![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
Vol. 283, Issue 1, 321-327, 1997
Department of Pharmacology, Gifu Pharmaceutical University, 5-6-1 Mitahorahigashi, Gifu 502, Japan (H.N., H.H., A.M., Y.U., N.I.) and Gifu College of Medical Technology, 795-1 Nagamine, Ichihiraga, Seki 501-32, Japan (K.K.)
 |
Abstract |
|---|
 Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
Five repeated topical applications of 2,4-dinitrofluorobenzene to the
ears of BALB/c mice resulted in contact dermatitis on the ears as well
as significant elevation in dinitrophenol-specific IgE antibody and
total IgE in the serum. FK-506 and cyclosporin A inhibited the
development of contact dermatitis in terms of skin thickness and
histopathological changes of skin lesions. On the contrary, these two
drugs potentiated dinitrophenol-specific and total IgE antibody
production without affecting IgG and IgM levels in serum. The
expression of interferon-
mRNA in reverse transcriptase-polymerase
chain reaction in the ear was inhibited by FK-506 and cyclosporin A. The expression of interleukin-4 mRNA, germline C
and productive C
in the auricular lymph node was not affected by these two drugs.
Contrary to the above in vivo findings, the
immunosuppressors, FK-506 and cyclosporin A, inhibited the production
of interferon- and interleukin-2 by cultured Th1 cells (1E10.H2
cells) and of interleukin-4 and -5 by Th2 cells (D10.G4.1 cells)
in vitro. These results indicated that FK-506 and
cyclosporin A selectively inhibited the Th1 cell-mediated contact
dermatitis and potentiated the Th2 cell-mediated IgE antibody production in vivo. This potentiation is probably due to
the down-regulation of interferon- production by Th1 cells after the
treatment with these drugs. However, because FK-506 and cyclosporin A
inhibited the production of cytokines by both Th1 and Th2 cells
in vitro and these two immunosuppressors showed higher
selectivity toward inhibiting Th1 cell-mediated reactions by
limitations in vivo experiments.
| |
Introduction |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
FK-506
and cyclosporin A are novel immunosuppressors derived from fungi. These
two agents inhibit T cell proliferation, cytotoxicity and cytokine
production without destroying cells (Kino et al., 1987
;
White, 1982). Their action is mainly based on the inhibition of
calcium/calmodulin-activated protein phosphatase, calcineurin (Liu
et al., 1991). Because a calcium/calmodulin-activated
calcineurin function is required to activate the cytoplasmic component
of the transcription NF-AT, both agents block the NF-AT activation and
transcription of many cytokine genes (McCaffrey et al.,
1993). This net result of the action is that these two
immunosuppressors interfere with T cell activation (Dumont et
al., 1990; Lin et al., 1991).
There are several different types of T cells, with a variety of
functions. One group interacts with B cells or mononuclear cells to
help them produce antibody or destroy intracellular pathogens. This
group of cells is called Th cells. Th cells can be divided into
different subsets depending on their cytokine profile. Th1 cells
produce IL-2 and IFN- but not IL-4 and IL-5 and are chiefly responsible for the delayed-type hypersensitivity response (Cher and
Mosmann, 1987; Mosmann et al., 1986). They can also help B cells to produce IgG2a but not much IgG1 and IgE, in the mouse. On the
contrary, Th2 cells produce IL-4 and IL-5 but not IL-2 and IFN-.
They are efficient helper cells for antibody production, especially
IgG1 and IgE.
Regarding the production of IgE, IL-4 and IL-13 play an important role
under the regulation with IFN-. Fuleinham et al. (1995) reported that cyclosporin A suppressed the T cell-dependent
IL-4-derived IgE synthesis in human lymphocytes. In addition, some
other investigators reported that suppressive effect of cyclosporin A
on the production of IgE (Toorenenbergen et al., 1996;
Vendeville et al., 1995; Puignero et al.,
1995). On the contrary, there are some reports to indicate the
potentiation of IgE production by immunosuppressors including
cyclosporin A and FK-506 (Bundick et al., 1995; Wheeler et al., 1995; Chang et al., 1993; Chen, 1988;
Wang et al., 1993). Because IgE is the most important
antibody in the development of allergic reaction and some
immunosuppressors including FK-506 and cyclosporin A are applied to
treat some allergic diseases, such as bronchial asthma and atopic
dermatitis, it is important to clarify the effect of FK-506 and
cyclosporin A on IgE antibody production in terms of the production of
cytokines.
Recently, we indicated that the repeated application of DNFB to the mouse ear results in a typical contact dermatitis in the ear and the simultaneous production of IgE antibody against DNFB (H. Nagai et al., submitted for publication). We demonstrated that contact dermatitis is mainly mediated by Th1 cells and that IgE antibody production is mediated by Th2 cells. Our study was therefore conducted to study the effect of FK-506 and cyclosporin A on Th2 cell-mediated IgE production and Th1 cell-mediated contact dermatitis by contact sensitization with DNFB in mice in vivo. In addition, we examined the effect of these drugs on the activation of Th1 and Th2 cells in vitro.
| |
Materials and Methods |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
Animals. Female BALB/c mice (7-10 wk old) obtained from Japan SLC, Inc. (Hamamatsu, Japan), 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 a guideline for the care and use of experimental animals made by the Japanese Association for Laboratory Animal Science in 1987.
Reagents and antibodies.
FK-506 and cyclosporin A were
provided by Fujisawa Pharmaceutical Co., Ltd., Osaka, Japan and Sandoz
Japan Co., Ltd., Tokyo, Japan, respectively. FK-506 was suspended in
0.5% sodium carboxymethylcellulose saline solution and orally
administered three times a week for 5 wk. Cyclosporin A was dissolved
in myglyol and administered orally three times a week for 5 wk. DNFB
was purchased from Nacalai Tesque Inc., Kyoto, Japan. Monoclonal
anti-mouse IgE antibody (LO-ME-3, Serotek Co., Ltd., Oxford, UK),
monoclonal anti-DNP IgE antibody (Clone SPE-7, Sigma Chemical Co.,
Ltd., St. Louis, MO), peroxidase-conjugated streptavidin (Dakopatts
a/s, Glostrup, Denmark), (for ELISA grade, Sigma Chemical Co., Ltd.),
DNP-BSA (LSL Co., Ltd., Tokyo, Japan), NHS-LC-Biotinylation kit (Pierce Chemical Co., Ltd., Rockford, IL), peroxidase labeled anti-mouse IgE
(Nordic Immunology Co., Ltd., Tilburg, Netherlands), goat anti-mouse
IgG and IgM, peroxidase labeled goat anti-mouse IgG and IgM (Organon
Teknika Co., Turnhout, Belgium), substrate kit (Sumitomo Bakelite Co.,
Ltd., Tokyo, Japan), ISOGEN (Nippon Gene Co., Ltd., Tokyo, Japan), PCR
primers (
-actin, IL-2, IFN-, IL-4, IL-5; Stratagene Co., Ltd., La
Jolla, CA), 1st-STRANDED TM cDNA Synthesis kit (Clontech Lab, Inc.,
Palo Alto, CA), PCR reagent kit (GeneAmp PCR Reagent Kit with AmpliTaq
DNA Polymerase, Perkin Elmer Japan Co., Ltd., Urayasu, Japan) were
purchased from the listed suppliers.
Procedure for painting ears with DNFB and for measuring the ear thickness. Mouse ears were painted with DNFB or vehicle once each week for 5 wk. A total of 25 µl of 0.15% DNFB in vehicle [acetone:olive oil (3:1)] was applied to each side of the ear. Ear thickness was measured using an engineering micrometer (R1-A, Ozaki MFG Co., Ltd., Tokyo, Japan) and is expressed as the increase in thickness from that at time 0.
Histopathological study of skin lesion. Mouse ears were removed 24 hr after the fifth painting with vehicle or DNFB and fixed with 10% neutral formalin. Ears were then cut into parasagittal slices, dehydrated and embedded in paraffin by standard procedures. Paraffin sections were stained with hematoxylin and eosin, then assessed by light microscopy.
Determination of hapten-specific IgE titer and immunoglobulin 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 immunoglobulin, serum was obtained from the mice 24 hr after the fifth 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, ICN Biomedicals, Costa
Mesa, CA) were coated with monoclonal anti-mouse IgE antibody and
incubated at 4°C overnight. The plates were blocked with PBS
containing 1% BSA and incubated at room temperature for 1 hr and
washed three times with T-PBS. Monoclonal anti-DNP IgE antibody was
sequentially diluted as the standard. Diluted serum samples at a volume
of 100 µl were added to each well and the plates were incubated at room temperature for 1 hr. After washing with T-PBS, 100 µl of diluted biotinylated DNP-BSA were added to each well and incubated at
room temperature for 1 hr. After extensive washing with T-PBS, 100 µl
of diluted peroxidase-conjugated streptavidin were added to each well.
The enzymatic reaction was stopped by adding 100 µl of stop solution
after an incubation at room temperature for 1 hr. The optical density
of the reaction mixture was read using an automatic ELISA plate reader
(Titertek Multiscan MCC/340, ICN Biochemicals, Costa Mesa, CA) at 450 nm.
To measure the total immunoglobulin concentrations (total IgE, G and
M), immunoplates were coated by incubating overnight at 4°C with
diluted monoclonal anti-IgE, goat anti-mouse IgG or goat anti-mouse IgM
antibody, respectively. 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 (monoclonal anti-DNP IgE,
clone SPE-7, Sigma), standard mouse IgG (Miles Scientific, Napervilla,
IL) and standard IgM (Miles). A total of 100 µl of serum sample was
added to the each well and incubated at room temperature for 1 hr.
ELISA was performed using peroxidase-conjugated anti-mouse IgE, IgG and
IgM. The ELISA data compared with the standards added to each plate
were analyzed out using the DELTA Soft program for the Macintosh
computer. The sIgE, tIgE, tIgG and tIgM titers are expressed in µg/ml
based on laboratory-generated standards and appropriate commercial
standards.
Analysis of cytokine mRNA expression in mouse auricular lymph
nodes and ears by RT-PCR.
Changes in cervical lymph node and
ear-derived cytokine mRNA levels were assessed by the RT-PCR using a
thermal cycler (Bio Metra Trio-Thermoblock, Bio Metra Co., Ltd.,
Gottingen, Germany). Using ISOGEN, total RNA was prepared from the ears
and lymph nodes of the mice 4 hr after being painted five times with
either vehicle or DNFB. The amount of total RNA in each sample was
measured spectrophotometrically at a wavelength of 260 nm and the
quality of the RNA 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 TM cDNA Synthesis kit. Each cDNA sample was
amplified in a total volume of 100 µl containing 0.5 µM of each
primer (RT-PCR primers set from the GeneAmp PCR Reagent kit). The
internal control was -actin. The mixture was overlaid with mineral
oil, then 30 and 35 PCR cycles proceeded in a thermal cycler. RT-PCR
was performed on -actin, IFN-, IL-2, IL-4 and IL-5. The PCR
products were resolved by electrophoresis and stained with ethidium
bromide to reveal the DNA. Samples were obtained from three mice. The ears and cervical lymph nodes were removed 24 hr after the fifth painting with DNFB. RT-PCR was semi-quantified by densitometrically scanning photo negatives produced using a Polaroid camera (Poraloid 665 film, Polaroid Corp., Cambridge, MA). For relative semiquantitation, the densitometry value of each cytokine was normalized to that of the
house keeping gene, -actin, which was not affected by DNFB within
the 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 germline and productive
C gene expression by RT-PCR.
RT-PCR proceeded as described
above to assess the changes in cervical lymph node-derived germline and
productive C mRNA levels. RNA (500n ng) was each of which was
reverse-transcribed for 60 min at 42°C using the 1st-STRANDED TM cDNA
Synthesis kit (Clontech Lab., Palo Alto, CA). The cDNA samples were
amplified in a total volume of 100 µl containing 0.5 mM of each 5
and 3 primers (primers sequences for germline C, 5 primer I:
ACTAGAGATTC ACAACG, 3 primer C2: AGCGATGAATGGAGTAGC; primers
sequences for productive C, 5 primer j4: TGGACTACTGGGGTCAAGG, 3
Primer C2: AGCGATGAATGGAG TAGC) with the GeneAmp PCR reagents
(GeneAmp PCR Reagent kit with AmppliTaq DNA Polymerase: Perkin-Elmer
Japan Co., Ltd., Urayasu, Japan).
-actin, germline C and productive C. Internal control was -actin. Each PCR product was resolved by electrophoresis and visualized with ethidium bromide to reveal the DNA. Auricular lymph
nodes were obtained 24 hr after the fifth painting with DNFB.
Cytokine production from cultured murine T cell clones stimulated
with chemicals.
The Th1 cell clone 1E10.H2 was kindly donated by
Prof. H. Oomori (Okayama University, Okayama, Japan). This clone is
specific for keyhole limpets hemocyanin in the context of
I-Ak. The Th2 clone D10.G4.1 (Dainippon Seiyaku,
Osaka, Japan) is specific for conalbumin in the context of
I-Ak. These two T cell clones were stimulated
every week with antigen and mitomycin C-treated syngeneic splenocytes
as antigen-presenting cells. The supernatant containing
-methyl-D-mannoside from rat spleen cells that had been
stimulated with concanavalin A for 48 hr was added at a ratio of 10%
as a source of lymphokines, to complete medium during cell culture.
Cells of each T cell clone were studied after stimulation with antigen
and mitomycin C treated splenocytes for 7 to 14 days. Cells were
collected by centrifugation and resuspended in RPMI 1640 medium
containing 10% fetal calf serum to a density of 3 ×105 cells/ml. Each cell was stimulated with 10 ng/ml PMA, 0.6 µg/ml calcium ionophore A23187 and 10 µg/ml
concanavalin A for 24 hr. The amount of cytokine in the supernatant was
measured by enzyme-linked immunmoassay. At least four doses of drugs
were tested and the IC50 value with 95%
confidence limits were calculated by linear regression.
Statistics. 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.
| |
Results |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
Effect of FK-506 and cyclosporin A on Th1 cell-mediated contact
dermatitis..
Repeated topical application (painting) of DNFB on
the ear skin provoked typical contact dermatitis in mice. Figure
1 shows the time course for the effect of
FK-506 and cyclosporin A on the changes in ear thickness due to
dermatitis. The thickness increased in proportion to the increase in
the number of exposure to DNFB. The ear thickness at time 0 was
21.68 ± 0.18 ×10
2 mm. Contact
dermatitis was detected 24 hr after the second, third, fourth and fifth
painting with DNFB. The increase in ear thickness at 24 hr after each
painting with DNFB was suppressed by FK-506 and cyclosporin A. Each
drug at a high dose suppressed the increment of the ear almost 50%.
Figure 2 shows the effect of FK-506 and cyclosporin A on the histopathological changes of the mice skin lesions
24 hr after the fifth painting with DNFB. Marked infiltration of
inflammatory cells such as neutrophils, eosinophils and monocytes and
hypertrophy of the epidermis was evident in the control group. FK-506
and cyclosporin A suppressed the infiltration of inflammatory cells and
hypertrophy of the epidermis. No significant histopathological change
was apparent in the mice given vehicle (data not shown). To investigate
the effect of these drugs on the expression of cytokine mRNAs, RT-PCR
concerning the IL-2, IFN-, IL-4 and IL-5 mRNAs in the ear and lymph
nodes was carried out. In the preliminary experiments, IFN- and IL-2
mRNA expression was detected in the ear and lymph node of control
animals. The expression of both mRNAs was increased after the antigen
application in the ear but not in the lymph node. Contrary, IL-4 and
IL-5 mRNAs were expressed in the lymph node but not in the ear by the
antigen challenge. As shown in figure 3,
the expression of IFN- mRNA in the ear was inhibited by FK-506 and
cyclosporin A whereas IL-4 mRNA in the lymph node was not affected.
|
|
|
Effect of FK-506 and cyclosporin A on the IgE antibody production
by contact sensitization with DNFB.
As indicated in figure
4, the levels of hapten sIgE and tIgE
significantly increased in the serum from mice painted five times with
DNFB. The tIgG level increased twice after the application of DNFB.
tIgM level was similar between vehicle and DNFB groups. The levels of
sIgE and tIgE but not IgG and IgM in the serum were potentiated by
FK-506 and cyclosporin A (fig. 5). To
examine the effect of these drugs on the IgE production in the B cells,
we investigated the expression of germline C and productive C
mRNAs in the cervical lymph node. As shown in figure
6, the expression of germline C and
productive C mRNAs in the cervical lymph node was not affected by
FK-506 and cyclosporin A.
|
|
|
Effect of FK-506 and cyclosporin A on the production of cytokines by Th1 and Th2 cell clones in vitro
After stimulation with PMA, calcium ionophore A23187 and concanavalin A, IFN- (average 150 ng/ml) and IL-2 (2 ng/ml) were produced from 3 ×105 1E10.H2 cells, whereas
IL-4 (150 ng/ml) and IL-5 (5 ng/ml) were produced from 3 ×105 D10 G4.1 cells. As shown in table
1, FK-506 and cyclosporin A inhibited the
production of IFN- and IL-2 by Th1 cells and IL-4 and IL-5 by Th2
cells in vitro. Two drugs showed more potent inhibition to
the production of Th1 cytokines when compared to that of Th2 cytokines.
|
| |
Discussion |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
In our study, we demonstrated that the novel immunosuppressors,
FK-506 and cyclosporin A, inhibited contact dermatitis and potentiated
IgE antibody formation caused by contact sensitization with DNFB in
mice. Our results also indicate that two drugs suppress the expression
of IFN- mRNA in the skin lesion. Because the expression of IL-4,
germline C and productive C mRNAs in the lymph node was not
affected by the administration of the two drugs, they did not affect
IL-4-induced IgE class switching. Contrary to above in vivo
experiments, in vitro experiments indicated that cytokine production is inhibited in both cultured Th1 and Th2 cells. Two immunosuppressors, therefore, showed different action on the function of Th2 cells in vitro and in vivo.
The immune system is regulated by various feedback mechanisms that
control the magnitude, type and specificity of the immunological reactions. In the present study, the production of IgE was
significantly enhanced by FK-506 and cyclosporin A. These drugs did not
affect the simultaneous production of IgG and IgM antibodies. Because the expression of IFN- and contact dermatitis are suppressed by
FK-506 and cyclosporin A, these data suggest the inhibition of mainly
Th1 cells activity in the ears. The suppressed IFN- mRNA expression
may also affect the production of IgE antibody. Many investigators
suggest an important role of the immunological balance of IL-4 and
IFN- in IgE production in mice and human (Parronchi et al.,
1992; Finkelman et al., 1988; Pene et al., 1988; Lack et al., 1994; Coffman and Carty, 1986). The
expression of IL-4, germline C and productive C mRNAs; however,
were not affected by the administration of these two drugs. These data suggest that Th2 activity and B cell function in IgE antibody production were not affected by FK-506 and cyclosporin A. Because our
RT-PCR experiments were semiquantitative, further quantitation will be
necessary to reach a conclusion. At least, because FK-506 and
cyclosporin A inhibited the expression of IFN- mRNA, the potentiating mechanism of IgE production may relate to the suppression of Th1 activity. Some clinical studies suggest that the down-regulation of IFN- in patients with bronchial asthma resulted in a high IgE
serum level without affecting IL-4 level (Mukoyama et al., 1995; Kou et al., 1994). The precise mechanism of high IgE
production by the down-regulation of IFN-, remains still obscure.
Further experiments will be necessary to elucidate the relationship
between the potentiation of the IgE production and down-regulation of IFN- production in vivo not in vitro.
Moreover, recently, Yamashita et al. (1996) reported that
FK-506 inhibited primary IgE production but not secondary IgE
production in mice. They pointed out the role of non-T cell-derived
IL-4 in the secondary IgE production. In our experiments, FK-506 and
cyclosporin A were administered immediately after the primary
immunization. The difference between their data and our results may be
based upon the different immunizing procedure, antigen and
administration timing. We are now examining the role of IL-4 in the
onset of this dermatitis and IgE production. The role of non-T
cell-derived IL-4 in the present system will be elucidated in the near
feature. In addition, some recent investigations indicated that IgE
antibody response can be introduced by the IL-4 independent mechanism
(Morawetz et al., 1996, Smiley et al., 1997).
These evidence suggest the possibility of another mechanism of IFN-
on the suppression of IgE response without affecting IL-4. Further
experiments will be necessary to elucidate the role of IFN- in IL-4
independent IgE antibody response.
Contrary to the above in vivo results, FK-506 and
cyclosporin A suppressed the production of cytokines in Th1 or Th2
cells. The activation of STAT families and transcription factors such as Fyn and Lck in Th1 and Th2 cells, may participate on the activation of T cells in a different mechanism (Tamura et al ., 1993;
Gajewski et al., 1990). However, the activation of
calcineurin is the common pathway to activate each subset of T cells
(Lin et al., 1991). Our results confirmed the inhibitory
effect of FK-506 or cyclosporin A on the activation of both T1 and Th2
cells in vitro. These drugs tended to favor Th1 over Th2
cells. Similar results regarding selective inhibition of Th1 cells by
these two immunosuppressors in vitro were reported by Naora
and Young (1994), Schmidt et al. (1994), Van Wauwe et
al. (1995) and McHugh et al. (1995). These results,
taken together with our in vitro and in vivo data
indicate that FK-506 and cyclosporin A have a selective inhibitory
action on Th1 cells, especially in vivo.
FK-506 and cyclosporin A are used to prevent or treat organ allograft
rejection. These two drugs are also effective for treating patients
with chronic severe allergic diseases such as bronchial asthma and
atopic dermatitis (Sepp and Fritsch, 1993; Alexander et al.,
1992). IgE antibody may play an important role in the development of
these conditions. It is important, therefore, to monitor the serum
level of IgE antibody during drug therapy.
As for IgE antibody production by repeated application of DNFB on the
mice skin, some investigators reported a failure of the elevation of
serum IgE level by topical application of DNFB (Dearman and Kimber,
1991, Dearman et al. 1996). The reasons of these
inconsistencies may be resulted from the difference of experimental conditions including animal strain, the sensitivity to identify the
level of serum IgE and other experimental protocols.
In conclusion, these results suggest that five exposures to the hapten caused contact dermatitis in the skin and IgE production in the serum. RT-PCR studies indicated that contact dermatitis was induced by the activation of Th1 cells and that IgE production was initiated by Th2 cell activation. The novel immunosuppressants, FK-506 and cyclosporin A, inhibited the Th1 cell-dependent reaction but enhanced the Th2 cell-dependent reaction in vivo. On the contrary, these two agents inhibited the activation of Th1 and Th2 cells in vitro. This evidence suggests that FK-506 and cyclosporin A potentiate IgE antibody production in vivo due to a relatively selective inhibitory action upon Th1 cells.
| |
Footnotes |
|---|
Accepted for publication June 24, 1997.
Received for publication February 5, 1997.
Send reprint requests to: Dr. H. Nagai, Department of Pharmacology, Gifu Pharmaceutical University, 5-6-1 Mitahorahigashi, Gifu 502, Japan.
| |
Abbreviations |
|---|
DNFB, dinitrofluorobenzene; DNP, dinitrophenol; NF-AT, nuclear factor of activated T cells; IL, interleukin; IFN, interferon; Th, T helper; BSA, bovine serum albumin; ELISA, enzyme-linked immunosorbent assay; T-PBS, phosphate buffered saline containing 0.2% Tween; RT-PCR, reverse transcriptase-polymerase chain reaction; PMA, phorbol myristate acetate, CMC, carboxymethylcellulose.
| |
References |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
a unifying hypothesis of cellular mechanisms of cyclosporin A.
Transplant. Proc.
20: 92-102, 1988[Medline]..
J. Immunol.
136: 949-959, 1986[Abstract]. regulates the isotype of Ig secreted during in vivo humoral immune responses.
J. Immunol.
140: 1022-1030, 1988[Abstract]. production by peripheral blood lymphocytes from patients with bronchial asthma.
Jpn. J. Allerg.
43: 482-491, 1994. decreases IgE production and normalizes airway function in a murine model of allergen sensitization.
J. Immunol.
152: 2546-2554, 1994[Abstract]. by peripheral mononuclear cells in asthmatic children.
Jpn. J. Allerg.
44: 77-85, 1995. and ) exert opposite regulatory effects on the development of cytolytic potential by Th1 and Th2 human T cell clones.
J. Immunol.
149: 2977-2983, 1992[Abstract]., and prostaglandin E2.
Proc. Natl. Acad. Sci. U.S.A.
85: 6880-6884, 1988This article has been cited by other articles:
|
|
 |
|
  R. Marsella Calcineurin Inhibitors: A Novel Approach to Canine Atopic Dermatitis J. Am. Anim. Hosp. Assoc., March 1, 2005; 41(2): 92 - 97. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. S. Exton, A. Elfers, W.-Y. Jeong, D. F. Bull, J. Westermann, and M. Schedlowski Conditioned suppression of contact sensitivity is independent of sympathetic splenic innervation Am J Physiol Regulatory Integrative Comp Physiol, October 1, 2000; 279(4): R1310 - R1315. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Nagai, Y. Ueda, H. Tanaka, Y. Hirano, N. Nakamura, N. Inagaki, K. Takatsu, and K. Kawada Effect of Overproduction of Interleukin 5 on Dinitrofluorobenzene-Induced Allergic Cutaneous Response in Mice J. Pharmacol. Exp. Ther., January 1, 1999; 288(1): 43 - 50. [Abstract] [Full Text]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
 |
 |
 |
|
 |
 |
 |
