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The Joy McCann Culverhouse Airway Disease Research Center, Division of Allergy and Immunology, Department of Internal Medicine, and
Department of Medical Microbiology and Immunology, University of South Florida, and James A. Haley Veterans Affairs Hospital, Tampa, FL 33612; and
Department of Medicine, University of Minnesota, Minneapolis, MN 55415
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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(MIP-1) in the lung tissue. A secondary
RSV infection persistently enhances airway responsiveness in
Df-sensitized mice, with a concomitant increase in
MIP-1 and RSV Ag load in lung tissues. Bulk cultures of thoracic
lymph node mononuclear cells demonstrate that acute RSV infection
augments both Th1- and Th2-like cytokines, whereas secondary and
tertiary infections shift the cytokine profile in favor of the Th2-like
cytokine response in Df-sensitized mice. The elevated
total serum IgE level in the Df-sensitized mice persists
following only RSV reinfection. Thus, recurrent RSV infections in
Df-sensitized mice augment the synthesis of Th2-like
cytokines, total serum IgE Abs, and MIP-1, which are responsible for
persistent airway inflammation and hyperresponsiveness, both of which
are characteristics of asthma. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, which down-regulates a Th2-like response. Therefore, a
RSV-induced Th2 response and the synergy between allergen sensitization
and RSV infection have remained a paradox. The mouse provides an excellent model to investigate the possibility of synergy between allergen sensitization and RSV infection at the immunologic level (14). The inhalation of dust mite allergens in the sensitized mouse induces eosinophil recruitment and the expression of cell adhesion molecules in lung tissue, leads to enhanced inflammation (15). Similarly, mice intranasally (i.n.) inoculated with human RSV develop lung disease characterized by pulmonary inflammation and airway responsiveness. Depending upon the dose of RSV and the age of the mice, they suffer either a mild infection with no overt signs of illness or a moderate/severe infection with weight loss, tachypnea, and patchy pneumonitis (16). The immunity to RSV infection in mice resembles the RSV-induced immunity found in humans (17, 18). RSV-infected mice exhibit increased production of IL-4 and eosinophils in lung tissues and an increase in airway hyperresponsiveness upon exposure to OVA (19), suggesting an interaction between RSV infection and allergen sensitization.
Genetically predisposed infants are exposed naturally to allergens
in utero and may be exposed to allergens and RSV immediately
following birth; we therefore hypothesized that RSV infection may act
as a cofactor in allergy-predisposed infants and expedite the
development of asthma. To test this hypothesis, we established a murine
model of mite allergen sensitization and RSV infection. In this model
mice infected with only live RSV (5 x 105
PFU/mouse) exhibit RSV replication and lung pathology, but show no
overt signs of illness (20). Analysis of cytokine mRNA
expression in RSV-infected mice showed elevated expression of IL-6,
IFN-, and eotaxin mRNAs in the lung; however, IL-10 and IL-13 mRNA
expression increased in spleen cells, suggesting a bias toward Th2-like
cytokine expression (21). Herein, we examined the effect
of repeated RSV infection in allergen-sensitized mice and compared the
airway reactivity, inflammation, and levels of ICAM-1, chemokines,
cytokines, and IgE Abs in groups of control, RSV-infected,
Dermatophagoides farinae (Df) allergen-sensitized, or
Df allergen-sensitized and RSV-infected mice. The results
indicate that repeated RSV infections play a critical role in the
induction of persistently elevated pulmonary inflammation and a
Th2-like response in allergen-sensitized mice.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Female BALB/c mice, 6–8 wk of age, and ICAM-1+/+ and ICAM-1-/- C57BL6 mice were obtained from The Jackson Laboratory (Bar Harbor, ME) and housed in pathogen-free conditions at the James A. Haley Veterans Affairs Hospital Animal Center. The procedures were reviewed and approved by the University of South Florida and the James A. Haley Veterans Affairs Hospital committee on animal research. The A2 strain of human RSV (American Type Culture Collection, Manassas, VA) was propagated in HEp-2 cells (American Type Culture Collection) in a monolayer culture as previously described (22). Mice were infected by an i.n. inoculation of RSV suspension (5 x 105 PFU/50 µl/mouse) under light anesthesia as described previously (20, 21). All experiments were repeated at least twice.
Immunization protocols
Groups of mice were either RSV-infected or sham-infected with
PBS or UV-inactivated RSV on days 4, 28, and 87 (Fig. 1
). Df allergen-sensitized
(AS) and Df allergen-sensitized plus RSV-infected (ASRSV)
groups were immunized one time i.p. on day -13 with 10 µg/mouse of
Df allergens (Greer Laboratories, Lenoir, NC) in 1 mg/mouse
aluminum hydroxide (alum). Then, these mice were challenged i.n., once
daily for 3 consecutive days (days 1–3, 25–27, and 84–86), with 50
µg of Df in 50 µl of PBS/mouse. To facilitate the
pulmonary aspiration of Df, mice were lightly anesthetized
as described above for RSV inoculation. ASRSV mice were inoculated with
RSV 24 h after the last i.n. challenge of Df on days 4,
28, and 87. Airway responsiveness (AR), serum total IgE, and
Df-specific IgE and IgG1 Ab levels were determined in all
groups of mice after 4, 10, and 17 days of primary and secondary RSV
infections and 4 days following the tertiary infection. Lungs, spleens,
and thoracic lymph nodes were removed from the mice 4 days after each
RSV infection to determine RSV Ag loads, cytokines, and chemokines.
Pathology and ICAM-1 immunohistochemistry in the lung tissues were only
examined 4 days following the primary infection.
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The total RSV Ag load in the lung homogenate was measured by an ELISA as previously described (20). Briefly, the supernatants of the lung homogenate were first incubated in goat anti-human RSV Ab (AB1128, Chemicon, Temecula, CA)-coated ELISA plates (Costar, Cambridge, MA) and then incubated with mouse anti-human RSV mAb (NCL-RSV3, Vector Laboratories, Burlingame, CA). Following incubation with peroxidase-labeled goat anti-mouse IgG Ab (Roche, Indianapolis, IN), immune complexes were detected using tetramethyl benzidine as the substrate. ODs were read at 450 nm using an automated ELISA reader. The results were considered positive if the mean OD of duplicate wells was >0.2 after subtraction of the background. The positive results were transformed to PFU per grams of lung by the standard curve obtained with virus of known titers (American Type Culture Collection).
Determination of AR
AR was measured in unrestrained mice by whole body plethysmography (Buxco, Troy, NY). As previously described (19), AR was expressed as an enhanced pause (Penh), a calculated value that closely correlates with pulmonary resistance measured by a conventional, two-chamber plethysmograph in ventilated animals. Using a four-chamber plethysmograph and PBS- and OVA-sensitized mice, similar results were obtained in three independent experiments, suggesting the specificity and low variances of this method. Groups of mice (n = 4) were exposed for 5 min to nebulized PBS and subsequently to increasing concentrations (6–50 mg/ml) of nebulized methacholine (MCh; Sigma, St. Louis, MO) in PBS using the ultrasonic nebulizer. After each nebulization, recordings were taken for 5 min. The Penh values measured during a 5-min sequence were averaged and are expressed for each MCh concentration as a percentage of baseline Penh values obtained following PBS exposure.
Immunohistochemical analysis
Mice were sacrificed with an overdose (0.6 g/kg) of pentobarbital 4 days after the primary RSV inoculation or 5 days after the final Df challenge, and lung sections were subjected to paraffin embedding as previously described (20, 21). Lung inflammation was assessed after staining the section with hematoxylin and eosin (HE) and scoring for severity on a scale of 0–3 indicating the degree of inflammation. The entire lung section was reviewed, and pathological changes were evaluated for epithelial damage, peribronchovascular cell infiltrate, and interstitial-alveolar cell infiltrate for the mononuclear cells and polymorphs. Epithelial damage was scored as: 0 = no damage, 1 = increased cytoplasm of epithelial cells without desquamation, 2 = epithelial desquamation without bronchial exudate composed of inflammatory cells, and 3 = bronchial exudate composed of desquamated epithelial cells and inflammatory cells. Peribronchovascular cell infiltrate was scored as: 0 = no infiltrate, 1 = infiltrate up to four cells, 2 = infiltrate 5–10 cells, and 3 = infiltrate >10 cells. Interstitial-alveolar cell infiltrate was scored as: 0 = no infiltrate, 1 = mild, generalized increase in cell mass of the alveolar septa without thickening of the septa or significant airspace consolidation, 2 = dense septal infiltrate with thickening of septa, and 3 = significant alveolar consolidation in addition to interstitial inflammation. Pathological scores were expressed as the mean ± SEM. The number of eosinophils was determined in 10 perivascular areas/section under oil immersion lens. The slides were coded and scored in blind fashion twice each by three different individuals. Intraobserver variation was <5%.
To examine ICAM-1 expression, slides were immunostained with ICAM-1 Ab. Briefly, paraffin-embedded sections were dewaxed in xylene and rehydrated in three graded concentrations of alcohol. Ag retrieval was performed by incubating sections immersed in 0.01 M sodium citrate buffer (pH 6.0) in a microwave oven (twice, 5 min each time). Endogenous peroxidase was blocked with 0.3% H2O2 in methanol (30 min). The sections were then incubated with a hamster anti-mouse ICAM-1 mAb (clone 01540D, PharMingen, San Diego, CA) at a 1/200 dilution (v/v), and incubated at 37°C for 18 h. Purified, monoclonal, hamster anti-trinitrophenol (PharMingen, San Diego, CA) was substituted for the primary Ab and served as a negative control. Secondary Ab staining and substrate reactions were performed using the Vectastain ABC kit (Vector Laboratories). Diaminobenzidine in H2O2 (Pierce, Rockford, IL) was used as a chromogen. Finally, sections were counterstained with hematoxylin and mounted. All the intermediate washings were performed with PBS buffer. In ICAM-1-stained sections, ICAM-1-positive cells in 200 epithelial cells/section were counted. These examined areas were selected randomly under a low power of magnification (x4) at which leukocyte subtypes and ICAM-1-positive epithelial cells were hardly distinguishable. The mean diameters of the selected blood vessels and bronchioles in each group were not statistically significant. Inter- and intraoperative variations were <10%. The results were expressed as the mean cell numbers of each group. The inflammation in the lungs of ICAM-1+/+ and ICAM-1-/- mice was quantified by scoring system as described above.
Analysis of cytokines and chemokines
Single-cell suspensions were prepared from the spleen (4 x
105/200 µl) and thoracic lymph node (2.5
x 105/200 µl) of BALB/c mice and cultured in
wells coated with either an anti-CD3 Ab (1 mg/ml; clone 17A2,
PharMingen) or Df allergen (100 µg/ml). The
productions of IL-4, IL-5, and IFN- from 24-h (anti-CD3) or 48-h
(Df) culture supernatant were determined by ELISA using
matched Ab pairs (PharMingen) as previously described
(23). To measure chemokine production, lungs were perfused
with ice-cold PBS via the right ventricle, removed aseptically, and
homogenized in 3 ml of lysis buffer (0.5% Triton X-100, 150 mM NaCl,
15 mM Tris, 1 mM CaCl2, and 1 mM
MgCl2, pH 7.4). The homogenates were incubated on
ice for 30 min, then centrifuged at 3000 rpm for 10 min. Supernatants
were collected and passed through a 0.2-µm pore size filter (Gelman
Sciences, Ann Arbor, MI). Concentrations of eotaxin, RANTES, and
MIP-1 in the supernatants were determined using ELISA kits (R&D
Systems, Minneapolis, MN). The results were expressed as the absolute
chemokine amount per wet lung weight (picograms per milligrams of
lung).
Quantification of total serum IgE and Df-specific serum Ig isotype
The total serum IgE level was determined by ELISA using a rat anti-mouse IgE Ab (clone R35-72, PharMingen) and biotin-conjugated rat anti-mouse IgE mAb (clone R35-92, PharMingen) as described previously (21). To measure the Df-specific serum Ig isotype, diluted sera were first incubated in Df extract (5 µg/ml)-coated ELISA plate followed by incubation with biotin-conjugated rat anti-mouse IgE mAb or IgG1 mAb (clone A85-1, PharMingen). The OD was read using an automatic ELISA plate reader at 405 nm. The total serum IgE level was expressed as micrograms per milliliter using mouse IgE standard (PharMingen). The Df-specific serum Ig levels were shown as an OD405 value.
Statistical analysis
Pairs of groups were compared by Wilcoxon rank-sum test. Differences between groups were considered significant at p < 0.05. Values for all measurements are expressed as the mean ± SEM. All analyses were performed on a Macintosh computer (Apple Computer, Cupertino, CA) with StatView II software (Abacus Concepts, Berkeley, CA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Intranasal administration of 50 µl of RSV (5 x
105 PFU/mouse) induces infection of lung cells
that reaches a peak at 4–6 days after infection. N protein mRNA is
detected in the lung tissues of all live RSV-infected BALB/c mice, but
is not detected in sham- or UV-inactivated RSV-infected mice by RT-PCR
on day 2 after infection (Fig. 2a). RSV Ag is detected
immunohistochemically in lung tissues on day 4 only in RSV-infected
mice using a polyclonal anti-RSV Ab (Fig. 2b) and not in
sham- or UV-inactivated RSV infected mice (not shown). Total RSV Ag
load in the lung tissues was determined by an ELISA 4 days after each
RSV inoculation. No RSV Ag was detected in the control and AS groups of
mice. Both RSV and ASRSV mice showed similar Ag loads after a primary
infection. Following secondary infection, the level of RSV Ag load in
the lung homogenate of both RSV and ASRSV mice significantly increased
compared with that after the primary infection (Fig. 2c).
However, after the tertiary infection, only RSV-infected mice showed a
decline in the level of RSV Ag load in their lung tissue, whereas ASRSV
mice showed a significant increase in the RSV Ag load. Plaque assays
were also performed in mice 4 days after infection; the number of
plaques in RSV and ASRSV mice were similar (mean, 2 x
103 to 104 PFU/g lung; data
not shown).
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The airways of RSV and AS mice, 4 days after primary RSV infection
and 5 days after the final Df challenge, were significantly
more reactive to 50 mg/ml of MCh than were control mice. ASRSV mice
showed a significant increase in MCh responsiveness compared with all
other groups (Fig. 3a). The
increase in AR in RSV mice was observed even 10 days after primary
infection and returned to a normal level by day 17. In addition, the
secondary RSV infection increased AR 4 days after inoculation, but
returned to a normal level by day 10. The tertiary RSV infection
slightly, but not significantly, increased AR 4 days after the
inoculation (Fig. 3b). ASRSV, compared with AS mice, also
exhibited a significant increase in AR 4 days after the secondary
infection. In contrast to the primary RSV infection, this increase in
AR in the ASRSV mice was detectable on day 17 after the secondary
inoculation (Fig. 3c). These results suggest that recurrent
RSV infections attenuate AR in control mice, whereas they increase AR
in allergen-sensitized mice.
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Lung inflammation was examined 4 days after the primary RSV
infection or 5 days after the final Df challenge by staining
with HE or anti-mouse ICAM-1 Abs. Representative pathological
features of HE staining are shown in Fig. 4. Epithelial shedding, mucous plugging,
and mononuclear cell (MNC) infiltration were observed in RSV mice (Fig. 4B) but not in controls (Fig. 4A). Lungs of AS
mice exhibited goblet cell metaplasia and cellular infiltration with
MNC and eosinophils (Fig. 4C). A bronchial exudate, composed
of both MNC and eosinophils, and massive tissue eosinophilic
infiltration were found in ASRSV mice (Fig. 4D). A
semiquantitative analysis using a scoring system for inflammatory cells
in the lung is shown in Table I. The mean
number of infiltrating eosinophils per 10 perivascular areas in ASRSV
mice was significantly higher than that in AS mice (AS, 353.6 ±
16.1 cells; ASRSV, 769.0 ± 20.0 cells; p <
0.01). The results indicate that the lungs of ASRSV mice show
evidence of additive and synergistic effects for epithelial damage and
cellular infiltration, respectively.
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Fig. 5A shows the results
of an immunohistochemical analysis for ICAM-1 in mouse lung tissues.
The number of ICAM-1-positive cells per 200 epithelial cells increased
significantly in both RSV and AS mice compared with that in control
mice (control, 5.0 ± 1.8 cells; RSV, 30.0 ± 4.3 cells; AS,
40.8 ± 2.0 cells; p < 0.01 both). ASRSV
(80.0 ± 20 cells), compared with AS, mice exhibited a 2-fold
increase in ICAM-1-positive epithelial cells. Fig. 5B shows
the quantitation of inflammation in the lungs of both
ICAM-1+/+ and
ICAM-1-/- mice. The
ICAM-1-/- mice exhibited
significantly lower inflammation in all counts than the
ICAM-1+/+ mice, suggesting an important role of
ICAM-1 in inflammation.
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The levels of eotaxin, RANTES, and MIP-1 in a whole lung
homogenate of BALB/c mice were measured 4 days after each inoculation
or 5 days after each final Df challenge. Eotaxin was
comparable between control and RSV-infected mice after all
inoculations. AS mice exhibited a significant increase in eotaxin only
after repeated Df allergen challenges (Fig. 6a). RANTES was significantly
enhanced after primary and tertiary RSV infections and only after
repeated Df allergen challenges (Fig. 6b).
Regardless of the primary or recall response, MIP-1 was
significantly increased in RSV and AS mice (Fig. 6c). Both
eotaxin and RANTES levels in AS mice were identical with those in ASRSV
mice (Fig. 6, a and b), whereas MIP-1 levels
in ASRSV mice were significantly higher than those in AS mice after all
RSV inoculations (Fig. 6c). This enhancement of MIP-1
after repeated infections was significantly higher than that after the
primary infection (primary, 1.3 ± 0.2-fold; secondary, 1.9
± 0.2-fold; tertiary, 2.5 ± 0.4-fold; p < 0.05
for both). These results suggest that RSV infection increases
Df allergen-induced MIP-1 production and that this
increase is further enhanced by recurrent RSV infections.
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Table II shows cytokine production
from anti-CD3-stimulated thoracic lymph node cultures. The primary
RSV infection produced maximal levels of IL-4, IL-5, and IFN-, with
their amounts decreasing following secondary and tertiary infections.
In marked contrast, all Df challenges increased IL-4 and
IL-5, but not IFN-. Compared with AS mice, ASRSV mice exhibited
significant reductions in IL-4 and IL-5 after primary RSV infection.
Secondary and tertiary RSV infections increased IL-4 and IL-5 in ASRSV
mice. Primary infection in AS mice decreased IFN-, but increased
IL-4 and IL-5, whereas secondary and tertiary infections induced
relatively less IFN- and higher levels of IL-4 and IL-5, leading to
a Th2 bias. The ratio of IL-4 to IFN- and that of IL-5 to IFN-
produced in anti-CD3-stimulated thoracic lymph node cultures from
four groups of mice is shown in Fig. 7.
Primary RSV infection induced significant increases in the IL-4:IFN-
and IL-5:IFN- ratios and drastic reductions in these ratios after
secondary and tertiary infections (Fig. 7, a and
b).
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shows the
Df-allergen specific cytokine production from thoracic lymph
node cell cultures. IL-4 and IL-5 were detected in AS and ASRSV mice,
but not in control and RSV mice. Compared with AS mice, ASRSV mice
exhibited significant increases in Df allergen-specific IL-4
and IL-5 productions only after repeated RSV infections. IFN- was
detected in all groups at comparable levels. After primary and tertiary
RSV infections, ASRSV mice showed a significant increase in
Df allergen-stimulated IFN- production compared with AS
mice. In contrast, primary RSV infection in AS mice did not alter the
IL-4:IFN- and IL-5:IFN- ratios; however, secondary and tertiary
infections in these mice resulted in significantly higher IL-4:IFN-
and IL-5:IFN- ratios in ASRSV mice compared with AS mice (Fig. 7
, c and d). Together, these results suggest a shift
toward a Th2-like response after repeated exposure to RSV.
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The serum levels of total IgE and Df-specific IgE and
IgG1 in different groups of mice are shown in Fig. 8. Neither control nor the RSV-infected
mice showed any detectable levels of total serum IgE and
Df-specific IgE or IgG1. ASRSV, compared with AS, mice
exhibited a significant increase in the total serum IgE level 10 days
after the primary RSV infection; however, it became comparable with
that in AS mice by day 17 (Fig. 8a). In contrast to the
primary infection, a significant increase in the total serum IgE level
in ASRSV mice occurred from days 4–17 compared with that in AS mice
after a secondary RSV infection. The difference in
Df-specific serum IgE and IgG1 levels between AS and ASRSV
mice was not statistically significant (Fig. 8, b and
c). These results suggest that RSV infections augment total
IgE Ab levels in Df-sensitized mice.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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in the airway. Furthermore, recurrent RSV infections shifted T cell
responses from a Th0-like response after primary infection toward a
Th2-like response after tertiary infection. This was accompanied by
increases in the RSV Ag load in the lung and the IgE levels in the
serum. Together, these RSV infection-induced changes in the lungs of an
allergen-sensitized mouse may contribute to the development of chronic
asthma. Our results demonstrate that the primary RSV infection in normal mice increases AR; however this increase is transient, as recurrent infections do not lead to any further enhanced AR. In marked contrast, in AS mice, after primary infection AR remained unchanged, while significant increases in AR were noted after secondary and tertiary infections. The differences compared with a previous report (21) were primarily noted with higher concentrations of Mch. This may be due to differences in the sensitization protocol, the allergen, the dose of virus, or the interval between the final challenge and measurement period. Our results are consistent with those of the epidemiological studies, which demonstrated that recurrent RSV infection is common in infancy (24). Also, two or more episodes of wheezing and lower respiratory tract illness are related to an increased risk of subsequent abnormal lung function (25).
The expression of ICAM-1 may constitute one of the most important mechanisms underlying the synergy between RSV infection and allergen sensitization. Df-sensitized and RSV-infected mice, compared with only Df-sensitized mice, showed a significant increase in ICAM-1-positive epithelial cells after the primary RSV infection. Consistent with previous reports (26, 27), both RSV infection and Df sensitization induced the expression of ICAM-1 in epithelial cells. ICAM-1 is implicated in the induction of Ag-induced, airway inflammation and AR in primate and murine models (7, 28). ICAM-1 knockout mice do not show increased AR following RSV infection (29), suggesting an important role of ICAM-1 in RSV-induced AR. Epithelial cells also enhance ICAM-1 expression after exposure to allergens or to RSV via the induction of inflammatory cytokines (26). We have recently found that ICAM-1 facilitates the initiation of RSV infection of epithelial cells (A. K. Behera, H. Matsuse, M. Kumar, X. Kong, F. Lockey, and S. S. Mohapatra, unpublished data). Together, these results indicate that ICAM-1 may constitute a pivotal player in furthering pulmonary inflammation during concomitant allergen sensitization and RSV infection.
Chemokines, such as MIP-1, RANTES, and eotaxin, play important roles
in a variety of immune and inflammatory responses, specifically,
eosinophil migration (6). Our results demonstrate that
primary RSV infection enhances the production of RANTES and MIP-1,
but not eotaxin, in lung tissue. Primary RSV infection in AS mice
enhances MIP-1. These results are consistent with human studies in
which RSV enhanced RANTES and MIP-1 production in cultured
epithelial cells in vitro (30). One human study reported a
significant increase in MIP-1 in nasopharyngeal secretions following
RSV infection (R. C. Welliver, American Academy of Allergy and
Immunology Meeting, 1999). A noteworthy distinction between
Df and RSV exposure is that Df challenges induced
eotaxin, whereas recurrent RSV infection induced enhanced RANTES
production. Interestingly, a significant increase in MIP-1 was found
in ASRSV mice compared with AS or RSV-infected mice, which suggests the
existence of a potential mechanism of synergy between allergen
sensitization and RSV infection. The mechanism of the increased
production of these chemokines upon exposure to RSV and/or AS may
involve a transcriptional up-regulation or might simply reflect a
greater number of cells in the airway rather than an increase in
chemokine production on a per cell basis. Because the number and type
of cells in the lung tissue were not determined in this study, the
basis for MIP-1 remains unclear.
Another important facet of this study is the immune response to RSV
vis-à-vis allergen exposure. Our results clearly demonstrate that
the primary infection, in the absence of allergen sensitization,
induces an aggressive cytokine response, typical of a Th0-like
response, and that further infections induce decreased Th2 or
increased Th1 dominance. However, in allergen-sensitized mice, the
primary infection induced IFN- production, which down-regulated
Th2-like cytokines. Further infections of these mice reversed the
cytokine dominance from Th1-like to Th2-like. Similarly, a comparison
between RSV and ASRSV mice demonstrated that a primary allergen
challenge in RSV mice suppresses both Th1- and Th2-like responses,
while repetitive allergen challenges induce an increase in the
generalized Th2-like response, but not in the Th1-like response, in
ASRSV mice.
Consistent with the Th2-like response, primary RSV infection enhances total serum IgE levels in AS mice, and the magnitude of increase is statistically significant. The increase in total serum IgE is higher after the secondary infection compared with that detected after the primary infection. Thus, the pattern of total serum IgE is correlated with the Th2-like cytokine response and AR observed during each period of infection. Despite increases in total IgE, Df-specific IgE is not significantly different between AS and ASRSV mice. This suggests that the enhanced IgE response observed in ASRSV mice is probably due to the increase in IgE Abs specific to RSV. This inference is consistent with previous reports that RSV is capable of inducing an IgE class switch (31, 32). The RSV G protein especially mounts an IgE response (33, 34). Several investigators reported allergen-specific IgE enhancement in animals with prior RSV (35, 36), influenza (37), or parainfluenza-3 viral (38) infection, suggesting that the virus-induced increase in allergen-specific IgE is possibly due to the facilitation of allergen uptake by virally damaged epithelial cells. It is likely that an increase in Df-specific IgE was not detected in the present study because mice were first sensitized and challenged with allergen and subsequently infected with RSV.
In conclusion, the effect of RSV infection varies depending upon the
inflammatory context of the lung. Both primary and recurrent RSV
infections contribute to and augment ongoing allergic inflammation;
however, in the absence of allergic sensitization, the effects of RSV
are transient. The finding that levels of RSV Ag, MIP-1, and
Th2-like cytokines increase upon recurrent infections is in agreement
with the report that MIP-1 increases the preponderance of Th2-like
cytokines (39). MIP-1 not only promotes eosinophilia
and pulmonary inflammation, but also delays RSV clearance from lung
tissue (40). Our results show that allergen sensitization
and/or RSV infection up-regulate the expression of several molecules,
either additively or synergistically, which leads to the persistence of
inflammation and, consequently, airway hyperreactivity. Thus, RSV
infection increases ICAM-1 expression on nasal epithelial cells
(41) and MIP-1 in the lung tissue, which may enhance
allergic inflammation by augmenting lymphocyte recruitment. Primary
infection also induces IFN-, which has been reported to increase the
survival of eosinophils and thus enhance inflammation in the lung
(42). Recurrent RSV infections produce less IFN- and
enhance a generalized Th2-like response, specifically with increased
IL-5 production. Th2 cells are involved in the pathogenesis of asthma,
resulting in eosinophila and mucous hypersecretion and airway
hyperresponsiveness (43). Overall, recurrent RSV
infections play a critical role in the development of persistent
inflammation and airway hyperresponsiveness in individuals with genetic
predisposition to atopy and, therefore, constitute a major risk factor
for asthma.
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Shyam S. Mohapatra, Division of Allergy and Immunology, Department of Internal Medicine, University of South Florida and James A. Haley Veterans Affairs Hospital, 13000 Bruce B. Downs Boulevard, Tampa, FL 33612.
3 Abbreviations used in this paper: RSV, respiratory syncytial virus; AR, airway responsiveness; AS, Df allergen-sensitized; ASRSV, Df allergen-sensitized plus RSV-infected; Df, Dermatophagoides farinae; HE, hematoxylin-eosin; i.n., intranasally; MCh, methacholine; MIP-1, macrophage inflammatory protein-1 ; MNC, mononuclear cell; Penh, enhanced pause.
Received for publication May 26, 1999. Accepted for publication April 4, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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expressed by a recombinant respiratory syncytial virus attenuates virus replication in mice without compromising immunogenicity. Proc. Natl. Acad. Sci. USA 96:2367. gene transfer protects BALB/c mice against respiratory syncytial virus infection. Vaccine 18:558.[Medline]
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by lower airway epithelial cells and eosinophils infected with respiratory syncytial virus. J. Virol. 72:4756.This article has been cited by other articles:
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 A. L. Miller, C. Gerard, M. Schaller, A. D. Gruber, A. A. Humbles, and N. W. Lukacs Deletion of CCR1 Attenuates Pathophysiologic Responses during Respiratory Syncytial Virus Infection J. Immunol., February 15, 2006; 176(4): 2562 - 2567. [Abstract] [Full Text] [PDF]
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 D. P. Ennis, J. P. Cassidy, and B. P. Mahon Acellular Pertussis Vaccine Protects against Exacerbation of Allergic Asthma Due to Bordetella pertussis in a Murine Model Clin. Vaccine Immunol., March 1, 2005; 12(3): 409 - 417. [Abstract] [Full Text] [PDF]
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 A. K. Behera, E. Hildebrand, J. Scagliotti, A. C. Steere, and L. T. Hu Induction of Host Matrix Metalloproteinases by Borrelia burgdorferi Differs in Human and Murine Lyme Arthritis Infect. Immun., January 1, 2005; 73(1): 126 - 134. [Abstract] [Full Text] [PDF]
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 B. J. Marsland, N. L. Harris, M. Camberis, M. Kopf, S. M. Hook, and G. Le Gros Bystander suppression of allergic airway inflammation by lung resident memory CD8+ T cells PNAS, April 20, 2004; 101(16): 6116 - 6121. [Abstract] [Full Text] [PDF]
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 Y. Kondo, H. Matsuse, I. Machida, T. Kawano, S. Saeki, S. Tomari, Y. Obase, C. Fukushima, and S. Kohno Regulation of Mite Allergen-pulsed Murine Dendritic Cells by Respiratory Syncytial Virus Am. J. Respir. Crit. Care Med., February 15, 2004; 169(4): 494 - 498. [Abstract] [Full Text] [PDF]
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 G. Zimmer, M. Rohn, G. P. McGregor, M. Schemann, K.-K. Conzelmann, and G. Herrler Virokinin, a Bioactive Peptide of the Tachykinin Family, Is Released from the Fusion Protein of Bovine Respiratory Syncytial Virus J. Biol. Chem., November 21, 2003; 278(47): 46854 - 46861. [Abstract] [Full Text] [PDF]
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C.-L. Chen, C.-T. Lee, Y.-C. Liu, J.-Y. Wang, H.-Y. Lei, and C.-K. Yu House Dust Mite Dermatophagoides farinae Augments Proinflammatory Mediator Productions and Accessory Function of Alveolar Macrophages: Implications for Allergic Sensitization and Inflammation J. Immunol., January 1, 2003; 170(1): 528 - 536. [Abstract] [Full Text] [PDF]
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A. K. Behera, M. Kumar, R. F. Lockey, and S. S. Mohapatra 2'-5' Oligoadenylate Synthetase Plays a Critical Role in Interferon-gamma Inhibition of Respiratory Syncytial Virus Infection of Human Epithelial Cells J. Biol. Chem., July 5, 2002; 277(28): 25601 - 25608. [Abstract] [Full Text] [PDF]
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,
sham-infected ICAM-1+/+; •, sham-infected
ICAM-1-/-; 
, RSV-infected
ICAM-1+/+; 
, RSV-infected
ICAM-1-/-.
