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Vol. 288, Issue 1, 43-50, January 1999
Department of Pharmacology, Gifu Pharmaceutical University, Gifu, Japan (H.N., Y.U., H.T., Y.H., N.N., N.I.); Institute of Medical Science, University of Tokyo, Tokyo, Japan (K.T.); and Gifu College of Medical Technology, Seki, Japan (K.K.)
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Abstract |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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The effect of overproduction of interleukin (IL) 5 on the allergic cutaneous response was investigated in transgenic mice overexpressing IL-5. Five repeated topical applications of 2,4-dinitrofluorobenzene (DNFB) to the ears of mice resulted in allergic dermatitis on the ears as well as significant elevation in dinitrophenol-specific IgE antibody and total IgE in the serum in both wild-type and transgenic mice. The development of dermatitis as measured by skin thickness and histopathological changes were potentiated in the transgenic mice. In IL-5 transgenic mice, significant accumulation of eosinophils in skin lesions was observed after five paintings of DNFB, and the magnitudes of eosinophilia and IL-5 messenger RNA expression were significantly higher than in wild-type mice. The dinitrophenol-specific and total IgE in the serum were higher in IL-5 transgenic mice. The late phase reaction of IgE antibody-mediated biphasic cutaneous response was potentiated in IL-5 transgenic mice. The magnitudes of vasopermeability increase by passive cutaneous anaphylaxis, serotonin, and platelet-activating factor were similar in both mice. These results indicate that overproduction of IL-5 resulted in the potentiation of DNFB-induced dermatitis by elevation of IgE production, IgE-mediated allergic late-phase cutaneous reaction, and eosinophilia in the skin lesion.
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Introduction |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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Interleukin (IL) 5 is
mainly produced by T lymphocytes and was originally identified by its
activity as a B cell growth factor (Kinashi et al., 1986
), an
IgA-enhancing factor (Yokota et al., 1987), and a differentiation
factor for the eosinophil lineage (Sanderson, 1992). Recently, much
attention has been paid to the role of IL-5 in pathophysiological
situations. Elucidating the role of IL-5 in allergic airway
inflammation might help define the pathophysiological role of
eosinophilia in allergic inflammation (Devos et al., 1995; van
Oosterhout et al., 1995).
Studies using several animal models of airway inflammation have
revealed the importance of IL-5 for eosinophil infiltration in airways.
Many of these studies employed anti-IL-5 monoclonal antibodies, soluble
receptor 
-chains, IL-5 transgenic mice, and receptor-deficient
mice (Akutsu et al., 1995; Yamaguchi et al., 1994; Nagai et al.,
1993; Lefort et al., 1996; Foster et al., 1996; Iwamoto and Takatsu,
1995). There are suitable methods to analyze the role of IL-5 in
allergic inflammation.
Many clinical investigators have reported the existence of IL-5 in
dermatitis lesions and overproduction of IL-5 by peripheral blood cells
from patients with atopic dermatitis (Yamada et al., 1995; Musial et
al., 1995; Leung, 1995; Tanaka et al., 1994). Many studies have
indicated that IL-5 is important for eosinophilia in allergic cutaneous
lesions. However despite extensive research, the detailed role of IL-5
in allergic skin disease is still obscure.
Therefore, the present study was conducted to investigate the effect of IL-5 overproduction during the onset and development of the allergic cutaneous response in transgenic mice expressing IL-5 under metallothionein promoter.
Recently, we showed that the repeated application of
dinitrofluorobenzene (DNFB) to the ears of mice results in a typical allergic dermatitis and the simultaneous production of IgE antibody against DNFB (Nagai et al., 1997a, b). We have previously demonstrated that the dermatitis consists of helper T 1 (Th1) cell-mediated contact
dermatitis and IgE antibody-mediated cutaneous response. In the present
study, we report that this DNFB-induced allergic cutaneous response is
accelerated by overproduction of IL-5.
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Materials and Methods |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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Animals.
Seven-week-old female transgenic mice weighing 20 to 25 g were prepared and maintained as previously described by
Tominaga et al. (1991). These mice were produced on C3H background,
were rederived as specific pathogen-free, and bred at our laboratory. Age-matched female wild-type C3H/HeN mice weighing 20 to 25 g were
originally obtained from Japan SLC, Inc. (Hamamatsu, Japan) and bred in
our laboratory. Mice were housed in plastic cages in an air-conditioned
room at 24°C, fed a standard laboratory diet, and given water ad
libitum. All experiments were carried out following the guideline for
the care and use of experimental animals made by the Japanese
Association for Laboratory Animal Science in 1987. The serum IL-5 level
in IL-5 transgenic mice was 814.38 ± 120.79 pg/ml (n = 24).
IL-5 was not detected in the serum of wild-type mice.
Reagents and Antibodies.
DNFB was purchased from Nacalai
Tesque Inc. (Kyoto, Japan). Monoclonal antimouse IgE antibody (LO-ME-3)
was from Serotek Co. (Oxford, UK). Peroxidase-conjugated streptavidin
was from Dakopatts a/s (Glostrup, Denmark). Bovine serum albumin (BSA)
[enzyme-linked immunosorbent assay (ELISA) grade] was obtained from
Sigma Chemical Co. (St. Louis, MO). Dinitrophenol (DNP)-BSA was
obtained from LSL Co. (Tokyo, Japan).
N-hydroxysuccinimide-long chain-biotinylation kit was
purchased from Pierce Chemical Co. (Rockford, IL). Peroxidase-labeled antimouse IgE was obtained from Nordic Immunology Co. (Tilburg, The
Netherlands). Goat antimouse IgG and IgM, peroxidase-labeled goat
antimouse IgG, and IgM was obtained from Organon Teknika Co. (Turnhout,
Belgium). The substrate kit was purchased from Sumitomo Bakelite Co.
(Tokyo, Japan). Isogen was from Nippon Gene Co. (Tokyo, Japan).
Polymerase chain reaction (PCR) primers [
-actin, interferon-
(IFN-), IL-4, and IL-5] were purchased from Stratagene Co. (La
Jolla, CA). The 1st STRAND complementary DNA (cDNA) synthesis kit was
from Clontech Lab. (Palo Alto, CA). GeneAmp PCR Reagent kit with
AmpliTaq DNA Polymerase was obtained from Perkin-Elmer Japan Co.
(Urayasu, Japan).
80°C
and used as a source of IgE. The maximum dilution of the preparation
that gave a positive passive cutaneous anaphylaxis (PCA) in Wistar rats
challenged with DNP-BSA was 1:1024.
Procedure for Allergic Dermatitis by Repeated Painting with DNFB.
Experiments were carried out by the method previously described
(Nagai et al., 1997a,b). Mouse ears were painted with DNFB or
just vehicle [acetone/olive oil (3:1)] once each week for 5 weeks.
Twenty-five microliters of 0.15% DNFB in vehicle was applied to each
side of the both ears. Ear thickness was measured using an engineering
micrometer (R1-A, Ozaki MFG Co., Tokyo, Japan) and expressed as the
increase in thickness from time zero. Mouse ears were removed 24 h
after the fifth painting and fixed with 10% neutral Formalin (Nacalai
Tesque, Kyoto, Japan). Ears were then cut into parasagittal
slices, dehydrated, and embedded in paraffin by standard procedures.
Paraffin sections were stained with H&E and assessed by light
microscopy. Individual inflammatory cell types were counted in
high-power fields (HPF; ×200) and expressed as cells per HPF with mean
(±S.E.M.) calculated.
Determination of Hapten-Specific IgE Titer and Ig Concentrations in Mouse Serum. Hapten-specific IgE (sIgE) and immunoglobulins concentrations [total IgE (tIgE), total IgG (tIgG), and total IgM (tIgM)] in mouse serum were measured using the ELISA described below. To measure the concentration of each Ig, serum was obtained from the mice 24 h after the every painting with DNFB.
We measured the hapten sIgE titer by a captured ELISA (Sakaguchi et al., 1989; Hirano et al., 1989). Briefly, immunoplates (EIA II Plus
microtitration plate; Flow Laboratories Inc. Mclean, VA) were
coated with monoclonal antimouse IgE antibody and incubated at 4°C
overnight. The plates were blocked with phosphate-buffered saline (PBS)
containing 1% BSA, incubated at room temperature for 1 h, and
washed three times with PBS containing 0.2% Tween 20 (T-PBS).
Monoclonal anti-DNP IgE antibody (monoclonal anti-DNP IgE, clone SPE-7;
Sigma Chemical Co.) was sequentially diluted as the standard. Diluted
serum sample (100 µl) was added to each well and the plates were
incubated at room temperature for 1 h. After washing with T-PBS,
100 µl of diluted biotinylated DNP-BSA was added to each well and
incubated at room temperature for 1 h. After extensive washing
with T-PBS, 100 µl of diluted peroxidase-conjugated streptavidin was
added to each well. The enzymatic reaction was stopped by adding 100 µl of stop solution after an incubation at room temperature for
1 h. The optical density of the reaction mixture was read using an
automatic ELISA plate reader (Titertek Multiscan MCC/340, Flow
Laboratories) at 450 nm.
To measure total Ig concentrations (tIgE, tIgG, and tIgM), immunoplates
were coated with diluted monoclonal anti-IgE, goat antimouse IgG, or
goat antimouse IgM antibody, respectively, by incubating overnight at
4°C. The plates were blocked as described above and washed three
times with T-PBS. Standard curves were generated as described above by
employing standard IgE, standard mouse IgG (Miles Scientific,
Naperville, IL), and standard IgM (Miles). Serum sample (100 µl) was
added to each well and incubated at room temperature for 1 h.
ELISA was performed using peroxidase-conjugated antimouse IgE, IgG, and
IgM. The ELISA data compared with the standards added to each plate
were analyzed using the Delta Soft program (Bio Metallics Inc., McLean,
VA) for the Macintosh computer (Apple Computer Inc., Cupertino,
CA). The sIgE, tIgE, tIgG, and tIgM titers are expressed in
micrograms per milliliter based on laboratory-generated standards and
appropriate commercial standards.
Analysis of Cytokine Messenger RNA (mRNA) Expression in Mouse
Auricular Lymph Nodes and Ears by Reverse Transcriptase-PCR (RT-PCR).
Changes in cervical lymph node and ear-derived cytokine mRNA
levels were assessed by RT-PCR using a thermal cycler (Bio Metra Trio-Thermoblock, Bio Metra Co., Göttingen, Germany).
Using Isogen, total RNA was prepared from the ears and lymph nodes of
the mice 4 h after being painted five times with either vehicle or
DNFB. The amount of total RNA in each sample was measured
spectrophotometrically at 260 nm and quality was checked by
electrophoresis. RT-PCR was performed as follows. Total RNA (500 ng)
was reverse-transcribed for 60 min at 42°C using the 1st STRAND cDNA
synthesis kit. Each cDNA sample was amplified in a total volume of 100 µl containing 0.5 µM each primer (RT-PCR primers set from the
GeneAmp PCR reagent kit). The internal control was -actin. The
mixture was overlaid with mineral oil and put in the thermal cycler for
35 cycles. RT-PCR was performed on actin, IFN-, IL-4, and
IL-5. The PCR products were resolved by electrophoresis and stained
with ethidium bromide. Samples were obtained from two mice whose ears
and cervical lymph nodes were removed 4 h after the fifth painting
with DNFB. RT-PCR was semiquantified by densitometrically scanning
photo negatives produced using a Polaroid camera (Poraloid 665 film; Polaroid Corp., Cambridge, MA). For semiquantitation, the densitometry value of each cytokine was normalized to that of the housekeeping gene,
-actin, which was not affected by the DNFB concentrations applied in
this study. In addition, a linear correlation between RNA input and PCR
product was examined. Fair linearity was obtained between the density
value of PCR product and RNA input. All PCR amplifications were
performed at least twice with multiple sets of experimental RNAs.
Purification of Total RNA and Analysis of Productive C
Gene
Expression by RT-PCR.
RT-PCR was performed as described above to
assess the changes in cervical lymph node-derived productive C mRNA
levels. RNA (500 ng) was reverse-transcribed for 60 min at 42°C using
the 1st STRAND cDNA synthesis kit. The cDNA samples were amplified in a
total volume of 100 µl containing 0.5 mM' and 3' primers (primers
sequences for productive C, 5' primer j4: TGGACTACTGG GGTCAAGG, 3'
primer C2: AGCGATGAATGGAGTAGC) with GeneAmp PCR reagents (GeneAmp
PCR Reagent kit with AmpliTaq DNA polymerase.
-actin and productive C. The internal
control was -actin. Each PCR product was resolved by electrophoresis
and visualized with ethidium. Auricular lymph nodes were obtained
4 h after the fifth painting with DNFB.
IgE-Mediated Biphasic Cutaneous Reaction.
This method was
described previously (Nagai et al., 1995). In brief, mice received two
i.v. injections (5-min interval) of 0.5 ml of monoclonal IgE
preparation. Twenty-four hours after the sensitization, a skin reaction
was elicited by applying 25 µl of 0.15% DNFB acetone-olive oil
solution to both sides of the both ears. The reaction was assessed by
measuring the ear thickness using an engineer's micrometer (Upright
Dial Gauge; Peacock, Tokyo, Japan) at different times after challenge.
Data were expressed as an increase in ear thickness after antigen
challenge by subtracting the value measured immediately before
challenge. Because we had previously confirmed that the vehicle for the
monoclonal IgE preparation did not affect the skin reaction in the
present experimental protocol, we injected saline into control mice
instead of the IgE preparation.
Vascular Permeability Increase by PCA, Serotonin, and
Platelet-Activating Factor (PAF) in Mouse Ear.
Mouse ear PCA was
carried out according to methods described previously (Inagaki et al.,
1988). Briefly, 10 µl of diluted anti-DNP monclonal IgE antibody was
injected into back skin under light anesthesia. After 48 h, PCA
was elicited by i.v. injection of 0.25 mg of DNP-BSA dissolved in 0.25 ml of 0.5% Evans blue saline solution. Thirty minutes after the
antigen challenge, mice were sacrificed by cervical dislocation and the
bluing skin area removed. To measure the amount of extravasated dye,
bluing skin was dissolved with 0.7 ml of 1 N KOH solution in a
stoppered tube at 37°C overnight and 9.3 ml of a mixture of 0.6 N
H3PO4 solution and acetone
(5:13) was added. After vigorous shaking, precipitates were filtered
off and the the amount of dye was measured colorimetrically at 620 nm.
Increase in vasopermeability by mediators was initiated by the
following. Mice back skin received 10 µl each of serotonin (3 × 10
6 g/ml) and PAF
(2x106 g/ml) solution. Immediately after the
injection of mediator, 0.5% Evans blue saline solution was injected
i.v. After 30 min, mice were sacrificed and the extravasated dye was
measured by the method described above.
Statistical Analyses. Results are expressed as the mean ± S.E.M. Data were evaluated by either Student's or Welch's t test after examining the variances using the F test. p < .05 was considered to be statistically significant.
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Results |
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Abstract
Introduction
Materials & Methods
Results
Discussion
References
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Dermatitis Caused by Repeated Painting with DNFB.
Repeated
topical application (painting) of DNFB on the ear provoked typical
contact dermatitis in mice. Figure 1
shows the time course for the changes in ear thickness due to
dermatitis in IL-5 transgenic and wild-type mice. The thickness
increased in proportion to the increase in the number of exposures to
DNFB. The ear thickness at time zero was 21.68 ± 0.18 × 102 mm. The maximum response of the dermatitis
was detected 24 h after the second, third, fourth, and fifth
painting with DNFB. Our previous studies have shown the response at
24 h consists of a mixture of contact dermatitis and IgE antibody-
mediated late-phase cutaneous response. The increase in ear thickness
at 24 h after each painting with DNFB was potentiated in IL-5
transgenic mice when compared with the response in wild-type mice. The
increased thickness of the ears in IL-5 transgenic mice was 29.44 ± 2.75 × 102 mm, and that in wild-type
mice was 16.89 ± 0.60 × 102 mm
24 h after the fifth painting. The maximum response was a 3-fold
increase in ear thickness. No significant increase in ear thickness was
apparent in the mice treated with vehicle and naive mice during the
experiment.
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and IL-5) in the ear was measured
by RT-PCR (Fig. 3). Semiquantitative data
were expressed in the bar graph. Each column represents the mean of three experiments. In the ear of IL-5 transgenic animals, IFN- mRNA
expression after antigen provocation was almost the same compared with
that in wild-type mice, whereas the expression of IL-5 mRNA was 14 times stronger than that in wild-type mice. IFN- mRNA in
lymph nodes was constitutively expressed and did not change after
antigen challenge in both wild-type and IL-5 transgenic mice (data not
shown). IL-4 mRNA was expressed in the lymph node but not in the ear
after antigen challenge in both animals (data of IL-4 mRNA in the skin
not shown).
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Ig Production by Contact Sensitization with DNFB.
As indicated
in Table 2, the levels of hapten sIgE and
tIgE in the serum from IL-5 transgenic mice after the fifth painting with DNFB was almost a 1.9 and 1.6 times increase from those in wild-type mice. The tIgG and tIgM levels were originally high in
transgenic mice when compared with those in wild-type mice. The tIgG
level in transgenic mice after DNFB treatment was significantly lower
than that in wild-type mice, but the tIgM level in transgenic mice was
higher than that in wild-type mice. To investigate the effect of IL-5
overproduction on the IgE production, we investigated the expressions
of IL-4 mRNA and productive C mRNA in the lymph node. As shown in
Fig. 4, the expression of both mRNAs in
the lymph node was similar in IL-5 transgenic and wild-type mice. In
addition, the expression of productive C mRNA in the spleen was also
examined. The magnitude of the expression of productive C mRNA in
transgenic and wild-type mice spleen was almost the same (data not
shown). The serum level of each Ig in the wild-type and transgenic mice
was not altered by the treatment with vehicle (data not shown).
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IgE-Mediated Biphasic Cutaneous Reaction. Mice were sensitized with anti-DNP IgE and challenged with antigen epicutaneously. Results of a time-course study are shown in Fig. 5. Intravenous application of IgE and the subsequent epicutaneous antigen challenge induced a cutaneous reaction in wild-type and IL-5 transgenic mice. A significant increase in ear thickness was observed at 1 and 24 h after antigen challenge in sensitized animals (p < .01 from normal animals). In control mice, which received an i.v. injection of saline instead of monoclonal IgE, epicutaneous application of DNFB did not cause a significant cutaneous reaction (data not shown). The magnitude of the response at 1 h after antigen challenge was identical for wild-type and IL-5 transgenic mice, whereas the magnitude of the late-phase response at 4 and 24 h after challenge in IL-5 transgenic mice was greater than in wild-type mice. As shown in the Fig. 6, marked eosinophilia, dermal edema, vascular dilatation, and epidermal hyperplasia were observed in the skin lesions of IL-5 transgenic mice 24 h after antigen application. In wild-type, C3H/HeN mice, many neutrophils and a few eosinophils were seen in the dermis and epidermis 24 h after antigen challenge.
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Increase in Vasopermeability by PCA, Histamine, Serotonin, and PAF in Mouse Ear. To examine the susceptibility of skin to allergic mediators, the increase in vasopermeability by PCA, histamine, serotonin, and PAF was compared in IL-5 transgenic and wild-type mice. The increase in vascular permeability caused by PCA, serotonin, and PAF was similar in IL-5 transgenic and wild-type mice (Fig. 7).
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Discussion |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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In the present study, we demonstrated that allergic contact
dermatitis caused by repeated sensitization with DNFB was potentiated in transgenic mice expressing IL-5 under a metallothionein promoter. The present results also show that the accelerated dermatitis is
associated with increases in IL-5 mRNA expression in the skin, serum
IgE level, IgE antibody-mediated late-phase cutaneous response, and
skin eosinophilia, but is not associated with the increase in IFN-
mRNA expression in the ear and susceptibility to allergic mediators.
The immunological mechanisms involved in the initiation and
development of allergic skin inflammation are poorly understood. Recent
data suggest that the local expression pattern of cytokines plays a
critical role in modulating the nature of tissue inflammation. Hamid et
al. (1994) reported that although acute and chronic atopic dermatitis
lesion are associated with increased activation of both IL-4 and IL-5
genes, initiation of acute skin inflammation in dermatitis is
associated with a predominance of IL-4 expression, whereas maintenance
of chronic inflammation is predominantly associated with increased IL-5
expression and eosinophil infiltration. Moreover, many investigators
report the existence of IL-5 in the lesions of dermatitis and
overproduction of IL-5 by the peripheral blood cells of patients with
allergic dermatitis (Yamada et al., 1995; Musial et al., 1995; Leung
1995; Tanaka et al., 1994). These reports suggest that the production
of IL-5 may play an important role in initiation or maintenance of
allergic dermatitis in humans. In the present study, we demonstrated
that IL-5 overproduction resulted in the potentiation of allergic
cutaneous inflammation.
IL-5 mRNA was detected in IL-5 transgenic mice ear skin, indicating a high production of IL-5 in the skin lesion of transgenic mice even without antigen stimulation. However, no significant histopathological changes were observed in the mice without antigen stimulation. Only a slight migration of eosinophils was observed in the ear skin of IL-5 transgenic mice. Overproduction of IL-5 without antigen stimulation did not cause dermatitis. However, when antigen was applied to the skin of mice, severe dermatitis with a significant elevation of IL-5 mRNA expression resulted. The magnitude of dermatitis was more severe in transgenic than in wild-type mice. These data suggest that overproduction of IL-5 is not an initiation factor but a potentiating factor for allergic dermatitis.
To investigate the mechanism for potentiation of dermatitis, we
examined the antigen-induced expression of cytokine mRNA in IL-5
transgenic mice. As reported previously, antigen stimulation caused
strong expression of IFN- in the skin lesions and IL-4 and IL-5
mRNAs in the lymph nodes in BALB/c mice (Nagai et al., 1997a). The
expression of IFN- in the lymph node was detected without antigen
stimulation and did not change with repeated antigen stimulation.
Simultaneous expression of IL-4 and IL-5 mRNAs in the skin lesion was
not detected or was very low even after antigen stimulation in BALB/c
mice. In the present study, a similar phenomenon was observed in
wild-type C3H/HeN mice. Simultaneously, antigen-induced strong
expression of IFN- and IL-5 mRNAs was observed in IL-5 transgenic
mice. However, the magnitude of IFN- mRNA was almost the same
between wild-type and IL-5 transgenic mice. Previous studies have shown
involvement of Th1-derived IFN- in contact dermatitis (Thomson et
al., 1993). Therefore, the present allergic contact dermatitis is due
to Th1 response in IL-5 transgenic mice and the allergic contact
dermatitis is a partial feature of Th2 (IL-5 transgenic) mice. In
addition, the present data indicate that the overproduction of IL-5
does not affect antigen-induced, Th1-derived IFN- mRNA expression in
the skin. To the contrary, the antigen stimulation significantly
potentiated the expression of IL-5 mRNA in IL-5 transgenic mice. It may
be closely related to the severe eosinophilia in the skin lesion.
However, the reason why antigen stimulation in IL-5 transgenic mice
amplifies the expression of IL-5 mRNA is still obscure. The antigen
stimulation may accumulate many kinds of IL-5 overexpressing cells
including mast cells, eosinophils, and T lymphocytes in the skin
lesion. Further experiments will be necessary to elucidate this
amplifying mechanism of IL-5 expression.
In the histopathological studies of skin lesions, marked eosinophilia
was observed in IL-5 transgenic mice. Many investigators have reported
that IL-5 plays a dominant role in eosinophilia in the skin, and severe
infiltration of eosinophils in skin lesions may accelerate dermatitis
(Yamada et al., 1995; Tanaka et al., 1994; Hamid et al., 1994). In the
present study, IgE-mediated late-phase cutaneous response was
potentiated in IL-5 transgenic mice, whereas IgE-mediated
immediate-phase reaction, PCA reaction, and mediator-mediated
permeability increase were not. These data show that the increase in
eosinophilic inflammatory response at 4 and 24 h in IL-5
transgenic mice caused the increase in ear thickness without a direct
relationship to the serum IgE level or immediate cutaneous reaction.
The relationship between IL-5 and late-phase cutaneous reaction has not
been clarified. But many reports indicate that late-phase cutaneous
response involves eosinophilia. Studies suggest the eosinophil is the
most influential cell in late-phase allergic reaction (Litchfield et
al., 1996). We observed marked eosinophilia in skin lesions 24 h
after the fifth antigen stimulation and in the IgE-mediated late-phase
cutaneous reaction in IL-5 transgenic mice. These results suggest that
activation of eosinophils is very important in the acceleration of
allergic dermatitis in IL-5 transgenic mice. Currently, we are
investigating the kinds of eosinophilic molecules expressed during
dermatitis by using in situ hybridization.
We also found potentiation of IgE production in IL-5 transgenic mice
after antigen stimulation. The reason for this acceleration is still
obscure. There is evidence to indicate a direct relationship between
overproduction of IL-5 and an acceleration of IgE antibody production.
Lim-Bo et al. (1995) reported that IL-5 potentiates the production of
IgE by mouse splenic lymphocytes stimulated with bacterial
lipopolysaccharide. Zhou et al. (1996) also reported a correlated
increase in IgE production to IL-5 mRNA-positive cells in
Nippostrongylus brasiliensis infected mice. Purkerson and
Isakson (1992) also reported that IL-5 in addition to IL-4 may
provide a signal to isotype switching to IgE in mice. These reports
have indicated a direct relationship between IL-5 and IgE production.
In the present study, we showed that the expression of both IL-4 mRNA
and the productive C gene are not altered by repeated antigen
stimulations in IL-5 transgenic mice. Sewell and Mu (1996) reported
that IL-4 and -5 are not always coordinately produced in parasitic
infestation and allergic disease. In addition, Brewer et al. (1996)
reported that there was no significant difference in IL-5 production
between IL-4-deficient and wild-type mice when mice were immunized
with alum and ovalbumin. These results, as well as our own, suggest
that IL-5 overproduction does not effect IgE production through IL-4 at
the gene level. To clarify this, we are currently investigating the
functions of T and B cells by using cultured cells from transgenic mice.
In conclusion, the present study indicates that an allergic cutaneous response caused by repeated painting with DNFB is potentiated in IL-5 transgenic mice when compared with wild-type mice. This potentiation may be related to the augmentation in IgE antibody production, IgE antibody-mediated late-phase cutaneous reaction, and eosinophilia in the skin lesion.
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Footnotes |
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Accepted for publication July 23, 1998.
Received for publication March 17, 1998.
Send reprint requests to: H. Nagai, Department of Pharmacology, Gifu Pharmaceutical University, 5-6-1 Mitahorahigashi, Gifu 502-8585, Japan. E-mail: nagai{at}gifu-pu.ac.jp
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Abbreviations |
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IL, interleukin; DNFB, dinitrofluorobenzene; DNP, dinitrophenol; PAF, platelet-activating factor; PCA, passive cutaneous anaphylaxis.
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References |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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in the treatment of atopic dermatitis: Influence on T-cell activation.
Allergy
50:
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, p < .01 (from
wild-type mice). WT, Wild-type mice; IL-5 Tg, IL-5 transgenic mice
