different and the macrophage percentage was significantly lower in dual responders due to the increased proportion of eosinophils (medians in single and dual responders were 2.3 and 3.0% neutrophils and 47 and 22% macrophages, respectively).

Integrin expression on blood and BAL eosinophils after segmental allergen challenge

Sample flow cytometry histograms are shown in Fig. 1 for eosinophil expression of total β1 (A), the activation-sensitive β1 epitope for mAb N29 (B–D), total β2 (E), the activation-sensitive β2 epitope for mAb24 (F), {alpha}M (G), and {alpha}D (H). Expression distributions were homogeneous in some samples (Fig. 1, B and F) and heterogeneous and asymmetric with one or more "shoulders" in others (Fig. 1, A, C, D, E, G, and H). β1 distributions were mostly heterogeneous on blood eosinophils and uniformly homogeneous on BAL eosinophils (Fig. 1A and Table II). N29 reactivity of blood eosinophils was variable (Fig. 1, B–D, and Table II). Some samples had two peaks (Fig. 1C) or a "shoulder" of reactivity (Fig. 1D). N29 reactivity of BAL eosinophils was uniformly homogeneous (Fig. 1, B–D, and Table II). β2 was heterogeneous on blood eosinophils, often with two or several distinct populations, whereas the peak of BAL eosinophils was mostly homogeneous and shifted to the right with higher fluorescence intensity compared with the most positive blood eosinophil population (Fig. 1E and Table II). mAb24 reactivity was homogeneous and low on blood eosinophils and mostly homogeneous and shifted to the right on BAL eosinophils (Fig. 1F and Table II). {alpha}M was mostly heterogeneous on blood eosinophils and homogeneous and shifted to the right on BAL eosinophils (Fig. 1G and Table II). {alpha}D was mostly heterogeneous on blood eosinophils and mostly homogeneous on BAL eosinophils (Fig. 1H and Table II). Thus, distributions on BAL eosinophils were typically homogeneous whereas distributions of β1, β2, {alpha}M, and {alpha}D on blood eosinophils were mostly heterogeneous.


Figure 1
View larger version (25K):
[in this window]
[in a new window]

  		FIGURE 1. Integrin expression on blood and BAL eosinophils before and after segmental Ag challenge. Representative flow cytometry histograms of expression of total β1 integrin (A), activation-sensitive β1 integrin mAb N29 epitope (B–D), total β2 integrin (E), activation-sensitive β2 integrin mAb24 epitope (F), {alpha}M integrin (G), or {alpha}D integrin (H) on blood eosinophils before segmental Ag challenge (green), blood eosinophils 48 h after segmental Ag challenge (blue), or BAL eosinophils 48 h after segmental Ag challenge (red), or isotype control (brown). For N29 epitope expression, three examples are shown: one homogeneous, symmetric peak on blood eosinophils before and after challenge and higher mean expression level after challenge than before (B); one peak on blood eosinophils before challenge and two peaks after challenge, and higher mean expression level after challenge (C), and an asymmetric peak with a "shoulder" to the right on blood eosinophils before and after challenge, and decrease in the size of the "shoulder" and lower mean expression level after challenge (D). The numbers of individuals with the various patterns are summarized in Table II.

 

View this table:
[in this window]
[in a new window]

  		Table II. Integrin expression on blood and BAL eosinophils as distribution patterns and percentage of positive cells before and 48 h after segmental Ag challenge

 
Expression data were scored either as percentage of positive cells or as expression level (specific gMCF) (Table II and Fig. 2). Fig. 3 shows a comparison between the results with these two scoring methods on blood eosinophils for N29 (Fig. 3A) and {alpha}D (Fig. 3B). At lower fluorescence intensity the percentage of positive cells and the expression level correlate, whereas at higher intensity the percentage of positive cells plateaus (Fig. 3, A and B). Because of its greater dynamic range, specific gMCF was chosen as the more informative measure of integrin expression and activation (Fig. 2). It should be emphasized that for the samples displaying heterogeneous bimodal or multimodal distributions of the β2 integrin subunits, neither gMCF nor percentage of positive cells fully captures the complexity of the distributions.


Figure 2
View larger version (40K):
[in this window]
[in a new window]

  		FIGURE 2. Integrin expression on blood and BAL eosinophils before and 48 h after segmental Ag challenge. Expression of total β1 integrin (A), activation-sensitive β1 integrin mAb N29 epitope (B), total β2 integrin (C), activation-sensitive β2 integrin mAb24 epitope (D), {alpha}M integrin (E), and {alpha}D integrin (F) on blood eosinophils before segmental Ag challenge (light gray), blood eosinophils 48 h after segmental Ag challenge (medium gray), and BAL eosinophils 48 h after segmental Ag challenge (dark gray) in all tested subjects, single responders, and dual responders. Values shown are specific gMCF (mean ± SEM). For n values and expression given as percentage of positive cells, see Table II. *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001 vs blood eosinophils before segmental challenge; {dagger}, p ≤ 0.05; {dagger}{dagger}{dagger}, p ≤ 0.001 vs blood eosinophils after segmental challenge; {ddagger}, p ≤ 0.05 vs single responders.

 

Figure 3
View larger version (9K):
[in this window]
[in a new window]

  		FIGURE 3. Integrin expression on blood eosinophils as the percentage of positive cells and the expression level. Blood eosinophil expression of activation-sensitive β1 integrin mAb N29 epitope (A) or {alpha}D integrin (B) before (open symbols) and 48 h after (filled symbols) segmental Ag challenge, expressed as the percentage of positive cells (y axes) or as expression level (specific gMCF) (x axes).

 
On circulating eosinophils of dual responders the expression level of the epitope for activation-sensitive anti-β1 mAb N29 (34, 40, 41) was higher 48 h after Ag segmental challenge as compared with the level before challenge (Fig. 2B). Circulating eosinophils also had significantly higher expression of the {alpha}D integrin subunit compared with before segmental challenge (Fig. 2F). {alpha}D and N29 epitope expression on blood eosinophils 48 h after segmental challenge correlated with each other (rs (Spearman rank correlation coefficient) = 0.73; p = 0.002). Thus, the results demonstrate that segmental Ag challenge causes significantly increased surface expression of the {alpha}D subunit of β2 integrins and, in dual responders, activation of β1 integrins at 48 h on blood eosinophils.

Eosinophils in BAL 48 h after segmental Ag challenge had significantly higher β2 and {alpha}M subunit expression than eosinophils in blood obtained before or 48 h after segmental challenge (Fig. 2, C and E). The expression of the epitope for the activation-sensitive anti-β2 mAb24 (32, 33) was significantly increased on BAL eosinophils; the binding of mAb24 to blood eosinophils was low in blood both before and after segmental challenge (Fig. 2D). {alpha}D expression was significantly increased on BAL eosinophils as compared with blood eosinophils before challenge but not as compared with blood also sampled at 48 h (Fig. 2F). In dual responders, BAL eosinophils had significantly more total and activated β1 (as ascertained by N29) than blood eosinophils before challenge (Fig. 2, A and B). {alpha}6 expression was lower on BAL eosinophils than on blood eosinophils after challenge (specific gMCF = 560 ± 140 (mean ± SEM) vs 920 ± 110, p ≤ 0.05)(data not shown). No differences were found in the expression of {alpha}L, {alpha}X, {alpha}4, or β7 (data not shown). Overall, the results indicate that {alpha}Mβ2 is up-regulated on BAL eosinophils and that BAL eosinophil β2 integrins are in a conformationally altered and activated state compared with β2 integrins on blood eosinophils.

To examine the specificity of the changes on blood eosinophils, circulating neutrophils and monocytes were also analyzed. Before segmental challenge, neutrophils had higher N29 reactivity than eosinophils and monocytes had even higher N29 reactivity (Figs. 4B and 5B; compare with Fig. 2B). At 48 h after segmental Ag challenge, circulating neutrophils and monocytes, like eosinophils, had significantly higher {alpha}D expression compared with before challenge (Figs. 4F and 5F). Monocytes from dual responders had higher {alpha}D after segmental challenge than those from single responders (Fig. 5F). N29 reactivity of neutrophils (as with eosinophils) increased significantly in dual responders upon segmental challenge, although to a lesser degree (1.4-fold for neutrophils, 1.7-fold for eosinophils) (Fig. 4B; compare with Fig. 2B), whereas N29 reactivity of monocytes did not increase significantly (Fig. 5B). Furthermore, {alpha}M on blood neutrophils and monocytes and β2 on blood monocytes increased significantly upon segmental challenge (Figs. 4E and 5, C and E) in contrast to blood eosinophil {alpha}M and β2, which did not (Fig. 2C,E).


Figure 4
View larger version (40K):
[in this window]
[in a new window]

  		FIGURE 4. Integrin expression on blood neutrophils before and 48 h after segmental Ag challenge. Expression of total β1 integrin (A), activation-sensitive β1 integrin mAb N29 epitope (B), total β2 integrin (C), activation-sensitive β2 integrin mAb24 epitope (D), {alpha}M integrin (E), and {alpha}D integrin (F) on blood neutrophils before segmental Ag challenge (light gray) and 48 h after segmental Ag challenge (medium gray) in all tested subjects, single responders, and dual responders. Values shown are specific gMCF (mean ± SEM). *, p ≤ 0.05 vs before segmental challenge.

 

Figure 5
View larger version (38K):
[in this window]
[in a new window]

  		FIGURE 5. Integrin expression on blood monocytes before and 48 h after segmental Ag challenge. Expression of total β1 integrin (A), activation-sensitive β1 integrin mAb N29 epitope (B), total β2 integrin (C), activation-sensitive β2 integrin mAb24 epitope (D), {alpha}M integrin (E), and {alpha}D integrin (F) on blood monocytes before segmental Ag challenge (light gray) and 48 h after segmental Ag challenge (medium gray) in all tested subjects, single responders, and dual responders. Values shown are specific gMCF (mean ± SEM). *, p ≤ 0.05; **, p ≤ 0.01 vs before segmental challenge; {ddagger}, p ≤ 0.05; {ddagger}{ddagger}, p ≤ 0.01 vs single responders.

 
Thus, we determined the following: 1) {alpha}D is up-regulated on all three leukocyte populations, not just eosinophils; 2) β1 at baseline is more activated on neutrophils and monocytes compared with eosinophils; 3) in dual responders we found an increased activation state of β1 on neutrophils despite the high baseline activation state; and 4) surface expression of {alpha}M is up-regulated on monocytes and neutrophils, but not on blood eosinophils.

Correlations between integrin expression and eosinophil numbers in BAL after segmental allergen challenge

As with the variability in eosinophil numbers in BAL after segmental Ag challenge in the 19 subjects (Table I), there was considerable variability in integrin activation or expression on BAL eosinophils (Fig. 2). BAL eosinophils from dual responders had significantly higher N29 epitope and total β2 expression than BAL eosinophils from single responders (Figs. 2 and 6). Grouping all subjects together, the reactivity of BAL eosinophils with the activation-sensitive mAbs, N29 against β1 and mAb24 against β2, correlated significantly with the percentage of eosinophils in BAL (Fig. 7). Surface expression of other integrin subunits on BAL eosinophils did not correlate with BAL eosinophil percentage (data not shown). These results indicate that activation of both β1 and β2 integrins are associated with eosinophil recruitment to the airway.


Figure 6
View larger version (8K):
[in this window]
[in a new window]

  		FIGURE 6. Differences in integrin activation and expression on BAL eosinophils between single and dual responders. BAL eosinophil expression 48 h after segmental Ag challenge of activation-sensitive β1 integrin mAb N29 epitope (A) or total β2 integrin (B) in single and dual responders. Bar, mean; *. p ≤ 0.05 vs single responders.

 

Figure 7
View larger version (10K):
[in this window]
[in a new window]

  		FIGURE 7. Correlations between integrin activation on BAL eosinophils and BAL eosinophil percentage. Correlations between BAL eosinophil expression of activation-sensitive β1 integrin mAb N29 epitope (A) or activation-sensitive β2 integrin mAb24 epitope (B) and the percentage of eosinophils in BAL 48 h after segmental Ag challenge. rs, Spearman rank correlation coefficient.

 
Correlations between integrin expression after segmental allergen challenge and the magnitude of the late-phase fall in FEV1 after whole-lung allergen challenge

We also analyzed for possible correlations between eosinophil integrins after segmental challenge and the magnitude of the maximum fall in FEV1 3–8 h after whole-lung allergen challenge (late-phase fall). {alpha}D expression and reactivity with N29 of blood eosinophils 48 h after segmental challenge each correlated with the magnitude of the late-phase fall in FEV1 after whole-lung challenge (Fig. 8, A and B). β2 expression and reactivity with mAb24 of BAL eosinophils 48 h after segmental challenge also correlated with the magnitude of the late-phase FEV1 fall (Fig. 8, C and D), as did {alpha}M expression of BAL eosinophils (rs = 0.83, p = 0.008) (data not shown). In addition, the late-phase FEV1 fall correlated with the percentage of eosinophils in BAL 48 h after segmental challenge (rs = 0.70, p = 0.001) (data not shown). Thus, the results indicate that greater {alpha}D up-regulation and β1 activation on blood eosinophils and greater {alpha}Mβ2 up-regulation and β2 activation on BAL eosinophils after segmental challenge in a subject are associated with a greater late-phase FEV1 fall in response to whole-lung challenge.


Figure 8
View larger version (16K):
[in this window]
[in a new window]

  		FIGURE 8. Correlations between integrin activation and expression on blood and BAL eosinophils and the magnitude of the fall in FEV1 3–8 h after whole-lung Ag challenge. Correlations between blood eosinophil expression of activation-sensitive β1 integrin mAb N29 epitope (A) or {alpha}D integrin (B) or BAL eosinophil expression of total β2 integrin (C) or activation-sensitive β2 integrin mAb24 epitope (D) 48 h after segmental Ag challenge and the magnitude of the late-phase fall in FEV1 3–8 h after whole-lung Ag challenge. rs, Spearman rank correlation coefficient.

 
Correlations between integrin expression and cytokine concentrations in BAL fluid after segmental allergen challenge

To identify factor(s) possibly responsible for the changes in integrins observed following segmental Ag challenge, the concentrations of IL-5, GM-CSF, IL-3, and IFN-{gamma} in BAL fluid were measured (Fig. 9). The concentration of all four cytokines was significantly higher 48 h after segmental challenge than in samples obtained immediately before segmental challenge (Fig. 9). There was a highly significant difference in BAL fluid IL-5 after segmental challenge between single and dual responders; median IL-5 concentration in BAL fluid from dual responders was ~40-fold greater than in BAL fluid from single responders (Fig. 9). Also IL-3 and IFN-{gamma} were significantly different between single and dual responders; medians from dual responders were ~10- and ~3-fold those from single responders for IL-3 and IFN-{gamma}, respectively (Fig. 9). GM-CSF was not different between single and dual responders (Fig. 9).


Figure 9
View larger version (40K):
[in this window]
[in a new window]

  		FIGURE 9. Cytokine concentrations in BAL fluid before and 48 h after segmental Ag challenge. Concentrations of IL-5 (A), GM-CSF (B), IL-3 (C), and IFN-{gamma} (D) in BAL fluid before (light gray bars) and 48 h after (medium gray bars) segmental Ag challenge from all tested subjects (n = 19), single responders (n = 10), or dual responders (n = 9). Values shown are medians with 25th and 75th percentiles of concentrations in recovered, unconcentrated BAL fluid. *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001 vs before segmental challenge; {ddagger}, p ≤ 0.05; {ddagger}{ddagger}{ddagger}, p ≤ 0.001 vs single responders.

 
The concentration of BAL fluid IL-5, but not that of the other cytokines, correlated significantly and strongly with the percentage of eosinophils in BAL 48 h after segmental Ag challenge (rs = 0.69, p = 0.001 for IL-5; rs = 0.23, p = 0.36 for GM-CSF; rs = 0.32, p = 0.19 for IL-3; and rs = 0.39, p = 0.11 for IFN-{gamma}). It is possible that one or both of the other IL-5 family cytokines would correlate with the BAL eosinophil percentage if a higher number of subjects were studied. IL-5 also correlated inversely with the percentage of macrophages (data not shown), reflecting the decreased macrophage percentage as eosinophil percentage increases. Further, BAL fluid IL-5 after segmental challenge correlated strongly with the magnitude of the late-phase fall in FEV1 after whole-lung challenge (rs = 0.79, p < 0.0001). IL-3 also correlated with late-phase FEV1 fall, but less well (rs = 0.51, p = 0.03). GM-CSF (rs = 0.37, p = 0.12) and IFN-{gamma} (rs = 0.36, p = 0.13) did not correlate with late-phase FEV1 fall.

Levels of β2 and reactivity with mAb24 of BAL eosinophils correlated significantly with the concentration of IL-5 in BAL fluid 48 h after segmental Ag challenge (Fig. 10), as did level of {alpha}M (rs = 0.70, p = 0.04) (data not shown). These integrins did not correlate with concentrations of the other cytokines (data not shown). There was no correlation with BAL fluid IL-5 before segmental challenge (data not shown). Integrins of blood eosinophils did not correlate with cytokine concentrations in BAL fluid 48 h after segmental Ag challenge (data not shown). Thus, a greater IL-5 concentration in BAL fluid in a subject is associated with greater {alpha}Mβ2 up-regulation and β2 activation on BAL eosinophils after segmental challenge.


Figure 10
View larger version (8K):
[in this window]
[in a new window]

  		FIGURE 10. Correlations between integrin activation and expression on BAL eosinophils and IL-5 in BAL fluid. Correlations between blood eosinophil expression of total β2 integrin (A) or activation-sensitive β2 integrin mAb24 epitope (B) and the concentration of IL-5 in BAL fluid 48 h after segmental Ag challenge. rs, Spearman rank correlation coefficient.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
We observed changes in integrins on eosinophils in the blood and airways of 19 subjects with mild allergic asthma after segmental bronchial allergen challenge that were related to the reactivities of subjects in the whole Ag challenge protocol. Eosinophils in blood 48 h after challenge had increased expression of the {alpha}D integrin subunit and, in asthmatics with a dual response phenotype, increased reactivity with the activation-sensitive anti-β1 integrin mAb N29 compared with before challenge. Eosinophils in BAL obtained 48 h after challenge had increased expression of β2 and {alpha}M integrin subunits and increased reactivity of activation-sensitive anti-β2 integrin mAb24 compared with eosinophils in blood. β2, {alpha}M, and mAb24 epitope expression on BAL eosinophils correlated with the concentration of IL-5 in BAL fluid obtained 48 h after segmental Ag challenge. In addition, {alpha}D expression and, in dual responders, N29 reactivity were higher on BAL eosinophils than on blood eosinophils before challenge. Integrin expression patterns on BAL eosinophils were more homogeneous and symmetric than on blood eosinophils. The observations of N29 epitope and {alpha}D expression are compatible with a scenario whereby extravasated eosinophils appearing in the airway are derived from a subpopulation of the total, more heterogeneous population of circulating eosinophils. However, the observations of {alpha}Mβ2 expression, β2 activation state, and IL-5 levels in BAL fluid indicate that, after leaving the circulation, eosinophils undergo IL-5-triggered activation of β2 and up-regulation of {alpha}Mβ2. Analysis of neutrophils and monocytes in the same blood samples revealed that {alpha}D expression increased on all three leukocyte types upon segmental challenge; thus, {alpha}D up-regulation may occur through a common mechanism. At baseline, N29 reactivity was higher on neutrophils than on eosinophils and even higher on monocytes. N29 reactivity increased significantly on neutrophils in dual responders upon challenge but to a relatively lesser degree than on eosinophils, whereas it did not change significantly on monocytes. The mechanisms responsible for maintaining the baseline β1 activation levels on the different leukocytes and those responsible for the elevation of activation state upon challenge are unknown. Finally, neutrophil and monocyte {alpha}M and monocyte β2 were up-regulated upon challenge, in contrast to eosinophil {alpha}M and β2, which were up-regulated in BAL but not in blood. Thus, circulating eosinophils are less sensitive than neutrophils or monocytes to increased surface expression of {alpha}Mβ2.

β1 activation on blood eosinophils, assessed by reactivity with N29, has been shown to correlate inversely with FEV1 in an inhaled corticosteroid withdrawal study (35). The present results complement the steroid withdrawal study by demonstrating persistent β1 activation in dual responders with more eosinophilic inflammation in the airway. Blood eosinophils from dual responders, but not from single responders, have also been shown to have enhanced activation of Fc{gamma}RII (CD32) as assessed with the phage mAb A17 6 h after challenge (42). Whether activation of β1 integrin and CD32 on circulating eosinophils are triggered by the same or different stimuli and signaling pathways is an interesting question that remains to be investigated.

The changes observed here in BAL eosinophils reproduce observations in the literature, made mostly on isolated eosinophils, that BAL eosinophils have higher {alpha}M and β2 than blood eosinophils (43, 44, 45, 46), activated {alpha}Mβ as monitored by anti-active {alpha}M mAb CBRM1/5 (22, 36), and increased {alpha}Mβ2-inhibitable adhesion to diverse ligands (22, 36). Our finding that dual responders have higher total numbers of cells and eosinophils and a higher proportion of eosinophils in BAL after segmental Ag challenge than single responders also is in accord with prior reports (47, 48, 49, 50). We also found that the β1 activation state and β2 surface expression were significantly higher on BAL eosinophils from dual responders than on those from single responders. Activation of β1 and β2 integrins, as assessed with the activation-sensitive mAbs N29 and mAb24, respectively, on BAL eosinophils correlated with eosinophil percentage in BAL, indicating that activation of both of these subfamilies of integrins is important for eosinophil recruitment. This idea is consistent with in vivo studies supporting the involvement of {alpha}4β1 in eosinophil appearance in the airway and with in vitro studies showing the involvement of both β1 and β2 integrins in eosinophil transendothelial migration (22, 51, 52, 53, 54, 55, 56). The evidence for involvement of both {alpha}4β1 and β2 integrins in eosinophil migration in vivo is further strengthened by a recent report that eosinophil recruitment to airway after Ag challenge was severely attenuated in both a conditional {alpha}4 integrin knockout mouse and a β2-deficient mouse (57). Finally, {alpha}D expression and β1 activation on blood eosinophils as well as β2 and {alpha}M expression and β2 activation on BAL eosinophils after segmental Ag challenge all correlated with the magnitude of the late-phase fall in FEV1 in response to whole-lung Ag challenge.

A number of mediators are increased in the airway after challenge (28, 29, 49, 58, 59). Of these, the most likely candidates to account for the up-regulation and activation of {alpha}Mβ2 are the IL-5 family cytokines (36, 60, 61). Concentrations of IL-5, GM-CSF, IL-3, and IFN-{gamma} in BAL fluid recovered 48 h after segmental Ag challenge were all increased significantly compared with those in BAL fluid recovered before challenge. IL-5 correlated with BAL eosinophil {alpha}M and β2 expression and β2 activation 48 h after segmental challenge. The median concentration of IL-5 in BAL fluid from dual responders after segmental challenge was ~600 pg/ml, which was >20-fold the median concentration of the other cytokines. Further, the IL-5 concentration in the five BAL samples containing eosinophils with the most highly activated β2 was 100 to >1000 pg/ml. During the recovery of BAL, the volume of the fluid lining the airway epithelium in vivo is estimated to become diluted 100-fold (62). Thus, the levels of IL-5 in the epithelial lining fluid of these subjects (all of whom were dual responders) are estimated to be ~10–100 ng/ml. Treatment of blood eosinophils in vitro with IL-5 at concentrations in this range is known to saturate the IL-5 receptor (63, 64, 65, 66, 67, 68) and lead to {alpha}Mβ2 up-regulation and activation and induction of adhesion to ICAM-1 and other substrates (36, 60, 61), priming and enhanced response to chemoattractants (69), enhanced viability (70), degranulation and granule protein release (30), IL-5 receptor {alpha} down-regulation (71), and enhanced expression of certain genes (72). In contrast, median concentrations of the other IL-5 family cytokines before dilution are estimated to be ~1–3 ng/ml. Thus, the concentration of IL-5 in the lining fluid in vivo, assuming that a significant portion of the IL-5 measured represents active IL-5, is estimated to be sufficient to cause {alpha}Mβ2 up-regulation and activation of β2 integrins. These results are in accord with the earlier observation that the IL-5 receptor is down-regulated specifically compared with the GM-CSF receptor on eosinophils recovered by BAL 48 h after segmental Ag challenge (30). We also found that BAL fluid IL-5 correlated with the percentage of eosinophils in BAL and with the magnitude of the late-phase fall in FEV1. A correlation between BAL fluid IL-5 and eosinophil recruitment to the airway after segmental Ag challenge has been reported previously (38, 73, 74, 75, 76) and is consistent with the observation that anti-IL-5 therapy causes a significant decrease in sputum eosinophils (1).

Our study has several limitations. The data sets were incomplete in that all integrins were not assayed from all samples. Sampling of blood and BAL was performed only at one time point (48 h) after segmental Ag challenge. Future experiments with blood sampling at various time points after segmental and/or whole-lung Ag challenge are required to record the time course of {alpha}D and N29 expression on blood eosinophils in dual and single responders and to learn when the values diverge. Further, the importance of the integrin expression heterogeneity observed on blood eosinophils, particularly regarding β2 and {alpha}M, is not known. Such heterogeneity, to our knowledge, has not been described or discussed before. Of note, we subjected whole blood to primary Ab incubation without previous cell isolation or centrifugation. Earlier studies on blood eosinophil integrin expression have been performed on a buffy coat preparation or purified eosinophils (43, 44, 45). Dextran sedimentation during buffy coat preparation has been shown to cause up-regulation of {alpha}M on blood eosinophils (and neutrophils) compared to that in whole, unfractionated blood (77). Eosinophils are likely similar to neutrophils, which are known to store a large proportion of {alpha}M in granules, wherefrom it is translocated to the plasma membrane after cell stimulation (78). Remarkably, eosinophils in BAL had strong homogeneous labeling for β2 and {alpha}M. One possibility is that higher reacting subpopulations of circulating leukocytes represent cells that have undergone "retrograde" migration back to the blood from tissues, as has been demonstrated for zebrafish neutrophils in vivo (79) and human neutrophils in vitro (80). Blood sampling at various time points after Ag challenge, including times beyond 48 h, may shed further light on circulating eosinophil subpopulations.

Our results and the literature are compatible with the schematic of eosinophil recruitment after segmental Ag challenge that is depicted in Fig. 11. At baseline, β1 and β2 integrins on a circulating eosinophil are in a low activation state, the level of eosinophil surface {alpha}D is low, and VCAM-1 is absent from endothelial surfaces of the bronchial circulation. Yet to be identified stimuli resulting from Ag challenge cause enhanced activation of β1 integrins and increased surface expression of {alpha}D on circulating eosinophils. Because increased expression of the β1 activation epitope occurs on most eosinophils, the activation likely takes place in the pulmonary circulation through which the eosinophils constantly pass or in response to the release of an activating substance into the circulation (Fig. 11). Such activation contrasts with the model for the recruitment of leukocytes (16), including eosinophils (20), in which integrin activation is assumed to occur locally and concurrently with rolling and tethering on endothelium. Interestingly, bone marrow-derived progenitor cells or circulating eosinophils of asthmatics have been shown to be activated upon Ag challenge also as measured by the up-regulation of IL-5 receptor {alpha} and CCR3, the activation of CD32, and greater responsiveness to chemoattractants (42, 81, 82, 83). Further, eosinophils from allergic asthmatics have been shown to have a greater capacity to adhere to and transmigrate through endothelium than eosinophils from normal donors (51, 84).


Figure 11
View larger version (29K):
[in this window]
[in a new window]

  		FIGURE 11. Sequential up-regulation and activation of eosinophil integrins during recruitment to airway after exposure to allergen. Schematic relating activation of β1 and β2 integrins to eosinophil trafficking during and after segmental Ag challenge. Left, An eosinophil (oval) with unactivated β1 integrins and low surface expression of {alpha}D is shown entering the pulmonary circulation of a segment subjected to Ag challenge. A yet to be identified stimulus or stimuli (X) cause(s) activation of β1 integrins and increased surface expression of {alpha}D during transit of the eosinophil (rectangle) through the vessel. Right, Concurrently, IL-4 and other mediators are released and specifically induce surface expression of VCAM-1 (discontinuous black line) on the endothelial cells of a bronchial blood vessel in the challenged segment. An eosinophil with activated β1 and up-regulated {alpha}D (rectangle) is shown entering the bronchial circulation. The eosinophil arrests and adheres (elongated rectangle) to VCAM-1. After exposure to IL-5 the eosinophil becomes responsive to chemotactic factors, migrates (arrow), and assumes a hyperadhesive airway phenotype (hexagon) in the lumen. The airway eosinophil displays activated β1, up-regulated {alpha}D, activated β2, and up-regulated {alpha}M and β2.

 
The question then arises as to how activated circulating eosinophils localize to the parts of the lung subjected to Ag challenge in preference to neutrophils and monocytes. As a parallel response to Ag challenge, mediators, including IL-4 and IL-13, are elaborated and activate the bronchial endothelium to specifically synthesize and express VCAM-1. VCAM-1 has been shown to be preferentially expressed in the asthmatic lung or after Ag challenge (85, 86). Further, VCAM-1 can support the adhesion of eosinophils but not neutrophils (9, 18, 22, 87, 88, 89, 90, 91). Circulating eosinophils with activated β1 and up-regulated {alpha}D are presumed to have a higher probability of arresting on VCAM-1 on the activated endothelium of the bronchial circulation (Fig. 11), because adhesion of purified blood eosinophils to VCAM-1 is mediated by {alpha}4β1 (22, 92), with a possible contribution by {alpha}Dβ2 (22, 31).

{alpha}M andβ2 are shown as being up-regulated and β2 integrins as being activated by IL-5, also elaborated in response to allergen. Eosinophils appearing in the airway lumen have a hyperadhesive phenotype that is marked by activated β1 integrins, up-regulated {alpha}Dβ2, activated β2 integrins, and up-regulated {alpha}Mβ2 (Fig. 11). Eosinophils, particularly after priming by IL-5 (69), responds to chemoattractants, such as eotaxin (9, 13) (Fig. 11). {alpha}Mβ2 is believed to be important for eosinophil migration (22, 51, 53, 93, 94, 95). Thus, the scenario has migrating eosinophils using activated {alpha}Mβ2 and possibly other integrins to interact with multiple substrates (22, 36, 92), including VCAM-1, ICAM-1, and extracellular matrix proteins, on endothelial cells, in connective tissue, and on epithelium of the bronchial wall.


   Acknowledgments
 
We thank Mary Jo Jackson and Erin Billmeyer for patient recruitment, screening, and assistance with bronchoscopy; Lin-Ying Liu, Sarah Panzer, Rebecca Lawniczak, and Rose DeGrauw for processing BAL samples and providing blood samples; Melissa Heim and Lisa Skoggs for assistance with ELISA; Nancy Hogg for providing mAb24; ICOS for providing anti-{alpha}D mAb 240I; Kathleen Schell, Joan Batchelder, and Joel Puchalski for help with flow cytometry data collection and analysis; Michael Evans for advice on statistics; and Julie Sedgwick and Lin-Ying Liu for advice.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
M.W.J., W.W.B., and D.F.M. have, with the help of the Wisconsin Alumni Research Foundation, a patent application pending on "Beta1 integrin activation as a marker for asthma," based on the associations between N29 reactivity and FEV1 reported by Johansson et al. 2006 (Ref. 35). W.W.B. has or has had research support from Novartis, Dynavax, Wyeth, Centocor, GlaxoSmithKline, Medicinova, Pfizer, and Biowa/MedImmune. E.A.B.K. and N.N.J. have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This work was supported by Specialized Center of Research Grant HL56396, Program Project Grant HL88594, and General Clinical Research Center Grant M01 RR03186 from the National Institutes of Health. Back

2 Address correspondence to Dr. Mats W. Johansson, Department of Biomolecular Chemistry, University of Wisconsin, 4285A Medical Sciences Center, 1300 University Avenue, Madison, WI 53706. E-mail address: mwjohansson{at}wisc.edu Back

3 Abbreviations used in this paper: BAL, bronchoalveolar lavage; AgPD20, provocative dose of Ag producing a 20% fall in FEV1; FEV1, forced expiratory volume in 1 s; gMCF, geometric mean channel fluorescence. Back

Received for publication November 20, 2007. Accepted for publication March 21, 2008.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

  1. Leckie, M. J., A. ten Brinke, J. Khan, Z. Diamant, B. J. O'Connor, C. M. Walls, A. K. Mathur, H. C. Cowley, K. F. Chung, R. Djukanovic, et al 2000. Effects of an interleukin-5 blocking monoclonal antibody on eosinophils, airway hyper-responsiveness, and the late asthmatic response. Lancet 356: 2144-2148. [Medline]
  2. Busse, W. W., R. F. Lemanske, Jr. 2001. Asthma. N. Engl. J. Med. 344: 350-362. [Free Full Text]
  3. Green, R. H., C. E. Brightling, S. McKenna, B. Hargadon, D. Parker, P. Bradding, A. J. Wardlaw, I. D. Pavord. 2002. Asthma exacerbations and sputum eosinophil counts: a randomised controlled trial. Lancet 360: 1715-1721. [Medline]
  4. Mattes, J., M. Yang, S. Mahalingam, J. Kuehr, D. C. Webb, L. Simson, S. P. Hogan, A. Koskinen, A. N. McKenzie, L. A. Dent, et al 2002. Intrinsic defect in T cell production of interleukin (IL)-13 in the absence of both IL-5 and eotaxin precludes the development of eosinophilia and airways hyperreactivity in experimental asthma. J. Exp. Med. 195: 1433-1444. [Abstract/Free Full Text]
  5. Flood-Page, P. T., A. N. Menzies-Gow, A. B. Kay, D. S. Robinson. 2003. Eosinophil’s role remains uncertain as anti-interleukin-5 only partially depletes numbers in asthmatic airway. Am. J. Respir. Crit. Care Med. 167: 199-204. [Abstract/Free Full Text]
  6. Flood-Page, P., A. Menzies-Gow, S. Phipps, S. Ying, A. Wangoo, M. S. Ludwig, N. Barnes, D. Robinson, A. B. Kay. 2003. Anti-IL-5 treatment reduces deposition of ECM proteins in the bronchial subepithelial basement membrane of mild atopic asthmatics. J. Clin. Invest. 112: 1029-1036. [Medline]
  7. Justice, J. P., M. T. Borchers, J. R. Crosby, E. M. Hines, H. H. Shen, S. I. Ochkur, M. P. McGarry, N. A. Lee, J. J. Lee. 2003. Ablation of eosinophils leads to a reduction of allergen-induced pulmonary pathology. Am. J. Physiol. 284: L169-L178.
  8. Shen, H. H., S. I. Ochkur, M. P. McGarry, J. R. Crosby, E. M. Hines, M. T. Borchers, H. Wang, T. L. Biechelle, K. R. O'Neill, T. L. Ansay, et al 2003. A causative relationship exists between eosinophils and the development of allergic pulmonary pathologies in the mouse. J. Immunol. 170: 3296-3305. [Abstract/Free Full Text]
  9. Wills-Karp, M., C. L. Karp. 2004. Eosinophils in asthma: remodeling a tangled tale. Science 305: 1726-1729. [Abstract/Free Full Text]
  10. Bochner, B. S., W. W. Busse. 2005. Allergy and asthma. J. Allergy Clin. Immunol. 115: 953-959. [Medline]
  11. Kay, A. B.. 2005. The role of eosinophils in the pathogenesis of asthma. Trends Mol. Med. 11: 148-152. [Medline]
  12. Liu, L. Y., S. K. Mathur, J. B. Sedgwick, N. N. Jarjour, W. W. Busse, E. A. Kelly. 2006. Human airway and peripheral blood eosinophils enhance Th1 and Th2 cytokine secretion. Allergy 61: 589-597. [Medline]
  13. Rothenberg, M. E., S. P. Hogan. 2006. The eosinophil. Annu. Rev. Immunol. 24: 147-174. [Medline]
  14. Kariyawasam, H. H., D. S. Robinson. 2007. The role of eosinophils in airway tissue remodelling in asthma. Curr. Opin. Immunol. 19: 681-686. [Medline]
  15. Liu, L. Y., M. E. Bates, N. N. Jarjour, W. W. Busse, P. J. Bertics, E. A. Kelly. 2007. Generation of Th1 and Th2 chemokines by human eosinophils: evidence for a critical role of TNF-{alpha}. J. Immunol. 179: 4840-4848. [Abstract/Free Full Text]
  16. Springer, T. A.. 1994. Traffic signals for lymphocyte recirculation and leukocyte emigration: the multistep paradigm. Cell 76: 301-314. [Medline]
  17. Hynes, R. O.. 2002. Integrins: bidirectional, allosteric signaling machines. Cell 110: 673-687. [Medline]
  18. Broide, D., P. Sriramarao. 2001. Eosinophil trafficking to sites of allergic inflammation. Immunol. Rev. 179: 163-172. [Medline]
  19. Wardlaw, A. J.. 2001. Eosinophil trafficking in asthma. Clin. Med. 1: 214-218. [Medline]
  20. Rosenberg, H. F., S. Phipps, P. S. Foster. 2007. Eosinophil trafficking in allergy and asthma. J. Allergy Clin. Immunol. 119: 1303-1310. [Medline]
  21. Tachimoto, H., B. S. Bochner. 2000. The surface phenotype of human eosinophils. Chem. Immunol. 76: 45-62. [Medline]
  22. Barthel, S. R., M. W. Johansson, D. M. McNamee, D. F. Mosher. 2008. Roles of integrin activation in eosinophil function and the eosinophilic inflammation of asthma. J. Leukocyte Biol. 83: 1-12. [Abstract/Free Full Text]
  23. Humphries, J. D., A. Byron, M. J. Humphries. 2006. Integrin ligands at a glance. J. Cell Sci. 119: 3901-3903. [Free Full Text]
  24. Palecek, S. P., J. C. Loftus, M. H. Ginsberg, D. A. Lauffenburger, A. F. Horwitz. 1997. Integrin-ligand binding properties govern cell migration speed through cell-substratum adhesiveness. Nature 385: 537-540. [Medline]
  25. Humphries, M. J.. 2004. Monoclonal antibodies as probes of integrin priming and activation. Biochem. Soc. Trans. 32: 407-411. [Medline]
  26. Xiao, T., J. Takagi, B. S. Coller, J. H. Wang, T. A. Springer. 2004. Structural basis for allostery in integrins and binding to fibrinogen-mimetic therapeutics. Nature 432: 59-67. [Medline]
  27. Calhoun, W. J., N. N. Jarjour, G. J. Gleich, C. A. Stevens, W. W. Busse. 1993. Increased airway inflammation with segmental versus aerosol antigen challenge. Am. Rev. Respir. Dis. 147: 1465-1471. [Medline]
  28. Liu, L., N. N. Jarjour, W. W. Busse, E. A. Kelly. 2004. Enhanced generation of helper T type 1 and 2 chemokines in allergen-induced asthma. Am. J. Respir. Crit. Care Med. 169: 1118-1124. [Abstract/Free Full Text]
  29. Kelly, E. A., W. W. Busse, N. N. Jarjour. 2000. Inhaled budesonide decreases airway inflammatory response to allergen. Am. J. Respir. Crit. Care Med. 162: 883-890. [Abstract/Free Full Text]
  30. Liu, L. Y., J. B. Sedgwick, M. E. Bates, R. F. Vrtis, J. E. Gern, H. Kita, N. N. Jarjour, W. W. Busse, E. A. Kelly. 2002. Decreased expression of membrane IL-5 receptor {alpha} on human eosinophils: I. Loss of membrane IL-5 receptor {alpha} on airway eosinophils and increased soluble IL-5 receptor {alpha} in the airway after allergen challenge. J. Immunol. 169: 6452-6458. [Abstract/Free Full Text]
  31. Grayson, M. H., M. Van der Vieren, S. A. Sterbinsky, M. W. Gallatin, P. A. Hoffman, D. E. Staunton, B. S. Bochner. 1998. {alpha}dβ2 integrin is expressed on human eosinophils and functions as an alternative ligand for vascular cell adhesion molecule 1 (VCAM-1). J. Exp. Med. 188: 2187-2191. [Abstract/Free Full Text]
  32. Dransfield, I., N. Hogg. 1989. Regulated expression of Mg2+ binding epitope on leukocyte integrin {alpha} subunits. EMBO J. 8: 3759-3765. [Medline]
  33. Lu, C., M. Shimaoka, Q. Zang, J. Takagi, T. A. Springer. 2001. Locking in alternate conformations of the integrin {alpha}Lβ2 I domain with disulfide bonds reveals functional relationships among integrin domains. Proc. Natl. Acad. Sci. USA 98: 2393-2398. [Abstract/Free Full Text]
  34. Wilkins, J. A., A. Li, H. Ni, D. G. Stupack, C. Shen. 1996. Control of β1 integrin function: localization of stimulatory epitopes. J. Biol. Chem. 271: 3046-3051. [Abstract/Free Full Text]
  35. Johansson, M. W., S. R. Barthel, C. A. Swenson, M. D. Evans, N. N. Jarjour, D. F. Mosher, W. W. Busse. 2006. Eosinophil β1 integrin activation state correlates with asthma activity in a blind study of inhaled corticosteroid withdrawal. J. Allergy Clin. Immunol. 117: 1502-1504. [Medline]
  36. Barthel, S. R., N. N. Jarjour, D. F. Mosher, M. W. Johansson. 2006. Dissection of the hyperadhesive phenotype of airway eosinophils in asthma. Am. J. Respir. Cell Mol. Biol. 35: 378-386. [Abstract/Free Full Text]
  37. Leitinger, B., N. Hogg. 2000. Effects of I domain deletion on the function of the β2 integrin lymphocyte function-associated antigen-1. Mol. Biol. Cell 11: 677-690. [Abstract/Free Full Text]
  38. Kelly, E. A., R. R. Rodriguez, W. W. Busse, N. N. Jarjour. 1997. The effect of segmental bronchoprovocation with allergen on airway lymphocyte function. Am. J. Respir. Crit. Care Med. 156: 1421-1428. [Abstract/Free Full Text]
  39. Lemanske, R. F., Jr. 1991. The late phase response: clinical implications. Adv. Intern. Med. 36: 171-193. [Medline]
  40. Ni, H., A. Li, N. Simonsen, J. A. Wilkins. 1998. Integrin activation by dithiothreitol or Mn2+ induces a ligand-occupied conformation and exposure of a novel NH2-terminal regulatory site on the β1 integrin chain. J. Biol. Chem. 273: 7981-7987. [Abstract/Free Full Text]
  41. Mould, A. P., M. A. Travis, S. J. Barton, J. A. Hamilton, J. A. Askari, S. E. Craig, P. R. Macdonald, R. A. Kammerer, P. A. Buckley, M. J. Humphries. 2005. Evidence that monoclonal antibodies directed against the integrin β subunit plexin/semaphorin/integrin domain stimulate function by inducing receptor extension. J. Biol. Chem. 280: 4238-4246. [Abstract/Free Full Text]
  42. Kanters, D., W. ten Hove, B. Luijk, C. van Aalst, R. C. Schweizer, J. W. Lammers, H. G. Leufkens, J. A. Raaijmakers, M. Bracke, L. Koenderman. 2007. Expression of activated Fc{gamma}RII discriminates between multiple granulocyte-priming phenotypes in peripheral blood of allergic asthmatic subjects. J. Allergy Clin. Immunol. 120: 1073-1081.
  43. Georas, S. N., M. C. Liu, W. Newman, L. D. Beall, B. A. Stealey, B. S. Bochner. 1992. Altered adhesion molecule expression and endothelial cell activation accompany the recruitment of human granulocytes to the lung after segmental antigen challenge. Am. J. Respir. Cell Mol. Biol. 7: 261-269. [Medline]
  44. Sedgwick, J. B., W. J. Calhoun, R. F. Vrtis, M. E. Bates, P. K. McAllister, W. W. Busse. 1992. Comparison of airway and blood eosinophil function after in vivo antigen challenge. J. Immunol. 149: 3710-3718. [Abstract]
  45. Kroegel, C., M. C. Liu, W. C. Hubbard, L. M. Lichtenstein, B. S. Bochner. 1994. Blood and bronchoalveolar eosinophils in allergic subjects after segmental antigen challenge: surface phenotype, density heterogeneity, and prostanoid production. J. Allergy Clin. Immunol. 93: 725-734. [Medline]
  46. Mengelers, H. J., T. Maikoe, L. Brinkman, B. Hooibrink, J. W. Lammers, L. Koenderman. 1994. Immunophenotyping of eosinophils recovered from blood and BAL of allergic asthmatics. Am. J. Respir. Crit. Care Med. 149: 345-351. [Abstract]
  47. Aalbers, R., H. F. Kauffman, B. Vrugt, M. Smith, G. H. Koeter, W. Timens, J. G. de Monchy. 1993. Bronchial lavage and bronchoalveolar lavage in allergen-induced single early and dual asthmatic responders. Am. Rev. Respir. Dis. 147: 76-81. [Medline]
  48. Shaver, J. R., J. J. O'Connor, M. Pollice, S. K. Cho, G. C. Kane, J. E. Fish, S. P. Peters. 1995. Pulmonary inflammation after segmental ragweed challenge in allergic asthmatic and nonasthmatic subjects. Am. J. Respir. Crit. Care Med. 152: 1189-1191. [Abstract]
  49. Gauvreau, G. M., R. M. Watson, P. M. O'Byrne. 1999. Kinetics of allergen-induced airway eosinophilic cytokine production and airway inflammation. Am. J. Respir. Crit Care Med. 160: 640-647. [Abstract/Free Full Text]
  50. Wood, L. J., R. Sehmi, S. Dorman, Q. Hamid, M. K. Tulic, R. M. Watson, R. Foley, P. Wasi, J. A. Denburg, G. Gauvreau, P. M. O'Byrne. 2002. Allergen-induced increases in bone marrow T lymphocytes and interleukin-5 expression in subjects with asthma. Am. J. Respir. Crit. Care Med. 166: 883-889. [Abstract/Free Full Text]
  51. Moser, R., J. Fehr, P. L. B. Bruijnzeel. 1992. Il-4 controls the selective endothelium-driven transmigration of eosinophils from allergic individuals. J. Immunol. 149: 1432-1438. [Abstract]
  52. Ebisawa, M., T. Yamada, C. Bickel, D. Klunk, R. P. Schleimer. 1994. Eosinophil transendothelial migration induced by cytokines: III. Effect of the chemokine RANTES. J. Immunol. 153: 2153-2160. [Abstract]
  53. Jia, G. Q., J. A. Gonzalo, A. Hidalgo, D. Wagner, M. Cybulsky, J. C. Gutierrez-Ramos. 1999. Selective eosinophil transendothelial migration triggered by eotaxin via modulation of Mac-1/ICAM-1 and VLA-4/VCAM-1 interactions. Int. Immunol. 11: 1-10. [Abstract/Free Full Text]
  54. Nagata, M., H. Yamamoto, K. Tabe, Y. Sakamoto. 2001. Eosinophil transmigration across VCAM-1-expressing endothelial cells is upregulated by antigen-stimulated mononuclear cells. Int. Arch. Allergy Immunol. 125: (Suppl. 1):7-11. [Medline]
  55. Norris, V., L. Choong, D. Tran, Z. Corden, M. Boyce, H. Arshad, S. Holgate, B. O'Connor, S. Millet, B. Miller, et al 2005. Effect of IVL745, a VLA-4 antagonist, on allergen-induced bronchoconstriction in patients with asthma. J. Allergy Clin. Immunol. 116: 761-767. [Medline]
  56. Yamamoto, H., M. Nagata, Y. Sakamoto. 2005. CC chemokines and transmigration of eosinophils in the presence of vascular cell adhesion molecule 1. Ann. Allergy Asthma Immunol. 94: 292-300. [Medline]
  57. Banerjee, E. R., Y. Jiang, W. R. Henderson, Jr, L. M. Scott, T. Papayannopoulou. 2007. {alpha}4 and β2 integrins have nonredundant roles for asthma development, but for optimal allergen sensitization only {alpha}4 is critical. Exp. Hematol. 35: 605-617. [Medline]
  58. Liu, L. Y., C. A. Swensen, E. A. Kelly, H. Kita, W. W. Busse. 2000. The relationship of sputum eosinophilia and sputum cell generation of IL-5. J. Allergy Clin. Immunol. 106: 1063-1069. [Medline]
  59. Kelly, E. A. B., W. W. Busse, N. N. Jarjour. 2003. A comparison of the airway response to segmental antigen bronchoprovocation in atopic asthma and allergic rhinitis. J. Allergy Clin. Immunol. 111: 79-86. [Medline]
  60. Zhu, X., N. M. Munoz, K. P. Kim, H. Sano, W. Cho, A. R. Leff. 1999. Cytosolic phospholipase A2 activation is essential for β1 and β2 integrin-dependent adhesion of human eosinophils. J. Immunol. 163: 3423-3249. [Abstract/Free Full Text]
  61. Wong, C. K., W. K. Ip, C. W. Lam. 2003. Interleukin-3,–5, and granulocyte macrophage colony-stimulating factor-induced adhesion molecule expression on eosinophils by p38 mitogen-activated protein kinase and nuclear factor-{kappa}B. Am. J. Respir. Cell Mol. Biol. 29: 133-147. [Abstract/Free Full Text]
  62. Rennard, S. I., G. Basset, D. Lecossier, K. M. O'Donnell, P. Pinkston, P. G. Martin, R. G. Crystal. 1986. Estimation of volume of epithelial lining fluid recovered by lavage using urea as marker of dilution. J. Appl. Physiol. 60: 532-538. [Abstract/Free Full Text]
  63. Chihara, J., J. Plumas, V. Gruart, J. Tavernier, L. Prin, A. Capron, M. Capron. 1990. Characterization of a receptor for interleukin 5 on human eosinophils: variable expression and induction by granulocyte/macrophage colony-stimulating factor. J. Exp. Med. 172: 1347-1351. [Abstract/Free Full Text]
  64. Plaetinck, G., J. Van der Heyden, J. Tavernier, I. Fache, T. Tuypens, S. Fischkoff, W. Fiers, R. Devos. 1990. Characterization of interleukin 5 receptors on eosinophilic sublines from human promyelocytic leukemia (HL-60) cells. J. Exp. Med. 172: 683-691. [Abstract/Free Full Text]
  65. Ingley, E., I. G. Young. 1991. Characterization of a receptor for interleukin-5 on human eosinophils and the myeloid leukemia line HL-60. Blood 78: 339-344. [Abstract/Free Full Text]
  66. Migita, M., N. Yamaguchi, S. Mita, S. Higuchi, Y. Hitoshi, Y. Yoshida, M. Tomonaga, I. Matsuda, A. Tominaga, K. Takatsu. 1991. Characterization of the human IL-5 receptors on eosinophils. Cell. Immunol. 133: 484-497. [Medline]
  67. Tavernier, J., R. Devos, S. Cornelis, T. Tuypens, J. Van der Heyden, W. Fiers, G. Plaetinck. 1991. A human high affinity interleukin-5 receptor (IL5R) is composed of an IL5-specific {alpha} chain and a β chain shared with the receptor for GM-CSF. Cell 66: 1175-1184. [Medline]
  68. Zhang, J., P. Wu, R. Kuvelkar, J. L. Schwartz, R. W. Egan, M. M. Billah, P. Wang. 1999. A scintillation proximity assay for human interleukin-5 (hIL-5) high-affinity binding in insect cells coexpressing hIL-5 receptor {alpha} and β subunits. Anal. Biochem. 268: 134-142. [Medline]
  69. Bates, M. E., V. L. Green, P. J. Bertics. 2000. ERK1 and ERK2 activation by chemotactic factors in human eosinophils is interleukin 5-dependent and contributes to leukotriene C4 biosynthesis. J. Biol. Chem. 275: 10968-10975. [Abstract/Free Full Text]
  70. Hall, D. J., J. Cui, M. E. Bates, B. A. Stout, L. Koenderman, P. J. Coffer, P. J. Bertics. 2001. Transduction of a dominant-negative H-Ras into human eosinophils attenuates extracellular signal-regulated kinase activation and interleukin-5-mediated cell viability. Blood 98: 2014-2021. [Abstract/Free Full Text]
  71. Liu, L. Y., J. B. Sedgwick, M. E. Bates, R. F. Vrtis, J. E. Gern, H. Kita, N. N. Jarjour, W. W. Busse, E. A. Kelly. 2002. Decreased expression of membrane IL-5 receptor {alpha} on human eosinophils: II. IL-5 down-modulates its receptor via a proteinase-mediated process. J. Immunol. 169: 6459-6466. [Abstract/Free Full Text]
  72. Bates, M. E., L. Y. Liu, S. Esnault, B. A. Stout, E. Fonkem, V. Kung, J. B. Sedgwick, E. A. Kelly, D. M. Bates, J. S. Malter, et al 2004. Expression of interleukin-5- and granulocyte macrophage-colony-stimulating factor-responsive genes in blood and airway eosinophils. Am. J. Respir. Cell Mol. Biol. 30: 736-743. [Abstract/Free Full Text]
  73. Zangrilli, J. G., J. R. Shaver, R. A. Cirelli, S. K. Cho, C. G. Garlisi, A. Falcone, F. M. Cuss, J. E. Fish, S. P. Peters. 1995. sVCAM-1 levels after segmental antigen challenge correlate with eosinophil influx, IL-4 and IL-5 production, and the late phase response. Am. J. Respir. Crit. Care Med. 151: 1346-1353. [Abstract]
  74. Sur, S., H. Kita, G. J. Gleich, T. C. Chenier, L. W. Hunt. 1996. Eosinophil recruitment is associated with IL-5, but not with RANTES, twenty-four hours after allergen challenge. J. Allergy Clin. Immunol. 97: 1272-1278. [Medline]
  75. Jarjour, N. N., W. J. Calhoun, E. A. Kelly, G. J. Gleich, L. B. Schwartz, W. W. Busse. 1997. The immediate and late allergic response to segmental bronchopulmonary provocation in asthma. Am. J. Respir. Crit. Care Med. 155: 1515-1521. [Abstract]
  76. Sedgwick, J. B., W. J. Calhoun, G. J. Gleich, H. Kita, J. S. Abrams, L. B. Schwartz, B. Volovitz, M. Ben-Yaakov, W. W. Busse. 1991. Immediate and late airway response of allergic rhinitis patients to segmental antigen challenge. Characterization of eosinophil and mast cell mediators. Am. Rev. Resp. Dis. 144: 1274-1281. [Medline]
  77. Berends, C., B. Dijkhuizen, J. G. de Monchy, J. Gerritsen, H. F. Kauffman. 1994. Induction of low density and up-regulation of CD11b expression of neutrophils and eosinophils by dextran sedimentation and centrifugation. J. Immunol. Methods 167: 183-193. [Medline]
  78. Todd, R. F., III, M. A. Arnaout, R. E. Rosin, C. A. Crowley, W. A. Peters, B. M. Babior. 1984. Subcellular localization of the large subunit of Mo1 (Mo1 {alpha}; formerly gp 110), a surface glycoprotein associated with neutrophil adhesion. J. Clin. Invest. 74: 1280-1290. [Medline]
  79. Mathias, J. R., B. J. Perrin, T. X. Liu, J. Kanki, A. T. Look, A. Huttenlocher. 2006. Resolution of inflammation by retrograde chemotaxis of neutrophils in transgenic zebrafish. J. Leukocyte Biol. 80: 1281-1288. [Abstract/Free Full Text]
  80. Buckley, C. D., E. A. Ross, H. M. McGettrick, C. E. Osborne, O. Haworth, C. Schmutz, P. C. Stone, M. Salmon, N. M. Matharu, R. K. Vohra, et al 2006. Identification of a phenotypically and functionally distinct population of long-lived neutrophils in a model of reverse endothelial migration. J. Leukocyte Biol. 79: 303-311. [Abstract/Free Full Text]
  81. Warringa, R. A., H. J. Mengelers, J. A. Raaijmakers, P. L. Bruijnzeel, L. Koenderman. 1993. Upregulation of formyl-peptide and interleukin-8-induced eosinophil chemotaxis in patients with allergic asthma. J. Allergy Clin. Immunol. 91: 1198-1205. [Medline]
  82. Sehmi, R., L. J. Wood, R. Watson, R. Foley, Q. Hamid, P. M. O'Byrne, J. A. Denburg. 1997. Allergen-induced increases in IL-5 receptor {alpha}-subunit expression on bone marrow-derived CD34+ cells from asthmatic subjects: a novel marker of progenitor cell commitment towards eosinophilic differentiation. J. Clin. Invest. 100: 2466-2475. [Medline]
  83. Sehmi, R., S. Dorman, A. Baatjes, R. Watson, R. Foley, S. Ying, D. S. Robinson, A. B. Kay, P. M. O'Byrne, J. A. Denburg. 2003. Allergen-induced fluctuation in CC chemokine receptor 3 expression on bone marrow CD34+ cells from asthmatic subjects: significance for mobilization of haemopoietic progenitor cells in allergic inflammation. Immunology 109: 536-546. [Medline]
  84. Moser, R., J. Fehr, L. Olgiati, P. L. Bruijnzeel. 1992. Migration of primed human eosinophils across cytokine-activated endothelial cell monolayers. Blood 79: 2937-2945. [Abstract/Free Full Text]
  85. Nakajima, H., H. Sano, T. Nishimura, S. Yoshida, I. Iwamoto. 1994. Role of vascular cell adhesion molecule 1/very late activation antigen 4 and intercellular adhesion molecule 1/lymphoycte function-associated antigen 1 interactions in antigen-induced eosinophil and T cell recruitment into the tissue. J. Exp. Med. 179: 1145-1154. [Abstract/Free Full Text]
  86. ten Hacken, N. H., D. S. Postma, F. Bosma, G. Drok, B. Rutgers, J. Kraan, W. Timens. 1998. Vascular adhesion molecules in nocturnal asthma: a possible role for VCAM-1 in ongoing airway wall inflammation. Clin. Exp. Allergy 28: 1518-1525. [Medline]
  87. Masinovsky, B., D. Urdal, W. M. Gallatin. 1990. IL-4 acts synergistically with IL-1 β to promote lymphocyte adhesion to microvascular endothelium by induction of vascular cell adhesion molecule-1. J. Immunol. 145: 2886-2895. [Abstract]
  88. Thornhill, M. H., D. O. Haskard. 1990. IL-4 regulates endothelial cell activation by IL-1, tumor necrosis factor, or IFN-{gamma}. J. Immunol. 145: 865-872. [Abstract]
  89. Thornhill, M. H., S. M. Wellicome, D. L. Mahiouz, J. S. Lanchbury, U. Kyan-Aung, D. O. Haskard. 1991. Tumor necrosis factor combines with IL-4 or IFN-{gamma} to selectively enhance endothelial cell adhesiveness for T cells: the contribution of vascular cell adhesion molecule-1-dependent and -independent binding mechanisms. J. Immunol. 146: 592-598. [Abstract]
  90. Weller, P. F., T. H. Rand, S. E. Goelz, G. Chi-Rosso, R. R. Lobb. 1991. Human eosinophil adherence to vascular endothelium mediated by binding to vascular cell adhesion molecule 1 and endothelial leukocyte adhesion molecule 1. Proc. Natl. Acad. Sci. USA 88: 7430-7433. [Abstract/Free Full Text]
  91. Schleimer, R. P., S. A. Sterbinsky, J. Kaiser, C. A. Bickel, D. A. Klunk, K. Tomioka, W. Newman, F. W. Luscinskas, M. A. Gimbrone, Jr, B. W. McIntyre, B. S. Bochner. 1992. IL-4 induces adherence of human eosinophils and basophils but not neutrophils to endothelium: association with expression of VCAM-1. J. Immunol. 148: 1086-1092. [Abstract]
  92. Barthel, S. R., D. S. Annis, D. F. Mosher, M. W. Johansson. 2006. Differential engagement of modules 1 and 4 of vascular cell adhesion molecule-1 (CD106) by integrins {alpha}4β1 (CD49d/29) and {alpha}Mβ2 (CD11b/18) of eosinophils. J. Biol. Chem. 281: 32175-32187. [Abstract/Free Full Text]
  93. Ebisawa, M., B. S. Bochner, S. N. Georas, R. P. Schleimer. 1992. Eosinophil transendothelial migration induced by cytokines: I. Role of endothelial and eosinophil adhesion molecules in IL-1 β-induced transendothelial migration. J. Immunol. 149: 4021-4028. [Abstract]
  94. Liu, L., L. Hakansson, P. Ridefelt, R. C. Garcia, P. Venge. 2003. Priming of eosinophil migration across lung epithelial cell monolayers and upregulation of CD11b/CD18 are elicited by extracellular Ca2+. Am. J. Respir. Cell Mol. Biol. 28: 713-721. [Abstract/Free Full Text]
  95. Ulfman, L. H., J. Alblas, C. W. van Aalst, J. J. Zwaginga, L. Koenderman. 2005. Differences in potency of CXC chemokine ligand 8-, CC chemokine ligand 11-, and C5a-induced modulation of integrin function on human eosinophils. J. Immunol. 175: 6092-6099. [Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 Am. J. Respir. Cell Mol. Bio.Home page
M. C. Seeds, K. K. Peachman, D. L. Bowton, K. L. Sivertson, and F. H. Chilton
Regulation of Arachidonate Remodeling Enzymes Impacts Eosinophil Survival during Allergic Asthma
Am. J. Respir. Cell Mol. Biol., September 1, 2009; 41(3): 358 - 366.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Johansson, M. W.
Right arrow Articles by Mosher, D. F.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Johansson, M. W.
Right arrow Articles by Mosher, D. F.
Right arrowPubmed/NCBI databases
*Compound via MeSH
*Substance via MeSH
Medline Plus Health Information
*Asthma

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS