Eur Arch Otorhinolaryngol (2011) 268:73–81
DOI 10.1007/s00405-010-1282-3
RHINOLOGY
Nuclear factor kappa B pathway down-regulates aquaporin 5
in the nasal mucosa of rats with allergic rhinitis
Weiwei Wang • Ming Zheng
Received: 24 November 2009 / Accepted: 6 May 2010 / Published online: 21 May 2010
Ó Springer-Verlag 2010
Abstract Nuclear factor kappa B (NF-jB) induces gene
transcription by binding CREB-binding protein (CBP). The
aim of the study was to detect the mechanisms by which
NF-jB pathway regulated aquaporin 5 (AQP5) in the nasal
mucosa of rats with allergic rhinitis (AR). Rats were
divided into control (group C), model (group M), low-dose
proline dithiocarbamate (PDTC) (group L) and high-dose
PDTC (group H) groups. AR model was established by the
sensitization with ovalbumin, then groups L and H were
treated with PDTC (50 or 100 mg/kg/day) for 5 days.
AQP5, interleukin-1b, NF-jBp65 and phosphorylated
cAMP-response element binding protein (p-CREB) were
detected by immunohistochemistry, Western blotting or
real-time PCR. AQP5 expression in group M was lower
than in group C, but in groups L and H it increased.
NF-jBp65 expression in group M was higher than group C,
but in groups L and H it reduced. p-CREB expression in
group M was lower than group C, but in groups L and H it
increased. Interleukin-1b gene level in group M was higher
than group C, but in groups L and H it was lower. These
data show that the NF-jB pathway could down-regulate
AQP5 by interleukin-1b which inhibited CREB phos-
phorylation or by NF-jBp65 which competitively bound
CBP.
Keywords Allergic rhinitis  Aquaporin 5 
Nuclear factor kappa B  Interleukin-1b 
cAMP-response element binding protein
W. Wang  M. Zheng (&)
Fujian Medical University, No. 88, Jiaotong Road,
Fuzhou 350004, Fujian, China
e-mail: zhengmingfujian@163.com
Introduction
Allergic rhinitis (AR) results from an IgE-mediated allergy
associated with nasal inflammation of variable intensity.
Over the past 20 years, it has become a global health
problem with an incidence of about 25–35%, even 44.2%
in some areas [1–3]. AR seriously affects the patients’
quality of life.
Aquaporins (AQPs) play an important role in maintain-
ing the fluid balance of the airway [4, 5]. AQPs are also
associated with water metabolism disorders in AR. AQP5 is
one member of the aquaporin family, and it is closely
related to secretion of serous glands [6]. Information
regarding the regulation of abundance and distribution of
AQP5 is relatively limited although current studies suggest
that the cAMP-PKA pathway regulates AQP5. Yang [7]
treated mouse lung epithelial cell lines with the cpt-cAMP,
and found that AQP5 mRNA and protein level increased
fourfold, and treatment with H-89, a protein kinase A
inhibitor, inhibited these changes. Sidhaye [8] observed that
exposure of lung epithelial cells to cAMP, or the cAMP
inducing drugs terbutaline and forskolin, led to increased
expression of AQP5 in the apical plasma membrane. These
effects were blocked by inhibition of PKA activation.
Parvin [9] cultivated duodenal slices with vasoactive
intestinal polypeptide (VIP) in vitro, and found the amount
of AQP5 significantly increased in the apical membrane,
and the addition of H-89 blocked this increased expression.
NF-jB is a protein with polytranscriptional regulatory
roles. It plays a vital role in the initiation and perpetuation
of allergic inflammation [10, 11]. In this study, we estab-
lished an AR model which was then treated with PDTC, a
NF-jB inhibitor, in order to determine the role of the NF-jB
pathway in the regulation of AQP5 in the nasal mucosa of
rat with AR. Description of this pathway could provide
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theoretical basis to study the water metabolism disorders of
AR.
Materials and methods
Tissue preparation
This study was done in accordance with the accepted
policy on the use of animals and all the procedures were
approved by the governmental animal care authorities. 120
SD rats (weight range 250–300 g) were randomly divided
into groups: control group (group C), model group (group
M), and PDTC-treated low-dose group (group L), and
PDTC-treated high-dose group (group H). Groups L and H
were, respectively, subdivided into groups L1d, L3d and
L5d and groups H1d, H3d and H5d to reflect the days of
treatment. Each group consisted of 15 rats.
The establishment of AR model followed a published
protocol [12]. Each rat of groups M, L and H was sensi-
tized by intraperitoneal injection of 1 ml saline containing
30 mg of AL (OH)3 and 0.3 mg of ovalbumin (Sigma
USA) once every other day for 14 days, then was sensi-
tized by dripping 20 ll of 10% ovalbumin into the bilateral
nasal cavities from the 15th to the 21st day once a day,
while group C was treated with normal saline without
ovalbumin.
After the establishment of AR, groups C and M were
killed by pentobarbitone (100 mg/kg). Groups L and H
were, respectively, injected intraperitoneally with 50 or
100 mg/kg/day of PDTC (Sigma, USA) for 5 days. Groups
L1d and H1d, groups L3d and H3d and groups L5d and
H5d were, respectively, killed by pentobarbitone (100 mg/kg)
on the first, third or fifth day. The nasal mucosa tissues of
the rats were collected and one part was stored at -80°C
and the others were fixed in 4% paraformaldehyde solution
at pH 7.4 over night at 4°C, then embedded in paraffin.
Immunohistochemistry
Paraffin-embedded nasal mucosa was sectioned at 5 lm.
Sections were mounted on poly-L-lysine-coated slides,
deparaffinized, then rehydrated through gradient ethanol.
Intrinsic peroxidase activity was blocked with H2O2 for
10 min at room temperature. The slices were block with
non-immune serum for 10 min, then sections were incu-
bated overnight at 4°C with a 1:250 dilution of goat anti-
AQP5 antibody (Santa Cruz, USA) or a 1:300 dilution of
rabbit anti-p-CREB antibody (Abcam, USA). Sections
were incubated for 10 min at room temperature with the
corresponding biotin-conjugated secondary antibody fol-
lowed by incubation with HRP-streptavidin for 10 min,
then were incubated with diaminobenzidine for 1 min, and
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Eur Arch Otorhinolaryngol (2011) 268:73–81
were stained with hematoxylin. The immunoreaction was
assessed with the Imagepro-Plus 12.0 which had been
programmed to measure the mean optical density (MOD)
of the positive areas.
Western blotting
Nucleoprotein and cytoplasmic protein were isolated from
mucosa samples for the detection of p-CREB (Ser133) and
NF-jB p65 protein using a Nuclear-Cytosol Extraction
Reagent Kit (Pierce USA). Total protein from mucosa
samples was isolated with lysis buffer (Beyond, China) to
detect AQP5. After denaturation, 100 lg of each analyzed
sample was prepared for 12% SDS gel electrophoresis.
Proteins were transferred after electrophoresis to PVDF
membranes (Millipore, USA). The membranes were
blocked with blocking buffer (Beyond, China) for 3 h at
room temperature, then were incubated overnight at 4°C
with a 1:200 dilution of goat anti-AQP5 antibody, a 1:250
dilution of rabbit anti-p-CREB antibody, a 1:500 dilution
of mouse anti-NF-jB p65 antibody, a 1:1,000 dilution of
mouse anti-b-actin antibody or a 1:300 dilution of anti-
histone H2A-1 antibody (Abcam, USA). The membranes
were then incubated for 1 h with the corresponding HRP-
conjugated secondary antibody at room temperature, and
antibody binding was detected with Beyond ECL Plus
(Beyond, China). The bands in the Western blot were
analyzed using the digitalized scientific software program
Quantity One (Silk Scientific Corporation, Orem, UT,
USA).
Real-time PCR
The total RNA from the nasal mucosa was isolated by
TRIzol Reagent (Invitrogen USA). 1 lg of total RNA was
reverse transcribed to cDNA with ReverAidTM First Strand
cDNA Synthesis Kit (Fermentas USA). Real-time PCR was
performed using PlatinumÒ SYBRÒ Green qPCR Super-
Mix-UDG kits (Invitrogen, USA) and a real-time thermal
cycler (ABI 7500, USA). The primers are as follows:
b-actin: 50 -accgtgaaaagatgacccagat-30 , 50 -agctgtggtggtgaa
gctgtag-30 ; AQP5: 50 -catcttctcctccaccgactct-30 , 50 -gggtgcttc
aaactcttcgtct-30 ; IL-1b: 50 -ctccacctcaatggacagaaca-30 , 50 -tg
cagccatctttaggaagaca-30 . The amplification reaction was
50°C for 2 min and 95°C for 2 min, which was followed by
40 cycles of 95°C for 15 s and 60°C for 1 min, and ter-
minated by a cooling step at 4°C. The mean CT derived
from group C was used as the calibrator. The equation used
was:
Ratio ðsample to calibratorÞ ¼ 2ðDDCTÞ ;
where DCT = CT (goal gene) - CT (reference gene), and
DDCT = DCT (sample) - DCT (calibrator).
Eur Arch Otorhinolaryngol (2011) 268:73–81
Statistical methods
All values were expressed as the mean ± SD. The data
were analyzed by a one-way ANOVA and a two-tailed
Student’s t test. A P value of \0.05 was considered
significant.
Results
Immunohistochemistry results show AQP5 expressed in
the cell membrane of the glandular epithelial cells and the
pseudostratified ciliated columnar epithelial cells from
Fig. 1. As seen in Fig. 2, the MOD of the immunohisto-
chemically positive areas of AQP5 in group C was 87.73%
greater than group M (P \ 0.01). There was no difference
between two groups although the AQP5 expression in
group L1d was elevated by 3.87% compared with group M
(P [ 0.05), while expression in groups L3d and L5d
increased by 23.57 and 34.31%, respectively (P \ 0.05).
The AQP5 expression in group H1d increased by 19.91%
compared with group M (P \ 0.05), and expression in
groups H3d and H5d was significantly increased 41.60 and
Fig. 1 The location and positive distribution of AQP5 in the nasal mucosa by immunohistochemistry. a–g or h Groups C, M, L1d, L3d, L5d,
H1d, H3d or H5d, respectively. Black arrows The positive expression of AQP5 in nasal mucosa (scale bars 200 lm)
Fig. 2 The MOD of the AQP5
immunohistochemically
positive area in nasal mucosa
*P \ 0.05 versus group M.
**P \ 0.01 versus group M.
#
P [ 0.05 versus group M
(mean ± SD n = 5)
75
68.72% (P \ 0.01). The AQP5 protein expression
increased in a time-dependent and dose-dependent manner
with the treatment with PDTC.
It can be seen that p-CREB is expressed in the nucleus
from Fig. 3. As seen in Fig. 4, the MOD of the immuno-
histochemically positive areas of p-CREB in group C is
131.87% higher compared with group M (P \ 0.01). The
MOD of the immunohistochemically positive areas of
p-CREB in group L1d increased by 30.67% compared with
group M (P \ 0.05), and groups L3d and L5d increased by
60.84 and 106.76%, respectively (P \ 0.01). The MOD of
p-CREB in groups H1d, H3d and H5d increased by 55.65,
68.34 and 116.31%, respectively, compared with group M
(P \ 0.01). The p-CREB protein increased in a time-
dependent and dose-dependent manner with the treatment
with PDTC.
As seen in Fig. 5, NF-jBp65 protein in group C was not
much in nucleus fraction of cells from the nasal mucosa so
that it could not be detected by Western blotting. As seen in
Fig. 6, significantly more NF-jBp65 was detected in group
M than that in group C (P \ 0.01). The value of NF-jBp65
was reduced in groups L1d, L3d and L5d by 15.56, 22.89
and 25.96%, respectively, compared with group M
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Fig. 3 The location and positive distribution of p-CREB (Ser133) in the nasal mucosa by immunohistochemistry. a–g or h Groups C, M, L1d,
L3d, L5d, H1d, H3d or H5d. Black arrows The positive expression of p-CREB in nasal mucosa (scale bars 200 lm)
Fig. 4 The MOD of the
p-CREB
immunohistochemically
positive area in nasal mucosa.
*P \ 0.05 versus group
M. **P \ 0.01 versus group
M (mean ± SD n = 5)
Fig. 5 Western blot analysis for NF-jBp65 protein in the nucleus.
Histone H2A-1 was used as loading control. Samples were analyzed
by immunoblotting with antibody against NF-jBp65 or histone
H2A-1
(P \ 0.05). Compared with group M, the value of
NF-jBp65 protein in groups H1d and H3d decreased by
19.89 and 26.37%, respectively (P \ 0.05), and expression
in group H5d was reduced by 40.72% (P \ 0.01).
Figure 7 shows NF-jBp65 protein in group C was not
much in cytoplasmic fraction of cells from the nasal
mucosa so that it could not be detected by Western blot-
ting. As seen in Fig. 8, we see a significant increase in the
expression of NF-jBp65 in group M compared with group
C (P \ 0.01). The expression of NF-jBp65 in groups L1d
and L3d decreased by 12.04 or 30.65%, respectively,
compared with group M (P \ 0.05), while the ratio
decrease by 41.95% in group L5d (P \ 0.01). NF-jB
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Eur Arch Otorhinolaryngol (2011) 268:73–81
expression in groups H1d and H3d decreased by 19.26 and
35.92%, respectively, compared with group M (P \ 0.05)
and expression in group H5d decreased by 56.96%
(P \ 0.01).
Figure 9 shows the p-CREB protein in the nasal mucosa,
which was detected by Western blotting. As seen from
Fig. 10, the expression of p-CREB (Ser133) in group C
was decreased by 66.77% compared with group M
(P \ 0.01). The expression of p-CREB (Ser133) in groups
L1d and L3d increased by 13.31 and 35.37%, respectively,
compared with group M (P \ 0.05), and expression in
group L5d increased by 47.3% (P \ 0.01). Compared with
group M, p-CREB (Ser 133) expression increased in group
H1d by 24.09% (P \ 0.01), and expression in groups H3d
and H5d increased by 44.92 and 63.62%, respectively
(P \ 0.01). We see the production of p-CREB (Ser133)
increased in a time-dependent and dose-dependent manner
with PDTC treatment of the AR rats.
Figure 11 shows the AQP5 protein in the nasal mucosa,
which was detected by Western blotting. From Fig. 12 we
see that the AQP5 level in group M decreased by 53.65%
compared with group C (P \ 0.01). No significant differ-
ence was observed between groups L1d and M, although the
Eur Arch Otorhinolaryngol (2011) 268:73–81 77

Fig. 6 The protein level of

NF-jBp65 in the nucleus by
Western blotting. The relative
ratio of the intensity of
NF-jBp65 bands in the nucleus
was normalized to the intensity
of histone H2A-1 bands.
*P \ 0.05 versus group M.
**P \ 0.01 versus group M
(mean ± SD n = 5)

(P \ 0.01). AQP5 gene expression increased in a time-

Fig. 7 Western blot analysis for NF-jBp65 protein in the cytoplasm.
b-actin was used as loading control. Samples were analyzed by
immunoblotting with antibody against NF-jBp65 or b-actin
AQP5 level in group L1d was elevated by 7.51% (P [
0.05), while group L3d increased by 32.34% (P \ 0.05) and
group L5d increased by 68.15% (P \ 0.01). AQP5
expression in group H1d was higher by 25.83% (P \ 0.05)
than that in group M (P \ 0.05). In groups H3d and H5d,
AQP5 expression increased by 71.01 and 90.77%, respec-
tively, compared with group M (P \ 0.01). The production
of AQP5 protein was up-regulated in a time-dependent and
dose-dependent manner with PDTC treatment of the AR
rats.
Figure 13 shows the amount of mRNA coding for AQP5
was detected by real-time PCR. From Fig. 14, AQP5 RNA
expression in group C was 154.84% higher than that in
group M (P \ 0.01). There was no difference between
groups L1d and M, although AQP5 mRNA expression in
group L1d increased by 3.68% (P [ 0.05). In groups L3d
and L5d, mRNA expression increased by 11.98 and
29.58%, respectively, compared with group M (P \ 0.05).
AQP5 mRNA expression in groups H1d and H3d increased
by 29.76 and 20.97%, respectively, compared with group
M (P \ 0.05), and in group H5d the increase was 47.46%
dependent and dose-manner with PDTC of AR rats.
Figure 15 shows the amount of mRNA coding for IL-1b
was detected by real-time PCR. As seen in Fig. 16, we can
see the mRNA of IL-1b in group M was 55.06 times that of
group C (P \ 0.01). The gene expression of IL-1b in
groups L1d and L3d decreased by 16.99 and 34.30%,
respectively (P \ 0.05) compared with group M, and
mRNA expression in group L5d decreased by 53.89%
(P \ 0.01). IL-1b mRNA expression in group H1d
decreased by 27.13% (P \ 0.05) compared to group M,
and mRNA expression in groups H3d and H5d decreased
by 44.31 and 63.41%, respectively (P \ 0.01). The level of
IL-1b mRNA decreased in a time-dependent and dose-
dependent manner with PDTC treatment of the AR rats.
Discussion
Since AQP1 was discovered, 13 mammalian AQPs have
been described that are distributed throughout the body
[13]. AQPs are a group of membrane transporter proteins
related to water transport. Most of them are selectively
distributed in the epithelial cells that are related to fluid
secretion or absorption and in endothelial cells with col-
laborative transcytosis which play an important role in fluid
transport and secretion of some glands [14, 15].
The main characteristic of AR is hyper-secretion of
glands. Song [6] stimulated mouse respiratory tract with

Fig. 8 The protein level of

NF-jBp65 in the cytoplasm by
Western blotting. The relative
ratio of the intensity of
NF-jBp65 bands in the
cytoplasm was normalized to
the intensity of b-actin bands.
*P \ 0.05 versus group M.
**P \ 0.01 versus group M
(mean ± SD n = 5)

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pilocarpine and found that the submucosal gland fluid
secretion rate in AQP5 null mice was 57% lower when
compared with wild type mice. They concluded that AQP5
was a key participant in the fluid secretion and a rate-
limiting barrier to the secretion of the submucosal serous
gland. The effective control of hypersecretion of glands
and understanding the mechanism of gland secretion are
the focus of the study of AR. In this study, the NF-jB
pathway was activated in AR rats, but the expression of
AQP5 was reduced, indicating that NF-jB pathway was
involved in the down-regulation of AQP5 in AR rats.
NF-jB pathway might down-regulate AQP5 by IL-1b
which inhibited CREB phosphorylation
In this experiment, NF-jBp65 protein in group C was so
little that it could not be detected by Western blotting, but a
significant amount of NF-jBp65 was seen in group M,
indicating that the NF-jB pathway was activated in AR. It
had been reported that the NF-jB pathway was activated in
AR, and the amount of NF-jBp65 in nucleus increased
[16, 17], which is consisted with our result.
NF-jB is an inducible cellular transcription factor that
regulates a wide variety of cellular genes including several
cytokines [18]. After the NF-jB pathway was activated, the
gene transcription of an array of inflammatory cytokines
including IL-1b, TNF-a is induced, which can reactivate
the NF-jB pathway as extracellular stimulation signals.
NF-jB plays an important role in the regulation of AR
cytokine networks [19, 20].
Fig. 9 Western blot analysis for p-CREB (Ser133) protein in the
nasal mucosa. Histone H2A-1 was used as loading control. Samples
were analyzed by immunoblotting with antibody against p-CREB
(Ser133) or histone H2A-1
Fig. 10 The protein level of
p-CREB (Ser133) in the nucleus
by Western blotting. The
relative ratio of the intensity of
NF-jBp65 bands in the nucleus
was normalized to the intensity
of histone H2A-1 bands.
*P \ 0.05 versus group M.
**P \ 0.01 versus group M
(mean ± SD n = 5)
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Eur Arch Otorhinolaryngol (2011) 268:73–81
In our result, the mRNA of IL-1b in group M was
upregulated, but the synthesis of p-CREB (Ser133) and
AQP5 in group M was down-regulated. After treatment
with PDTC, the NF-jB pathway was inhibited, the pro-
duction of NF-jBp65 and IL-1b was down-regulated, but
the synthesis of p-CREB (Ser133) and AQP5 was upreg-
ulated. This indicated that the NF-jB pathway inhibited the
biosynthesis of p-CREB and AQP5. Jambal [21] found
that p-CREB (Ser133) decreased when mouse pancreatic
b-cell lines were exposed to IL-1b and genes regu-
lated by the cAMP-PKA pathway were down-regulated.
Lahiri [22] studied the effects of IL-1b on CRE-dependent
gene expression in human bronchial smooth muscle
cells, finding that IL-1b could suppress cAMP-response
element-dependent gene expression. As mentioned above,
cAMP-PKA pathway regulated AQP5. PKA induced gene
expression by phosphorylating its major target protein
CREB at Ser133 in the nucleus [23]. The AQP5 50 -flanking
region contains consensus binding sites for CREB [24].
The cAMP-PKA pathway could regulate the expression of
AQP5 by CREB phosphorylation. The NF-jB pathway was
activated in AR, which up-regulated IL-1b mRNA and the
production of IL-1b. We speculate that the NF-jB pathway
might down-regulate AQP5 by IL-1b which inhibited
CREB phosphorylation at Ser133.
NF-jB pathway might down-regulate AQP5 by
competitive binding to CREB-binding protein (CBP)
Under physical circumstances, NF-jB is present as a het-
erodimer comprising a 50 and a 65 kDa subunit, and is
normally sequestered in the cytoplasm through being
associated with an IjB (inhibitor of NF-jB) protein in an
inactive state [25]. Upon stimulation of cells with various
cytokines, IjB proteins are phosphorylated at specific
serine residues, poly-ubiquitinated, then degraded. Freed
from IjB, the active NF-jB is able to translocate to the
nucleus, where it induces gene transcription by binding
transcription co-factor CBP in a selective manner [26, 27].
CBP plays an important role in gene transcription or
Eur Arch Otorhinolaryngol (2011) 268:73–81 79

regulation by binding to specific NF-jBp65 binding sites of
corresponding genes [28].
In this study, much NF-jBp65 was detected in group
M by Western blotting. This indicated that plenty of
Fig. 11 Western blot analysis for AQP5 protein in the nasal mucosa.
b-actin was used as loading control. Samples were analyzed by
immunoblotting with antibody against AQP5 or b-actin
NF-jBp65 went into the nucleus from the cytoplasm to
induce gene transcription after the NF-jB pathway was
activated. At the same time, the expression of AQP5 or
p-CREB in group M was lower than that in group C. After
treatment with PDTC, there was significant up-regulation
of AQP5 and p-CREB (Ser133). This response might be
related to CBP. As an important transcription co-factor,
CBP is a protein that can specifically bind CREB at serine
133, after phosphorylation of serine 133, promoting gene
transcription [23] and participate in a number of signal
transduction pathways [29]. Many transcription factors
with inducing activity are regulated by CBP, including
NF-jB and CREB [30]. These transcription factors do not
have transcription activity until CBP combines with the

Fig. 12 The protein level of

AQP5 in nasal mucosa by
Western blotting. The relative
ratio of the intensity of AQP5
bands in the nucleus was
normalized to the intensity of
b-actin bands. *P \ 0.05 versus
group M. **P \ 0.01 versus
group M. #P [ 0.05 versus
group M (mean ± SD n = 5)

Fig. 13 The real-time

polymerase chain reaction
comparative analysis of AQP5
in the nasal mucosa. Curves
corresponded to AQP5
amplification from every group.
The x-axis represented the
number of cycles, and the y-axis
represented normalized
fluorescence

Fig. 14 The mRNA level of

AQP5 in the nasal mucosa by
real-time PCR. *P \ 0.05
versus group M. **P \ 0.01
versus group M. #P [ 0.05
versus group M (mean ± SD
n = 5)

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Fig. 15 The real-time
polymerase chain reaction
comparative analysis of IL-1b
in the nasal mucosa. Curves
corresponded to IL-1b
amplification from every group.
The x-axis represented the
number of cycles, and the y-axis
represented normalized
fluorescence
Fig. 16 The mRNA level of IL-1b in the nasal mucosa by real-time
PCR. *P \ 0.05 versus group M. **P \ 0.01 versus group M
(mean ± SD n = 5)
promoter region of the gene. Interaction with CBP appears
to be necessary to optimize the transcriptional activity of
NF-jB [18]. CBP can also combine with other transcrip-
tion factors, such as CREB, besides NF-jBp65. The
interaction of NF-jBp65 with CBP involves the KIX
region of CBP, which is the same region responsible for
binding the transcriptionally active serine-133-phosphory-
lated form of CREB [31]. Because CBP is present in
limiting amounts in the nucleus, competition between
NF-jB and CREB for binding CBP has been postulated to
be important in regulating the transcriptional activity of
these factors [30]. That NF-jBp65 competitively binds
CBP with CREB can lead to the reduction in gene tran-
scription-dependent CREB [31].
We propose the following hypothesis, the activation of
the NF-jB pathway in AR is due to an increase in the
intracellular concentration of NF-jBp65. NF-jBp65 com-
petitively binds CBP, resulting in less available CBP to
combine with p-CREB so that AQP5 transcription, which
is dependent on the cAMP-PKA-CREB pathway, is down-
regulated.
Conclusion
According to our findings in the present study, the NF-jB
pathway down-regulated the expression of AQP5 in rats
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Eur Arch Otorhinolaryngol (2011) 268:73–81
with AR by inhibiting CREB phosphorylation or by
NF-jBp65 binding CBP in competition with p-CREB,
resulting in hypersecretion during inflammation because of
the clearance dysfunction of the gland secretions.
Acknowledgments This study was carried out in the neurobio-
logical laboratory of Fujian Medical University, so we warmly thank
Prof. Weiwang, the director of the neurobiological Laboratory. This
study was approved by Fujian Medical University on Principles of
Laboratory Animal Care and was performed in accordance with the
German Law on protection of Animals.
Conflict of interest statement The authors do not have any pos-
sible conflicts of interest.
References
1. Cagnani CE, Solé D, Dı́az SN et al (2009) Allergic rhinitis update
and its impact on asthma (ARIA 2008). Latin American per-
spective. Rev Alerg Mex 56:56–63
2. Weber RW (2008) Allergic rhinitis. Prim Care 35:1–10
3. Sakashita M, Hirota T, Harada M et al (2009) Prevalence of
Allergic Rhinitis and Sensitization to Common Aeroallergens in a
Japanese Population. Int Arch Allergy Immunol 151:255–261
4. Song Y, Jayaraman S, Yang B, Matthay MA, Verkman AS (2001)
Role of aquaporin water channels in airway fluid transport,
humidification, and surface liquid hydration. J Gen Physiol
117:573–582
5. Boucher RC (1999) Molecular insights into the physiology of the
‘thin film’ of airway surface liquid. J Physiol 516:631–638
6. Song Y, Verkman AS (2001) Aquauaporin-5 dependent fluid
secretion in airway submucosal glands. J Biol Chem 276:41288–
41292
7. Yang F, Kawedia JD, Menon AG (2003) Cyclic AMP regulates
aquaporin 5 expression at both transcriptional and post-tran-
scriptional levels through a protein kinase A pathway. J Biol
Chem 278:32173–32180
8. Sidhaye V, Hoffert JD, King LS (2005) cAMP has distinct acute
and chronic effects on aquaporin-5 in lung epithelial cells. J Biol
Chem 280:3590–3596
9. Parvin MN, Kurabuchi S, Murdiastuti K et al (2005) Subcellular
redistribution of AQP5 by vasoactive intestinal polypeptide in the
Brunner’s gland of the rat duodenum. Am J Physiol Gastrointest
Liver Physiol 288:G1283–G1291
10. Wan F, Lenardo MJ (2010) The nuclear signaling of NF-kappaB:
current knowledge, new insights, and future perspectives. Cell
Res 20:24–33
Eur Arch Otorhinolaryngol (2011) 268:73–81
11. Kumar A, Takada Y, Boriek AM, Aggarwal BB (2004) Nuclear
factor-kappaB: its role in health and disease. J Mol Med
82:434–448
12. Fan EZ, Xi L, Han DM, Zhang SZ, Li Y, Zhang L (2009)
Evaluation of the safety of aluminium adjuvant in the preparation
of allergic rhinitis animal model. Zhonghua Er Bi Yan Hou Tou
Jing Wai Ke Za Zhi 44:664–668
13. Ishibashi K, Hara S, Kondo S (2009) Aquaporin water channels
in mammals. Clin Exp Nephrol 13:107–117
14. King LS, Yasui M (2002) Aquaporins and disease: lessons from
mice to humans. Trends Endocrinol Metab 13:355–360
15. Connolly DL, Shanahan CM, Weissberg PL (1998) The aqu-
aporin. A family of water channel proteins. Int J Biochem Cell
Biol 30:169–172
16. Li J, Luo L, Wang X, Liao B, Li G (2009) Inhibition of
NF-kappaB expression and allergen-induced airway inflamma-
tion in a mouse allergic asthma model by andrographolide. Cell
Mol Immunol 6:381–385
17. Zhou LF, Zhu Y, Cui XF, Xie WP, Hu AH, Yin KSn (2006)
Arsenic trioxide, a potent inhibitor of NF-jB, abrogates allergen-
induced airway hyperresponsiveness and inflammation. Respir
Res 7:146
18. Sheppard KA, Rose DW, Haque ZK et al (1999) Transcriptional
activation by NF-kappaB requires multiple coactivators. Mol Cell
Biol 19:6367–6378
19. Xu R, Xu G, Shi J, Wen W (2007) A correlative study of
NF-kappaB activity and cytokines expression in human chronic
nasal sinusitis. J Laryngol Otol 121:644–649
20. Osada R, Takeno S, Hirakawa K, Ueda T, Furukido K, Yajin K
(2003) Expression and localization of nuclear factor-kappa sub-
units in cultured human paranasal sinus mucosal cells. Rhinology
41:80–86
81
21. Jambal P, Masterson S, Nesterova A et al (2003) Cytokine-
mediated down-regulation of the transcription factor cAMP-
response element-binding protein in pancreatic beta-cells. J Biol
Chem 278:23055–23065
22. Lahiri T, Moore PE, Baraldo S et al (2002) Effect of IL-1 on
CRE-dependent gene expression in human airway smooth muscle
cells. Am J Physiol Lung Cell Mol Physiol 283:L1239–L1246
23. Parker D, Ferreri K, Nakajima T et al (1996) Phosphorylation of
CREB at Ser-133 induces complex formation with CREB-bind-
ing protein via a direct mechanism. Mol Cell Biol 16:694–703
24. Krane CM, Towne JE, Menon AG (1999) Cloning and charac-
terization of murine Aqp5: evidence for a conserved aquaporin
gene cluster. Mamm Genome 10:498–505
25. Vallabhapurapu S, Karin M (2009) Regulation and function of
NF-kappaB transcription factors in the immune system. Annu
Rev Immunol 27:693–733
26. Haglund K, Dikic I (2005) Ubiquitylation and cell signaling.
EMBO J 24:3353–3359
27. Hayden MS, Ghosh S (2004) Signaling to NF-kappaB. Genes
Dev 18:2195–2224
28. Abraham E (2000) NF-kappaB activation. Crit Care Med
28:N100–N104
29. Chan HM, La Thangue NB (2001) p300/CBP proteins: HATs for
transcriptional bridges and scaffolds. J Cell Sci 114:2363–2373
30. Shenkar R, Yum HK, Arcaroli J, Kupfner J, Abraham E (2001)
Interactions between CBP, NF-kappaB, and CREB in the lungs
after hemorrhage and endotoxemia. Am J Physiol Lung Cell Mol
Physiol 281:L418–L426
31. Zhong H, Voll RE, Ghosh S (1998) Phosphorylation of NF-kappa
B p65 by PKA stimulates transcriptional activity by promoting a
novel bivalent interaction with the coactivator CBP/p300. Mol
Cell 1:661–671
123