1 Department of Physiology, Brody School of Medicine at East Carolina University, Greenville, North Carolina 27858; and 2 Department of Biochemisty and Molecular Biology, Mayo Clinic, Scottsdale, Arizona 85259
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ABSTRACT |
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Allergen-induced recruitment of T
lymphocytes and eosinophils to the airways is associated with increased
expression of the transcription factor GATA-3. In this study, the
relationship between airway inflammation and GATA-3 expression in the
lungs was investigated using ragweed-sensitized C57BL/6J mice.
Intratracheal ragweed challenge increased both the number of
GATA-3-expressing cells in the perivascular and peribronchial regions
and the amount of expression per cell. Interleukin (IL)-4 and IL-5
levels in bronchoalveolar lavage fluid were upregulated in parallel
with GATA-3 expression. GATA-3 mRNA and protein colocalized to
eosinophils. Eosinophils isolated from the lungs and stimulated with
phorbol 12-myristate 13-acetate and/or A-23187 released IL-5. The
release was inhibited by actinomycin D, which indicates that de novo
synthesis of the cytokine was involved. Western blot analysis of
proteins from isolated eosinophils demonstrated expression of the p50
subunit of nuclear factor-B, a transcription factor that is
implicated in control of GATA-3 expression. These data provide evidence
that allergen challenge increases GATA-3 and proinflammatory cytokine expression by pulmonary eosinophils, which could provide positive feedback for the inflammatory response.
asthma; airway inflammation; major basic protein; transcription; interleukin
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INTRODUCTION |
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ASTHMA IS CHARACTERIZED by pulmonary inflammation, reversible airflow obstruction, and airway hyperresponsiveness (1, 2, 7, 10). The asthma phenotype evolves from a complex cascade of immunological responses to aeroallergens, which leads to leukocyte recruitment to the airways and subsequent remodeling of the inflamed tissue (11, 14, 18). The recruitment of eosinophils is associated with increased production of the Th2 cytokines [interleukin (IL)-4 and IL-5], and ectopic expression within the airways of these two factors has been shown to induce eosinophil recruitment (21, 31). The biological role of IL-5 is limited to control of eosinophil proliferation, differentiation, activation, and survival (6, 9, 23, 34, 37, 43). In contrast, the biological significance of IL-4 is broader and includes stimulation of CD4+ T-cell differentiation and proliferation (40), B-cell class switching (25), and the induction of eosinophil-specific chemokine secretion (38). Thus IL-4 and IL-5 are key factors in asthma pathogenesis.
Studies suggest that CD4+ T cells are the primary source of IL-4 and IL-5 in allergic inflammation (1, 15, 17, 32). However, there is evidence that eosinophils are capable of providing positive feedback to the inflammatory response via production and secretion of these cytokines (16). In murine lungs infected with Shistosoma mansoni, eosinophils are reported to be the dominant source of IL-4 and IL-5 and to cause prolonged eosinophil recruitment and continued granuloma formation (33). To date, a similar role for eosinophils in the development of allergic asthma has not been established.
Transcription of both the IL-4 and IL-5 genes is regulated in part by the helix-loop-helix transcription factor GATA-3 (3, 30, 46). In murine T lymphocytes, GATA-3 is a major positive regulatory factor for IL-4 production (46, 47), and GATA-3 is required for IL-5 expression in T cells (45, 46). Ouyang and colleagues (27) recently established that GATA-3 is part of a master switch for Th2 commitment in T cells. This response to GATA-3 is independent of STAT-6, which indicates that the effect is not mediated by the IL-4/STAT-6 signaling pathway.
Evidence supporting a role for GATA-3 in the development of asthma is accumulating. Biopsies from human asthmatic patients have demonstrated a significant increase in the number of GATA-3 mRNA-positive cells in the airways compared with tissue from nonasthmatic individuals (26). Using a murine model of asthma, Zhang and colleagues (45) demonstrated that T-cell-specific expression of a GATA-3 dominant-negative mutant during the sensitization and challenge processes attenuated allergen-induced airway eosinophilia. These observations together with studies correlating eosinophilic inflammation with Th2 cytokine production (24) suggest that GATA-3 plays a key role in eosinophilic inflammation of the airways.
In the current study, ragweed (RW)-sensitized C57BL/6J mice were used to define the pattern of GATA-3 expression in the lungs over a 72-h period after allergen challenge. Maximum expression of GATA-3 was observed 12-24 h after RW challenge and corresponded with the appearance of IL-4 and IL-5 in bronchoalveolar lavage (BAL) fluid. Greater than 95% of the GATA-3 mRNA expressing cells in the lungs were eosinophils. Furthermore, in vitro stimulation of RW-primed lung eosinophils demonstrated transcriptional dependence for Th2 cytokine production.
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MATERIALS AND METHODS |
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Animals. Male 6-10-wk-old C57BL/6J mice were purchased from Jackson Laboratories (Bar Harbor, ME). NJ.1638 mice were obtained from J. Lee and N. Lee (Mayo Clinic, Scottsdale, AZ). All mice were maintained by the Department of Comparative Medicine at the Brody School of Medicine at East Carolina University. Animals were fed Purina laboratory chow and sterilized tap water ad libitum. All studies were approved by the Animal Welfare and Use Committee at the Brody School of Medicine.
Sensitization and challenge of animals. A 14-day protocol was used. Mice were sensitized by intraperitoneal injection of RW on days 0 and 4 (80 µg/injection, lot 56-29; endotoxin content <2.3 ng/mg RW; Greer Laboratories, Lenoir, NC). Sensitization solution consisted of 1 mg of RW in 1 ml of 0.9% NaCl (Baxter, Deerfield, IL) plus 333 µl of Imject alum (Pierce, Rockford, IL). On day 11, animals were anesthetized with ketamine (90 mg/kg body wt) and xylazine (10 mg/kg body wt) and challenged by intratracheal administration of RW (10 µg of RW in 0.1 ml of 0.9% NaCl).
Lung lavage, tissue fixation, and sectioning.
Lungs were lavaged with a single 1.0-ml aliquot of PBS (GIBCO-BRL,
Grand Island, NY). Samples were centrifuged at 2,000 rpm for 5 min, and
the supernatants were removed and stored at 80°C. Lungs were
removed, infused with 4% paraformaldehyde (in PBS) for 30 min, rinsed
with PBS, and immersed in 0.5 M sucrose (in PBS) overnight at 4°C.
Lungs were inflated with and embedded in optimal cutting temperature
compound (Sakura FineTek, Torrance, CA) and stored at
80°C.
Cytokine determination. Concentrations of IL-4 and IL-5 were determined using two-site immunoenzymetric assay kits (Endogen, Cambridge, MA) according to the manufacturer's instructions. The lower limits of detection were 1 pg/ml for both IL-4 and IL-5.
In situ hybridization.
In situ hybridization was performed as described by Helton and
colleagues (13). Lung sections (10-20 µm) were
placed on Superfrost Plus microscope slides (VWR Scientific, West
Chester, PA), acetylated using 0.25% acetic acid anhydride, and
dehydrated in ethanol. Sections were hybridized using synthetic DNA
oligonucleotides with the following sequences: GATA-3 antisense
(5'-CTCGGCTGTGCTCGCGCCT-3') and GATA-3 sense
(5'-AGGGCGCGGAGCACAGCCGAG-3'). Oligonucleotides were synthesized by
GIBCO-BRL and labeled on the 3' end using -35S-labeled
dATP (1,075 Ci/mmol, New England Nuclear, Boston, MA) and terminal
deoxynucleotidyl transferase (Boehringer Manheim, Indianapolis, IN) to
a specific activity of ~0.9-1.8 µCi/mmol. Sections were
incubated with 1 × 106 counts/min of
-35S-labeled dATP per 25 µl of hybridization buffer
for 12 h at 37°C. Slides were dipped in Kodak NTB-3
emulsion (Sigma Chemical, St. Louis, MO), exposed for 72 h, and
processed using Dektol and Kodak fixer (Sigma). Quantification of the
mRNA hybridization signal was performed using NIH Image 1.60 software
(W. Rasband, National Institute of Mental Health) and a Macintosh G3
computer. Integrated density for GATA-3 mRNA expression in the lung
sections was determined using the NIH imager program. The area
of the cell being scanned and the mean density of the scanned region
were multiplied to yield the integrated density. Numbers of GATA-3
mRNA-expressing cells were normalized per millimeter of basement
membrane for airways and blood vessels. Enumeration of GATA-3 mRNA
values for the alveolar region were normalized per square millimeter of
distal lung area.
Immunofluorescence and immunocytochemistry. Frozen lung sections were analyzed for eosinophil major basic protein (MBP) and GATA-3 protein expression using anti-mMBP monoclonal antibody and anti-GATA-3 polyclonal antibody (Santa Cruz Biotech, San Diego, CA). Briefly, endogenous peroxidase activity was quenched with 3% H2O2 (Sigma) and 80% methanol (J. T. Baxter, Phillipsburg, NJ) in PBS. Lung sections were permeabilized with pepsin (Zymed Laboratories, San Francisco, CA) and 0.01% Triton X (Sigma). Sections were blocked with 10% mouse serum (Vector Laboratories, Burlingame, CA) and incubated with anti-GATA-3 Ab (1:50 dilution; 200 µg/ml) that was biotinylated with the DAKO-ARK biotinylation kit (DAKO, Carpinteria, CA). GATA-3 antibody staining was visualized by the streptavidin-diaminobenzidine system (DAKO) as described by the manufacturer. Subsequently, cellular autofluorescence was blocked by incubation of sections with 1% Chromotrope-R (Sigma), and slides were incubated in rat anti-mMBP Ab (1:3,000 dilution; 1.4 µg/ml) before the addition of Alexa-488 conjugated goat anti-rat IgG antibody (1:5,000 dilution; 0.5 µg/ml; Molecular Probes, Eugene, OR).
Isolation of peripheral blood and airway eosinophils. Isolation of peripheral blood eosinophils was performed by collecting 0.5 ml of peripheral blood from the tail vein of NJ.1638 (T-cell-specific and IL-5 transgenic) mice (22). White blood cells were isolated by centrifugation on a discontinuous Percoll gradient with subsequent hypotonic lysis of remaining red blood cells. Lung eosinophils were isolated from RW-challenged C57BL/6J mice. The lungs were perfused with ice-cold PBS introduced through the right ventricle and were excised, minced, digested in collagenase (1% collagenase in PBS; Sigma), and filtered through a 100-µm mesh. White blood cells were isolated on a discontinuous Percoll gradient. Eosinophils were enriched from both populations by the negative-selection process using anti-CD90 and anti-CD45R antibodies to deplete the B- and T-cell populations using the MACS magnetic bead separation method per the manufacturer's suggested protocol (Miltenyi Biotechnical, Auburn, CA). Eosinophil fractions were routinely enriched to <98%.
In vitro stimulation of eosinophils.
Purified peripheral blood and lung eosinophils were resuspended in
RPMI-1640 (GIBCO-BRL) and 5% fetal calf serum (GIBCO-BRL) at a cell
density of 1 × 106 cells/ml for both peripheral blood
and lung eosinophils. The cells were stimulated with 107
M phorbol 12-myristate 13-acetate (PMA) and 10
7 M A-23187
(Sigma). Actinomycin D was added at a concentration of
10
6 M when indicated. Samples were stimulated in 96-well
plates at 37°C for 30 min, 1 h, and 16 h.
Western blot analysis of GATA-3 and nuclear factor-B.
Purified peripheral blood eosinophils from NJ.1638 mice were collected
by centrifugation and resuspended in extraction buffer that contained
10 mM Tris · HCl, pH 7.4, 0.32 M sucrose, 2 mM EDTA, 1 mM EGTA,
0.5 mM dithiothreitol, 0.5 mM phenylmethylsulfonyl fluoride, and 2 µg/ml pepstatin A. The cells were disrupted by sonication on ice
twice for 20 s. Lysates were centrifuged for 1 h at 100,000 g. The supernatants were collected and protein concentrations were determined (Bio-Rad). Proteins were separated by
12% SDS-PAGE and transferred to 0.45-µm polyvinylidene difluoride membranes (Immobilon). GATA-3 and the p50 subunit of nuclear factor (NF)-
B were identified by immunoblotting with goat anti-GATA-3 Ab
and goat anti-NF-
B p50 Ab (both from Santa Cruz Biotechnology), respectively, followed by horseradish peroxidase-conjugated donkey anti-goat IgG Ab. Immunoreactive protein complexes were detected by
using enhanced chemiluminescence detection reagents.
Statistical analysis. All experimental values within the individual data sets were compared by ANOVA and Tukey's HSD post hoc test (P < 0.05). Values presented in this report represent means ± SE.
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RESULTS |
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GATA-3 mRNA-positive cells are recruited to the airways after RW
allergen challenge.
In situ hybridization revealed that GATA-3 mRNA expression increased
over the 24 h after RW challenge. The GATA-3-positive cells were
located along the basement membrane of the airways and blood vessels
and were broadly dispersed in the alveolar region (Fig.
1). A significant increase in the number
of GATA-3-positive cells was observed in the airways and blood vessels
12 h after RW challenge (Table 1).
In contrast, an increase in expression in the alveolar region was
delayed. In sensitized mice, a significant increase in the number of
GATA-3 mRNA-positive cells was not observed until 24 h after RW
challenge. Interestingly, recruitment to the airways and blood vessels
was only observed after the airway challenge, whereas sensitization
alone elicited expression of GATA-3 in the alveolar region. Kinetics of
the GATA-3 mRNA-positive cell distribution reflected the pattern of
eosinophil recruitment beginning with extravasation through the blood
vessels at 12 h, localization around vessels and large airways by
24 h, and a minor increase in alveolar eosinophilia at these time
points.
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Expression of GATA-3 mRNA and GATA-3 protein colocalizes with
MBP-positive cells.
Combined in situ hybridization for GATA-3 mRNA and immunofluorescence
for MBP protein demonstrated that GATA-3 mRNA colocalized with MBP in
the airways (Fig. 2). Of the GATA-3
mRNA-positive cells, >95% costained for MBP. Colocalization was also
observed at the protein level (Fig. 3).
These results indicate that the majority of cells expressing GATA-3 in
the airways after allergen challenge are eosinophils. To confirm this
finding, peripheral blood eosinophils from nonsensitized and
nonchallenged NJ.1638 mice were isolated, and Western blot analysis was
performed (Fig. 4). Expression of GATA-3
and the p50 subunit of NF-B, the upstream modulator of GATA-3, were
detected.
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IL-4 and IL-5 levels in BAL fluid increase after RW challenge.
IL-4 and IL-5 concentrations in the BAL fluid of RW-challenged mice
were increased significantly 24 h postchallenge (Table 2). Comparison of the IL-4 and IL-5
levels with the expression of GATA-3 (shown in Fig. 5) indicates that
the three parameters changed in parallel, and that eosinophils were a
proble source of IL-4 and IL-5 in the lungs after allergen challenge.
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Nonspecific activation of airway and peripheral blood eosinophils
in vitro leads to de novo production of IL-4 and IL-5.
The ability of eosinophils to produce the Th2 cytokines IL-4 and IL-5
was assessed in vitro using cells isolated from peripheral blood of
NJ.1638 mice and the lungs of RW-challenged wild-type mice. Incubation
with PMA and A-23187 for 18 h induced the release of both IL-4 (Fig.
6A) and IL-5 (Fig.
6B). Release of preformed IL-4 and IL-5 was unlikely,
because exposure of the cells to PMA or A-23187 for 30-120 min did
not lead to the release of these cytokines (data not shown). In
addition, incubation of the cells with the RNA polymerase II inhibitor
actinomycin D abolished IL-4 and IL-5 release, which suggests that
cytokine production is a function of transcriptional activation of
these Th2 cytokine genes.
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DISCUSSION |
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The results of this study provide evidence that allergen challenge induces expression of GATA-3 and GATA-3-responsive genes in pulmonary eosinophils. A role for GATA-3 in the development of asthma was initially demonstrated by Nakamura and colleagues (26), who reported a significant elevation of GATA-3 mRNA in lung tissue from asthma patients compared with lung tissue from nonasthmatic subjects. Further evidence of a role for GATA-3 in allergen-induced lung inflammation was provided by Zhang and colleagues (45), who found a dominant negative mutant expressed for GATA-3 in mice and demonstrated that the pulmonary response to allergen challenge was attenuated. Expression of the GATA-3 mutant was restricted to T cells and was induced during both the sensitization and airway-challenge periods. The results of these studies were therefore consistent with the known importance of Th2 lymphocytes in asthma and the role of GATA-3 in the growth of this cell type. The results of the present study indicate that GATA-3 plays a broader role in allergic inflammation. Expression by pulmonary eosinophils and the capacity of that cell type to produce IL-5 provide a mechanism for positive feedback in the inflammatory response at the site of the allergen challenge.
There is considerable evidence that GATA-3 regulates IL-5 gene expression in human and murine T cells. For example, expression of IL-5 in EL-4 and myeloid cells requires the presence of GATA-3 (3, 36, 44), mutation of the GATA-3 binding site in EL-4 cells abolishes antigen or cAMP-induced IL-5 production (44), GATA-3 binds to the IL-5C promoter site in EL-4 cells (20), and retroviral infection with GATA-3 cDNA enhances IL-5 production in developing Th1 T-cell clones (8). The role of GATA-3 in the regulation of IL-4 expression is more controversial. In contrast to IL-5, early studies of IL-4 expression demonstrated that ectopic expression of GATA-3 did not induce IL-4 production (46), and GATA-3 antisense oligonucleotides did not inhibit IL-4 promoter activity (30). However, recent studies suggest that GATA-3 plays a more direct role in IL-4 transcription. Retroviral expression of GATA-3 in committed Th1 cells enhances IL-4 production (8), and expression of a dominant negative mutant for GATA-3 inhibits allergen-induced IL-4 production (45). In addition, introduction of GATA-3 into STAT-6-deficient T cells induces DNase I hypersensitive sites in the IL-4 locus and triggers STAT-6 independent IL-4 production, which is a novel concept in the paradigm of IL-4 gene regulation (27). Taken together, the evidence indicates that GATA-3 is an important factor in the regulation of both IL-4 and IL-5.
The molecular mechanism by which GATA-3 enables the polarization of
T-cell subsets into the Th2 category has also been described by Ouyang
and colleagues (28), who observed that developing Th1
cells could inhibit GATA-3 expression through an IL-12-dependent mechanism. Conversely, GATA-3 expression was found to inhibit IL-12
signaling in T cells. These findings have been further substantiated by
the demonstration that ectopic expression of GATA-3 in developing Th1
cells inhibits interferon- production and commitment to the Th1
phenotype (8). Collectively, these findings indicate that the role of GATA-3 in both the upregulation and downregulation of gene
expression is critical to the development of a Th2 immune response.
The present study demonstrates that murine tissue eosinophils express
GATA-3 after allergic sensitization and challenge. A previous report by
Zon and colleagues (48) demonstrated GATA-3 expression in
the HL-60-C15 and 3+C5 sublines of the human eosinophil-like cell
line, HL-60. GATA-3 has also been shown to downregulate the expression
of genes in HL-60 cells (35). The finding that eosinophils express GATA-3 in vivo is therefore a logical outcome of the present study.
The observation that GATA-3 is expressed and is functionally active in eosinophils has important implications in the progression and self-perpetuation of allergic reactions that lead to asthma. Production of IL-5 by tissue eosinophils should contribute to a microenvironment that supports the localization, longevity, activation, and proliferation of these cells, perhaps independent of Th2 cells. However, its importance in human asthma is unknown. The acute sensitization and challenge protocol used in this study results in a high degree of eosinophil accumulation in the airways with minimal recruitment of lymphocytes. Therefore, the role that eosinophil GATA-3 expression plays in allergic airway inflammation is likely to be amplified in this model. In contrast, chronic allergic airway inflammation such as is observed in human asthma is characterized by significant accumulation of lymphocytes in the airways. Nakamura and colleagues (26) observed that 60-90% of the GATA-3-positive cells in human lung tissue were CD3-positive T cells; however, <15% of the GATA-3-positive cells were identified as eosinophils. Thus in human asthma, eosinophil expression of GATA-3-responsive genes is not likely to be the primary source of proinflammatory cytokines leading to airway inflammation but rather may provide a redundant source of Th2 cytokine to support the chronic inflammatory process.
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FOOTNOTES |
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Address for reprint requests and other correspondence: M. R. Van Scott, Dept. of Physiology, Brody School of Medicine at East Carolina Univ., 6N98 Brody Bldg., Greenville, NC 27858 (E-mail: vanscottmi{at}mail.ecu.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
10.1152/ajplung.00158.2001
Received 7 May 2001; accepted in final form 1 October 2001.
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REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Bentley, AM,
Menz G,
Storz C,
Robinson DS,
Bradley B,
Jeffery PK,
Durham SR,
and
Kay AB.
Identification of T lymphocytes, macrophages, and activated eosinophils in the bronchial mucosa in intrinsic asthma. Relationship to symptoms and bronchial responsiveness.
Am Rev Respir Dis
146:
500-506,
1992[ISI][Medline].
2.
Bjornsdottir, US,
and
Cypcar DM.
Asthma: an inflammatory mediator soup.
Allergy
54 Suppl49:
55-61,
1999[ISI][Medline].
3.
Blumenthal, SG,
Aichele G,
Wirth T,
Czernilofsky AP,
Nordheim A,
and
Dittmer J.
Regulation of the human interleukin-5 promoter by Ets transcription factors. Ets1 and Ets2, but not Elf-1, cooperate with GATA3 and HTLV-I Tax1.
J Biol Chem
274:
12910-12916,
1999
4.
Boushey, HA,
and
Fahy JV.
Targeting cytokines in asthma therapy: round one.
Lancet
356:
2114-2116,
2000[ISI][Medline].
5.
Bryan, SA,
O'Connor BJ,
Matti S,
Leckie MJ,
Kanabar V,
Khan J,
Warrington SJ,
Renzetti L,
Rames A,
Bock JA,
Boyce MJ,
Hansel TT,
Holgate ST,
and
Barnes PJ.
Effects of recombinant human interleukin-12 on eosinophils, airway hyper-responsiveness, and the late asthmatic response.
Lancet
356:
2149-2153,
2000[ISI][Medline].
6.
Coffman, RL,
Seymour BW,
Hudak S,
Jackson J,
and
Rennick D.
Antibody to interleukin-5 inhibits helminth-induced eosinophilia in mice.
Science
245:
308-310,
1989[ISI][Medline].
7.
Eum, SY,
Haile S,
Lefort J,
Huerre M,
and
Vargaftig BB.
Eosinophil recruitment into the respiratory epithelium following antigenic challenge in hyper-IgE mice is accompanied by interleukin 5-dependent bronchial hyperresponsiveness.
Proc Natl Acad Sci USA
92:
12290-12294,
1995[Abstract].
8.
Ferber, IA,
Lee HJ,
Zonin F,
Heath V,
Mui A,
Arai N,
and
O'Garra A.
GATA-3 significantly downregulates IFN- production from developing Th1 cells in addition to inducing IL-4 and IL-5 levels.
Clin Immunol Immunopathol
91:
134-144,
1999.
9.
Finkelman, FD,
Katona IM,
Urban JFJ,
Holmes J,
Ohara J,
Tung AS,
Sample JV,
and
Paul WE.
IL-4 is required to generate and sustain in vivo IgE responses.
J Immunol
141:
2335-2341,
1988
10.
Frigas, E,
Loegering DA,
Solley GO,
Farrow GM,
and
Gleich GJ.
Elevated levels of the eosinophil granule major basic protein in the sputum of patients with bronchial asthma.
Mayo Clin Proc
56:
345-353,
1981[ISI][Medline].
11.
Fujita-Yamaguchi, Y,
Sacks DB,
Mcdonald JM,
Sahal D,
and
Kathuria S.
Effect of basic polycations and proteins on purified insulin receptor.
Biochem J
263:
813-822,
1989[ISI][Medline].
12.
Grimaldi, JC,
Yu NX,
Grunig G,
Seymour BW,
Cottrez F,
Robinson DS,
Hosken N,
Ferlin WG,
Wu X,
Soto H,
O'Garra A,
Howard MC,
and
Coffman RL.
Depletion of eosinophils in mice through the use of antibodies specific for C-C chemokine receptor 3 (CCR3).
J Leukoc Biol
65:
846-853,
1999[Abstract].
13.
Helton, TE,
Daunais JB,
and
McGinty JF.
Convulsant doses of cocaine alter immediate early gene and opioid peptide expression in rat limbic forebrain.
Brain Res Mol Brain Res
20:
285-288,
1993[ISI][Medline].
14.
Hogan, SP,
and
Foster PS.
Cytokines as targets for the inhibition of eosinophilic inflammation.
Pharmacol Ther
74:
259-283,
1997[ISI][Medline].
15.
Kapsenberg, ML,
Hilkens CM,
Jansen HM,
Bos JD,
Snijders A,
and
Wierenga EA.
Production and modulation of T-cell cytokines in atopic allergy.
Int Arch Allergy Immunol
110:
107-113,
1996[ISI][Medline].
16.
Kay, AB,
Barata L,
Meng Q,
Durham SR,
and
Ying S.
Eosinophils and eosinophil-associated cytokines in allergic inflammation.
Int Arch Allergy Immunol
113:
196-199,
1997[ISI][Medline].
17.
Krakauer, T.
IL-10 inhibits the adhesion of leukocytic cells to IL-1-activated human endothelial cells.
Immunol Lett
45:
61-65,
1995[ISI][Medline].
18.
Kung, TT,
Jones H,
Adams GK,
Umland SP,
Kreutner W,
Egan RW,
Chapman RW,
and
Watnick AS.
Characterization of a murine model of allergic pulmonary inflammation.
Int Arch Allergy Immunol
105:
83-90,
1994[ISI][Medline].
19.
Leckie, MJ,
ten Brinke A,
Khan J,
Diamant Z,
O'Connor BJ,
Walls CM,
Mathur AK,
Cowley HC,
Chung KF,
Djukanovic R,
Hansel TT,
Holgate ST,
Sterk PJ,
and
Barnes PJ.
Effects of an interleukin-5 blocking monoclonal antibody on eosinophils, airway hyper-responsiveness, and the late asthmatic response.
Lancet
356:
2144-2148,
2000[ISI][Medline].
20.
Lee, HJ,
O'Garra A,
Arai K,
and
Arai N.
Characterization of cis-regulatory elements and nuclear factors conferring Th2-specific expression of the IL-5 gene: a role for a GATA-binding protein.
J Immunol
160:
2343-2352,
1998
21.
Lee, JJ,
McGarry MP,
Farmer SC,
Denzler KL,
Larson KA,
Carrigan PE,
Brenneise IE,
Horton MA,
Haczku A,
Gelfand EW,
Leikauf GD,
and
Lee NA.
Interleukin-5 expression in the lung epithelium of transgenic mice leads to pulmonary changes pathognomonic of asthma.
J Exp Med
185:
2143-2156,
1997
22.
Lee, NA,
McGarry MP,
Larson KA,
Horton MA,
Kristensen AB,
and
Lee JJ.
Expression of IL-5 in thymocytes/T cells leads to the development of a massive eosinophilia, extramedullary eosinophilopoiesis, and unique histopathologies.
J Immunol
158:
1332-1344,
1997[Abstract].
23.
Lopez, AF,
Sanderson CJ,
Gamble JR,
Campbell HD,
Young IG,
and
Vadas MA.
Recombinant human interleukin 5 is a selective activator of human eosinophil function.
J Exp Med
167:
219-224,
1988[Abstract].
24.
Louis, R,
Lau LC,
Bron AO,
Roldaan AC,
Radermecker M,
and
Djukanovic R.
The relationship between airway inflammation and asthma severity.
Am J Respir Crit Care Med
161:
9-16,
2000
25.
Lutzker, S,
Rothman P,
Pollock R,
Coffman R,
and
Alt FW.
Mitogen- and IL-4-regulated expression of germ-line Ig--2b transcripts: evidence for directed heavy chain class switching.
Cell
53:
177-184,
1988[ISI][Medline].
26.
Nakamura, Y,
Ghaffar O,
Olivenstein R,
Taha RA,
Soussi-Gounni A,
Zhang DH,
Ray A,
and
Hamid Q.
Gene expression of the GATA-3 transcription factor is increased in atopic asthma.
J Allergy Clin Immunol
103:
215-222,
1999[ISI][Medline].
27.
Ouyang, W,
Lohning M,
Gao Z,
Assenmacher M,
Ranganath S,
Radbruch A,
and
Murphy KM.
STAT6-independent GATA-3 autoactivation directs IL-4-independent Th2 development and commitment.
Immunity
12:
27-37,
2000[ISI][Medline].
28.
Ouyang, W,
Ranganath SH,
Weindel K,
Bhattacharya D,
Murphy TL,
Sha WC,
and
Murphy KM.
Inhibition of Th1 development mediated by GATA-3 through an IL-4-independent mechanism.
Immunity
9:
745-755,
1998[ISI][Medline].
29.
Persson, CG,
Erjefalt JS,
Korsgren M,
and
Sundler F.
The mouse trap.
Trends Pharmacol Sci
18:
465-467,
1997[ISI][Medline].
30.
Ranganath, S,
Ouyang W,
Bhattarcharya D,
Sha WC,
Grupe A,
Peltz G,
and
Murphy KM.
GATA-3-dependent enhancer activity in IL-4 gene regulation.
J Immunol
161:
3822-3826,
1998
31.
Rankin, JA,
Picarella DE,
Geba GP,
Temann UA,
Prasad B,
DiCosmo B,
Tarallo A,
Stripp B,
Whitsett J,
and
Flavell RA.
Phenotypic and physiologic characterization of transgenic mice expressing interleukin 4 in the lung: lymphocytic and eosinophilic inflammation without airway hyperreactivity.
Proc Natl Acad Sci USA
93:
7821-7825,
1996
32.
Robinson, DS,
Durham SR,
and
Kay AB.
Cytokines. 3. Cytokines in asthma.
Thorax
48:
845-853,
1993[ISI][Medline].
33.
Rosenwasser, LJ.
Promoter polymorphism in the candidate genes, IL-4, IL-9, TGF-1, for atopy and asthma.
Int Arch Allergy Immunol
118:
268-270,
1999[ISI][Medline].
34.
Rothenberg, ME,
Luster AD,
and
Leder P.
Murine eotaxin: an eosinophil chemoattractant inducible in endothelial cells and in interleukin 4-induced tumor suppression.
Proc Natl Acad Sci USA
92:
8960-8964,
1995[Abstract].
35.
Sadat, MA,
Kumatori A,
Suzuki S,
Yamaguchi Y,
Tsuji Y,
and
Nakamura M.
GATA-3 represses gp91phox gene expression in eosinophil-committed HL-60-C15 cells.
FEBS Lett
436:
390-394,
1998[ISI][Medline].
36.
Siegel, MD,
Zhang DH,
Ray P,
and
Ray A.
Activation of the interleukin-5 promoter by cAMP in murine EL-4 cells requires the GATA-3 and CLEO elements.
J Biol Chem
270:
24548-24555,
1995
37.
Snapper, CM,
and
Paul WE.
Interferon- and B cell stimulatory factor-1 reciprocally regulate Ig isotype production.
Science
236:
944-947,
1987[ISI][Medline].
38.
Stellato, C,
Matsukura S,
Fal A,
White J,
Beck LA,
Proud D,
and
Schleimer RP.
Differential regulation of epithelial-derived C-C chemokine expression by IL-4 and the glucocorticoid budesonide.
J Immunol
163:
5624-5632,
1999
39.
Stelts, D,
Egan RW,
Falcone A,
Garlisi CG,
Gleich GJ,
Kreutner W,
Kung TT,
Nahrebne DK,
Chapman RW,
and
Minnicozzi M.
Eosinophils retain their granule major basic protein in a murine model of allergic pulmonary inflammation.
Am J Respir Cell Mol Biol
18:
463-470,
1998
40.
Swain, SL,
Weinberg AD,
English M,
and
Huston G.
IL-4 directs the development of Th2-like helper effectors.
J Immunol
145:
3796-3806,
1990
41.
Wills-Karp, M.
Immunologic basis of antigen-induced airway hyperresponsiveness.
Annu Rev Immunol
17:
255-281,
1999[ISI][Medline].
42.
Yamaguchi, Y,
Ackerman SJ,
Minegishi N,
Takiguchi M,
Yamamoto M,
and
Suda
Mechanisms of transcription in eosinophils: GATA-1, but not GATA-2, transactivates the promoter of the eosinophil granule major basic protein gene.
Blood
91:
3447-3458,
1998
43.
Yamaguchi, Y,
Hayashi Y,
Sugama Y,
Miura Y,
Kasahara T,
Kitamura S,
Torisu M,
Mita S,
Tominaga A,
and
Takatsu K.
Highly purified murine interleukin 5 (IL-5) stimulates eosinophil function and prolongs in vitro survival. IL-5 as an eosinophil chemotactic factor.
J Exp Med
167:
1737-1742,
1988[Abstract].
44.
Zhang, DH,
Cohn L,
Ray P,
Bottomly K,
and
Ray A.
Transcription factor GATA-3 is differentially expressed in murine Th1 and Th2 cells and controls Th2-specific expression of the interleukin-5 gene.
J Biol Chem
272:
21597-21603,
1997
45.
Zhang, DH,
Yang L,
Cohn L,
Parkyn L,
Homer R,
Ray P,
and
Ray A.
Inhibition of allergic inflammation in a murine model of asthma by expression of a dominant-negative mutant of GATA-3.
Immunity
11:
473-482,
1999[ISI][Medline].
46.
Zhang, DH,
Yang L,
and
Ray A.
Differential responsiveness of the IL-5 and IL-4 genes to transcription factor GATA-3.
J Immunol
161:
3817-3821,
1998
47.
Zheng, W,
and
Flavell RA.
The transcription factor GATA-3 is necessary and sufficient for Th2 cytokine gene expression in CD4 T cells.
Cell
89:
587-596,
1997[ISI][Medline].
48.
Zon, LI,
Yamaguchi Y,
Yee K,
Albee EA,
Kimura A,
Bennett JC,
Orkin SH,
and
Ackerman SJ.
Expression of mRNA for the GATA-binding proteins in human eosinophils and basophils: potential role in gene transcription.
Blood
81:
3234-3241,
1993[Abstract].