Extract
Note: Please read the complete
full text with Figures and Tables at
Introduction
Water transport across epithelial and endothelial
barriers in bronchopulmonary tissues occurs during airway
hydration, alveolar fluid transport, and submucosal gland
secretion. Many of the tissues involved in these processes
are highly water permeable and express aquaporin (AQP)
water channels[1]. AQP1 is expressed in microvascular
endothelia throughout the lung and airways,
AQP3 in basal epithelial cells in large airways and throughout the
nasopharynx, AQP4 at the basolateral plasma membrane
surface in epithelia throughout the trachea and the small and
large airways, and AQP5 in type I alveolar epithelial cells and
submucosal gland acinar cells[2]. In the lungs,
AQP1 and AQP5 provide the principal route for osmotically-driven
water transport[3]. AQP5 deletion in submucosal glands in
the upper airways reduced fluid secretion and increased
protein content[4].
Chronic obstructive pulmonary disease (COPD) is a
major cause of chronic morbidity and mortality throughout the
world, and is usually complicated by mucus hypersecretion
in the airways leading to repeated
infection[5] and irrever-sible airflow
limitation[6]. Mucus consists mainly of water
(95%) combined with salts, lipids, proteins, and glycoprotein.
Abnormal mucus secretions have been proposed to promote
bacterial adhesion and inhibit bacterial clearance by
impeding cilia function[7], which may lead to exacerbations in COPD
patients. So far, studies about mucus are focused on the
regulation of mucin synthesis. However, the secretion of
mucin is intimately related to the secretion of water and ions
as well, for which little study has been done.
Thirteen mucin genes have been
discovered[8]. Mucin 5AC (MUC5AC), a secreted
mucin, colocalizes with glycoprotein-positive goblet cells and submucosal gland mucus
cells. Thus, MUC5AC is the marker of mucus production in
the epithelium airways[9]. In this study, we hypothesized
that an abnormal gene expression of AQP may exist and
become complicated by mucus hypersecretion in the airways
of COPD patients. To this end, we examined the expression
levels of AQP5, MUC5AC, and mucin in bronchial tissues of
Chinese COPD patients to investigate the relationship
of AQP5 and mucus overproduction.
Materials and methods
Bronchial tissues Bronchial tissues obtained from
fiberoptic bronchoscopy and bronchial biopsy in West China
Hospital (Sichuan, China) between April and July 2004 were
prepared as described[10]. Twenty-five patients were
diagnosed as COPD patients according to the Global Initiative
for Chronic Obstructive Lung Disease (GOLD) 2004
(http://www.goldcopd.com), and another 20 were the control
patients without COPD. Patients with asthma, cystic fibrosis,
bronchiectasis, and diffuse panbronchiolitis were excluded.
The COPD patients had history of productive coughing for
3 consecutive months each year for the past 2 years, with a
forced expiratory volume in 1 s (FEV1) <80% of the predictive
value, a FEV1/forced vital capacity (FVC)
ratio <70%, and a reversibility in
FEV1 <10% after inhalation
of 400 µg salbutamol. According to
FEV1% predicted value, there were 8 mild, 16 moderate, and 1 severe patient, respectively. The
control patients had foreign body in the bronchus or benign
bronchial stenosis and needed treatment with fiberoptic
bronchoscopy. Eight healthy volunteers were also included
in the control group (Table 1). All patients were ex-smokers
or non-smokers, and the ex-smokers had given up smoking
for more than 5 years. All patients had no respiratory tract
infections within 2 weeks preceding the operation. The sites
of biopsy were randomized to either the right or the left lower
lobe, and 5 to 6 endobronchial biopsy specimens were taken
from the tertiary carinae of the right or left lower lobes under
direct visualization. The ethics committee of the West China
Hospital, Sichuan University approved the study, and all
patients gave written informed consent.
RT-PCR To examine the AQP5 mRNA levels in the human
airways, semiquantitative RT-PCR was performed. Total RNA
was isolated with Trizol reagent in accordance with the
manufacturer's protocol (Invitrogen, Carlsbad, CA, USA).
The AQP5 primer sequences were as
follows: upstream, 5'-AAG GCC GTG TTC GCA GAG TT-3' and downstream,
5'-TGG TCA GCT CCA TGG TCT TC-3'. The MUC5AC primer
sequences were as follows[11]: upstream, 5'-TCC GGC CTC
ATC TTC TCC-3' and downstream, 5'-ACT TGG GCA CTG GTG CTG-3'. The GADPH primer sequences were as follows:
upstream, 5'-GAG CGA GAT CCC TCC AAA-3' and down-stream, 5'-ACT GTG GTC ATG AGT CCT TC-3'. The RT-PCR
reagents were purchased from Promega Corporation
(Madi-son, MI, USA). The reaction mixture contained 10 µL Avian
Myeloblastosis Virus (AMV)/Thermus flacus (Tfl) 5×reaction
buffer, 0.2 mmol/L deoxy-ribonucleoside
triphosphate (dNTP) mixture, 1 µmol/L of each primer, 1 mmol/L
MgSO4, 5U AMV reverse transcriptase, 5U Tfl DNA poly-merase, and 1 µg
RNA sample in 50 µL volume. The first-strand cDNA was
synthesized at 48 oC for 45 min, followed by denature of the
template at 94 oC for 2 min. Amplication was achieved with
40 cycles of denaturing (94 oC, 30 s), annealing
(57 oC, 1 min), and extension (68
oC, 2 min). The final extension was performed at
68 oC for 7 min. The PCR products were analysed
by 1.5% agarose gel electrophoresis, and the expressions
were measured by densitometry using Molecular Analysis
Software (BioRad, Hercules, CA, USA). Relative
quantita-tions of gene expressions were normalized to GAPDH with
each sample.
In situ hybridization The bronchial sections were fixed
with 4% neutral buffered formaldehyde in 0.1% diethyl
pyrocarbonate (DEPC). Six 4-µm sections were cut from each
bronchial tissue block. A digoxigenin-labeled 24 bp RNA
probe was designed by Shenergy Biocolor Company
(Shanghai, China). The AQP5 primer sequence was 5'-AGA
GCA GGT AGA AGT AAA GGA TGG CAG-3'. The probe was
diluted in 50 pmol/µL. ISH reagents were purchased from
Pan Path (Amsterdam, the Netherlands). Bound probes were
detected with alkaline phosphatase-conjugated anti-digoxin
(DIG) antibodies, and Nitroblue
tetrazolium/5-bromo-4-chloro-3-indolyl phosphate was used as substrate. The
sections were counterstained with methyl green.
Hybridizations of several sections with
AQP5 sense probes were used as negative controls.
The samples were dewaxed in xylene and dehydrated in a
graded ethanol series prehybridization. Tissues were
incubated for 15 min at 37 oC in 1 mg/mL proteinase K, and
then blocked for 10 min in 0.1 mol/L triethanolamine and 0.25%
acetic solution before equilibration in 2×standard saline
citrate (SSC). The sections were prehybridized with 50%
formamide in 2×SSC for 4 h at 50 oC. Hybridization was
carried out overnight at 42 oC and then the slides were washed
for 15 min in 4×SSC at 37 oC and exposed for 30 min to 10
µg/mL RNase in 2×SSC at 37 oC. After incubation for 30 min
with normal goat blood serum, alkaline
phosphatase-conjugated anti-DIG antibodies and Nitroblue
tetrazolium/5-bromo-4-chloro-3-indolyl phosphate were added as substrates. The
sections were counterstained with methyl green.
Histochemical analysis The bronchial sections were fixed
with 4% neutral buffered formaldehyde. Six 4-µm sections
were cut from each bronchial tissue block. Antihuman
AQP5 polyclonal antibody and MUC5AC polyclonal antibody were
purchased from Santa Cruz Biotechnology (Santa Cruz, CA,
USA) and diluted at 1:100. The sections were stained with
diaminobenzidine (DAB).
The samples were dewaxed in xylene and dehydrated in a
graded ethanol series. Endogenous peroxidase activity was
inhibited by incubating the slides with 3%
H2O2 for 30 min, followed by washing in phosphate-buffered solution (PBS)
and microwave heating (700 W) for 15 min in 10 mmol/L citric
acid (pH 6.0). The sections were inactivated with normal
goat serum for 20 min at 37 oC. After repeated washing with
PBS, the sections were incubated with the
AQP5 and MUC5AC antibodies, placed in a humid chamber at 4
oC overnight, washed in PBS, and incubated at 37
oC for 30 min with biotinylated rabbit antigoat immunoglobulin, followed by
repeated washing in PBS. The tissue was incubated with
Streptavidin Peroxidase (SP) reagents for 30 min at 37
oC and then the sections were stained with freshly prepared DAB.
After repeated washing with PBS and rinsing with tap water,
the slides were counterstained with hematoxylin, and alcian
blue-periodic acid-Schiff (AB-PAS) staining was performed
as usual.
Image analysis All the images were generated by Spot
Advanced Software (Diagnostic Instruments, MI, USA) and
analyzed by IMAGE-Pro plus Version 4.5 (Media Cybernetics,
HWY, USA). Two pathologists blinded to the patients'
clinical data reviewed all the images and made the analysis.
High-power fields of bronchial tissues were randomly sampled
and calculations were performed on 6 images per section
examined at ×200 magnification. The mean of the 6 sections
from each tissue was used for the statistical analysis. For
the in situ hybridization and immunohistochemistry, the
optical density (OD) of the positively-stained cells was
normalized to the OD of the background in the same sections.
For AB-PAS staining, the positively-stained areas in the
submucosal glands were used to evaluate proliferations of the
submucosal glands, normalized to areas of the bronchial
tissues in the same fields of view.
Statistical analysis All data were expressed as the mean±
SEM. Statistical analyses were performed using SPSS 12.0
software (SPSS Inc, IL, USA). Statistical significance was
analyzed with one-way ANOVA, followed by Student-Newman-Keuls test to isolate significant difference.
χ2-test and the Pearson correlation analysis were also used. A
P-value less than 0.05 (2-tailed test) was considered
statistically significant.
Results
AQP5 and MUC5AC mRNA expression in bronchial
tissues To investigate the expression of
AQP5 and MUC5AC mRNA, RT-PCR was performed. The signals of
APQ5 and MUC5AC were detected in the bronchial tissues of all the
patients. Bands of AQP5, MUC5AC, and GAPDH PCR
products were located at 756, 682, and 298 bp, respectively (Figure 1).
Compared with the control group, attenuated expressions of
AQP5 were detected in the mild, moderate, and severe COPD
groups (P<0.01, P<0.01). The mean OD of
AQP5 were 0.68± 0.01 in the control group, 0.62±0.03 in the mild COPD group,
and 0.56±0.04 in the moderate and severe COPD group
(Figure 1A). The upregulated expressions of MUC5AC were
also found in 2 COPD groups (P<0.01,
P<0.01). The mean OD of MUC5AC were 0.43±0.03 in the control group,
0.48±0.05 in the mild COPD group, and 0.54±0.04 in the moderate
and severe COPD group (Figure 1B). In the lung tissues, the
differences in AQP5 in the groups were not noted and there
was no MUC5AC expression in the lung tissues in all the
groups (data not shown).
The influence of smoking on the expression of
AQP5 and MUC5AC mRNA was also evaluated. There were 17
ex-smokers and 8 non-smokers in the COPD group, and 10
ex-smokers and 10 non-smokers in the control group. In the
COPD group, the mean OD of AQP5 were 0.58±0.03 for the
ex-smokers and 0.63±0.04 for the non-smokers, and the mean OD of
MUC5AC were 0.53±0.04 for the ex-smokers and 0.49±0.03
for the non-smokers. Both differences were significant
(P<0.05, P<0.05), but in the control group, the mean OD of
AQP5 was 0.67±0.03 for the ex-smokers and 0.68±0.04 for the
non-smokers, and the mean OD of MUC5AC was 0.45±0.04
for the ex-smokers and 0.43±0.03 for the non-smokers,
without significant differences
(P>0.05, P>0.05).
To identify the cell-specific expression of
AQP5, in situ hybridization was performed.
AQP5 expression was detected mainly in the submucosal gland cells, type I alveolar cells,
and secondarily in the airway epithelial cells from all groups
of patients. The positive signals (in purple) were expressed
mainly in the nucleus and secondarily on the cell membrane.
The COPD groups showed attenuated expressions and less
positively-stained cells of submucosal glands and epithelial
airway cells than the control group. The mean OD
was 1.37±0.05 in the control group, 1.35±0.02 in the mild COPD
patients, and 1.31±0.02 in the moderate and severe COPD
patients; the differences were significant
(P<0.01, P<0.01; (Figure 2A_2C). The differences of
AQP5 expression in type I alveolar cells of all groups were not significant
(P>0.05, P>0.05; data not shown). Hybridization with the sense probe
revealed no staining, confirming the specificity of the
assay (Figure 2D).
AQP5 and MUC5AC protein expression in bronchial
tissues To examine the correlation between
AQP5 expression and MUC5AC synthesis, immunohistochemical staining for
AQP5 and MUC5AC were performed.
AQP5 was expressed mainly in submucosal glands, type I alveolar cells, and
secondarily at the apical membranes of cellula
columno-epithelialis from patients in all the groups. The positive
signals (in brown) were expressed mainly in the cytoplasm and
secondarily on the cell membrane. The COPD groups showed
attenuated expressions of AQP5 and less positively-stained
cells of submucosal glands and epithelial airway cells than
the control group. The mean OD was 1.36±0.02 in the
control group, 1.34±0.01 in the mild COPD patients, and
1.30±0.02 in the moderate and severe COPD patients; the
differences were significant (P<0.01,
P<0.01; Figure 3A_3C). The differences of
AQP5 expression in type I alveolar cells of all
groups were not significant (P>0.05,
P>0.05; data not shown).
MUC5AC was expressed throughout the bronchial
tissues from patients in all groups, but no positive signals were
found in the lung tissue of any group. The positive signals
(in brown) were expressed mainly in the cytoplasm and
secondarily on the cell membrane. The COPD groups showed
stronger expression and more positively-stained cells than
the control group. The mean OD was 1.33±0.01 in the
control group, 1.37±0.01 in the mild COPD patients, and
1.41± 0.03 in moderate and severe COPD patients; the differences
were significant (P<0.01, P<0.01; Figure 4A_4C). When PBS
was used instead of the AQP5 and MUC5AC antibodies, no
staining resulted, confirming the specificity of the assay
(Figures 3D,4D).
The results of the immunohistochemical staining for
AQP5 and MUC5AC were used to evaluate the influence of
smoking. In the COPD group, the mean OD of
AQP5 was
1.31±0.02 for the ex-smokers and 1.34±0.02 for the
non-smokers, and the mean OD of MUC5AC was 1.42±0.02 for
the ex-smokers and 1.37±0.03 for the non-smokers. Both
differences were significant (P<0.05,
P<0.01), but in the control group, the mean OD of
AQP5 was 1.36±0.03 for the
ex-smokers and 1.35±0.02 for the non-smokers. The mean
OD of MUC5AC was 1.31±0.02 for the ex-smokers and
1.34±0.02 for the non-smokers, without significant differences
(P>0.05, P>0.05).
Mucin production in bronchial tissues To examine
whether the decreased expression of
AQP5 was associated with mucus hypersecretion, AB-PAS staining were performed.
The sialic acid in the submucosal gland acinar cells was
stained blue, while polysaccharide and neutral mucin were
stained purple. Submucosal glands in bronchial tissues from
all groups produce mucin. The ratios of the positively-stained
areas in the submucosal glands to the areas of bronchial
tissues in the same fields of view were 0.26±0.01 in the
control group, 0.31±0.02 in the mild COPD patients, and
0.38±0.04 in the moderate and severe COPD patients;
significant differences were noted
(P<0.01, P<0.01; Figure 5A_5C).
Smoking resulted in increased positively-stained areas
in the submucosal glands. In the COPD group, they were
0.37±0.02 for the ex-smokers and 0.32±0.02 for the
non-smokers, 0.25±0.01 for the ex-smokers and 0.26±0.01 for the
non-smokers in the control group. The difference in the
COPD group was significant (P<0.01), and that of the
control group was to the contrary (P>0.05).
Correlation analysis The downregulation of
AQP5
expression correlated significantly with mucus
overproduc-tion, the data of FEV1/FVC and
FEV1% predicted values,
V50% predicted value, and
V25% predicted value. The expression
levels of MUC5AC mRNA negatively correlated with the data
of FEV1/FVC and FEV1% predicted values,
V50% predicted value, and
V25% predicted value (Table 2).
Discussion
The discovery of the AQP family of water channel
proteins has provided insight into molecular mechanisms of
membrane water permeability. The specific expression
pattern of AQP5 in the lungs suggests that it plays a role in the
regulation of mucus secretion. In this study, we evaluated
the expression of AQP5, MUC5AC, and mucin in the airways
in bronchial tissues. Attenuated AQP5 mRNA expression
was demonstrated in the airways of COPD patients.
Con-comitantly, MUC5AC and mucus secretion increased greatly
in the airways of COPD patients. Smoking attenuated the
expressions of AQP5 and increased the staining of MUC5AC
and mucin in submucosal glands in the COPD groups, while
there were no significant differences in the control group.
The downregulation of AQP5 expression correlated
significantly with mucus overproduction, the data of
FEV1/FVC and FEV1% predicted values,
V50% predicted value, and
V25% predicted value. The expression levels of MUC5AC mRNA
negatively correlated with the data of
FEV1/FVC and FEV1% predicted values,
V50% predicted value, and
V25% predicted value. These results suggest that the attenuated gene
expression of AQP5 existed and was complicated by mucus
hypersecretion in the airways of Chinese COPD patients.
Smoking results in airway remodeling in patients with COPD.
Mucus protects the underlying airway epithelium from
dehydration, pathogens, and chemical and particulate
irritants, but mucus hypersecretion in the airways leads to
the obstruction of airflow. Abnormal mucus secretions have
been proposed to promote dysfunction of antibiosis enzymes
and antibacterial peptides[7]. Submucosal glands are thought
to play an important role in the regulation of mucus volume
and composition, and are important in antimicrobial
defense[12]. Immunocytochemistry of human and rat airways indicated
the expression of AQP water channels in glandular
epithelia[13,14]. In the current study, we found the expression of
AQP5 at the apical membranes of cellular columnoepithelialis and
submucosal glands, the sites of active mucus secretion. This
confirmed the importance of AQP5 in the regulation of
mucus secretion. AQP5 was also found expressed in the apical
surface of type I pneumocytes[15], which involved fluid
movement across epithelial and endothelial barriers occurring in
interstitial and alveolar pulmonary edema. Song
et al[16] showed that gland fluid from
AQP5-null mice had a substantially higher protein concentration and mildly elevated
chloride concentration compared with fluid from wild-type mice.
Ma et al[17] found that
AQP5 deletion resulted in impaired secretion of fluid from salivary glands. Furthermore, Krane
et al[18] showed that the decreased rate of fluid secretion
onto the airway surface in AQP5-null mice could increase
airway reactivity in response to bronchoconstricting agents.
We found for the first that the expression of
AQP5 was attenuated throughout the bronchial tissues from patients with
COPD. Moreover, the attenuated expression of
AQP5 was related to the severity of airflow obstruction. Thus,
AQP5 deletion may be one of the causes of hypertonic mucus and
airway hyperresponsiveness in patients with COPD.
Smoking is a main factor in the pathogenesis of COPD.
We evaluated the influence of smoking on the expressions
of AQP5, MUC5AC, and mucus in submucosal glands of
ex-smokers and non-smokers in the control and COPD groups,
respectively. Interestingly, smoking attenuated the
expressions of AQP5 and increased the staining of MUC5AC and
mucin in submucosal glands in the COPD group, while there
were no significant differences in the control group.
Although all ex-smokers had given up smoking for more than 5
years, the influence of smoking continued to exist in
patients with COPD. This might be the result of airway
remodeling in these patients, but it seemed that smoking did not
play any role in the control group. Only 15% of cigarette
smokers develop clinically-significant
COPD[19]. Whether AQP5 gene polymorphisms are involved in the pathogenesis
of COPD needs further investigation.
In COPD patients, the impaired hypertonic mucus by
AQP5 deletion may be a functional rather than anatomical defect as
we found that greater areas of submucosal glands were noted
in the COPD group than in the control group, and the
expression levels of AQP5 mRNA significantly correlated with the
patients' data of FEV1/FVC and
FEV1% predicted values,
V50%, predicted value, and
V25% predicted value. The
AQP5 deletion and MUC5AC increase may reflect, at least in part,
the airflow obstruction. Towne et
al[20] showed that tumor necrosis
factor-α (TNF-α) inhibited the expression of
AQP5 mRNA and protein in mouse lung epithelia cells. Recently,
Lora et al[21] demonstrated that
TNF-α triggered mucus production in the epithelium airways through an
IκB kinase β-dependent mechanism. So TNF-α may regulate mucus
secretion in 2 ways. In a cultured human airway epithelial cell
line, the expression of AQP3 was upregulated with the
treatment of corticosteroid[22], although the use of corticosteroids
in stable COPD patients is still controversial. It will be
helpful to investigate the possible regulators, including
TNF-α and corticosteroid, for the gene expression of
AQP5, MUC5AC, and mucin secretion in the epithelium airways
of COPD patients.
In summary, the current study demonstrates that
attenuated gene expression of AQP5 existed and was complicated
by mucus hypersecretion in the airways of Chinese COPD
patients. The results suggest the importance of
AQP5 for fluid secretion of submucosal gland airways. Treatment
targeting AQP5 may be an effective approach to COPD therapy.
References
1 Wang F, Feng XC, Li YM, Yang H, Ma TH. Aquaporins as
potential drug targets. Acta Pharmacol Sin 2006; 27: 394_401.
2 Verkman AS, Matthay MA, Song Y. Aquaporin water channels
and lung physiology. Am J Physiol Lung Cell Mol Physiol 2000;
278: L867_79.
3 Verkman AS. Aquaporin water channels and endothelial cell
function. J Anat 2002; 200: 617_27.
4 Borok Z, Verkman AS. Lung edema clearance: 20 years of progress
interview: role of aquaporin water channels in fluid transport in
lung and airways. J Appl Physiol 2002; 93: 2199_206.
5 Miller AL, Strieter RM, Gruber AD, Ho SB, Lukacs NW. CXCR2
regulates respiratory syncytial virus-induced airway
hyperreactivity and mucus overproduction. J Immunol 2003; 170:
3348_56.
6 Kuperman DA, Huang X, Koth LL, Chang GH, Dolganov GM.
Zhu Z, et al. Direct effects of interleukin-13 on epithelial cells
cause airway hyperreactivity and mucus overproduction in asthma.
Nat Med 2002; 8: 885_9.
7 Jayaraman S, Joo NS, Reitz B, Wine JJ, Verkman AS. Submucosal
gland secretions in airways from cystic fibrosis patients have
normal Na+ and pH but elevated viscosity. Proc Natl Acad Sci
USA 2001; 98: 8119_23.
8 Rose MC, Piazza FM, Chen YA, Alimam MZ, Bautista MV, Letwin
N, et al. Model systems for investigating mucin gene expression
in airway diseases. J Aerosol Med 2000; 13: 245_61.
9 Zuhdi Alimam M, Piazza FM, Selby DM, Letwin N, Huang L,
Rose MC, et al. Muc-5/5ac mucin messenger RNA and protein
expression is a marker of goblet cell metaplasia in murine airways.
Am J Respir Cell Mol Biol 2000; 22: 253_60.
10 Hoshino M, Nakamura Y, Sim JJ, Shimojo J, Isogai S. Bronchial
subepithelial fibrosis and expression of matrix
metalloproteinase-9 in asthmatic airway inflammation. J Allergy Clin Immunol
1998; 102: 783_8.
11 Desseyn JL, Buisine MP, Porchet N, Aubert JP, Laine A.
Genomic organization of the human mucin gene MUC5B. cDNA
and genomic sequences upstream of the large central exon. J Biol
Chem 1998; 273: 30 157_64.
12 Boucher RC. Molecular insights into the physiology of the `thin
film' of airway surface liquid. J Physiol 1999; 516: 631_8.
13 Kreda SM, Gynn MC, Fenstermacher DA, Boucher RC, Gabriel
SE. Expression and localization of epithelial aquaporins in the
adult human lung. Am J Respir Cell Mol Biol 2001; 24: 224_34.
14 King LS, Nielsen S, Agre P. Aquaporins in complex tissues. I.
Developmental patterns in respiratory and glandular tissues of
rat. Am J Physiol 1997; 273: C1541_8.
15 Jiao G, Li E, Yu R. Decreased expression of
AQP1 and AQP5 in acute injured lungs in rats. Chin Med J 2002; 115: 963_7.
16 Song Y, Verkman AS. Aquaporin-5 dependent fluid secretion in
airway submucosal glands. J Biol Chem 2001; 276: 41288_92.
17 Ma T, Song Y, Gillespie A, Carlson EJ, Epstein CJ, Verkman AS.
Defective secretion of saliva in transgenic mice lacking
aquapo-rin-5 water channels. J Biol Chem 1999; 274: 20071_4.
18 Krane CM, Fortner CN, Hand AR, McGraw DW, Lorenz JN,
Wert SE, et al. Aquaporin-5 deficient mouse lungs are
hyper-responsive to cholinergic stimulation. Proc Natl Acad Sci USA
2001; 98: 1059_63.
19 Bascom R. Differential susceptibility to tobacco smoke: possible
mechanisms. Pharmacogenetics 1991; 1: 101_6.
20 Towne JE, Krane CM, Bachurski CJ, Menon AG. Tumor necrosis
factor-α inhabits aquaporin 5 expression in mouse lung epithelial
cells. J Biol Chem 2001; 276: 18657_64.
21 Lora JM, Zhang DM, Liao SM, Burwell T, King AM, Barker PA,
et al. Tumor necrosis factor-alpha triggers mucus production in
airway epithelium through an IkappaB kinase beta-dependent
mechanism. J Biol Chem 2005; 280: 36510_7.
22 Tanaka M, Inase N, Fushimi K, Ishibashi K, Ichioka M, Sasaki S,
et al. Induction of aquaporin 3 by corticosteroid in a human
airway epithelial cell line. Am J Physiol 1997; 273: L1090_5.
|
