A Study of the Cough Reflex in Idiopathic
Pulmonary Fibrosis
Benjamin D. M. Hope-Gill, Simon Hilldrup, Christine Davies, Russell P. Newton, and Nicholas K. Harrison
Respiratory Unit and Pathology Department, Morriston Hospital; and Biological Sciences, University of Wales, Swansea, United Kingdom
Little is known about the pathogenesis of cough in idiopathic pulmonary fibrosis (IPF). We hypothesized that abnormalities of respiratory
tract tachykinin-containing sensory nerves may be implicated. We
studied cough response to capsaicin, substance P (SP), and bradykinin
in 10 healthy control subjects and 10 patients with IPF. Six patients
were tested before and after steroid therapy. Induced sputum cell
counts and neurotrophic factor levels were also measured in 13 patients and 13 control subjects. The results show that cough sensitivity
to capsaicin was greater in patients (p ⬍ 0.01). Neither SP nor bradykinin induced cough in normal subjects. SP and bradykinin induced
cough in 7/10 patients (p ⬍ 0.002) and 2/10 patients (not significant)
with IPF, respectively. Prednisolone caused a reduction in cough
sensitivity to capsaicin (p ⬍ 0.05) and SP (p ⬍ 0.05) in all six patients
treated. There were significantly more neutrophils (p ⫽ 0.001) and
higher levels of nerve growth factor (p ⬍ 0.01) and brain-derived
neurotrophic factor (p ⬍ 0.01) in patient’s sputa. These findings
suggest functional upregulation of lung sensory neurones in IPF. The
cough response to inhaled SP in most patients may reflect disrupted
respiratory epithelium. The response to corticosteroids demonstrates
that the cough is amenable to therapy.
Keywords: idiopathic pulmonary fibrosis; cough; neurotrophins
Idiopathic pulmonary fibrosis (IPF) is a condition characterized
by fibroproliferation and modest mononuclear inflammation of
the pulmonary interstitium. Patients typically present with worsening shortness of breath. However, an irritating, nonproductive
cough is an additional distressing feature in 73 to 86% of cases
(1, 2). Little is known about the pathogenesis of this cough,
which frequently proves resistant to conventional antitussive
therapies. Anecdotal evidence suggests that corticosteroids may
be beneficial, but there have been no formal studies to confirm
this. One previous study reported enhanced cough reflex sensitivity to inhaled capsaicin in patients with IPF. By simulating a
restrictive defect in normal subjects, they demonstrated that this
was not due to greater deposition of aerosolized particles in
proximal airways (3). This suggests there is something intrinsic
to the mechanism of disease in IPF that enhances the cough
reflex.
Cough is mediated by the interaction of sensory afferent
nerves, central cough reflexes, and local axon reflexes (4). Various peptides have been implicated in the modulation of the
sensory afferents, and it is known that airway sensory fibers can
mediate neurogenic inflammation as an effector function within
the respiratory epithelium (5). Previous studies suggest there
are two principal cough receptors: rapidly adapting receptors,
(Received in original form April 30, 2003; accepted in final form July 31, 2003)
Supported by a grant from Iechyd Morgannwg Health R&D Consortium. Astra
Zeneca Pharmaceuticals donated the Omeprazole used in this study.
Correspondence and requests for reprints should be addressed to Nicholas K.
Harrison, M.D., F.R.C.P., Respiratory Unit, Morriston Hospital, Swansea, SA6 6NL
UK. E-mail: resp.unit@swansea-tr.wales.nhs.uk
Am J Respir Crit Care Med Vol 168. pp 995–1002, 2003
Originally Published in Press as DOI: 10.1164/rccm.200304-597OC on August 13, 2003
Internet address: www.atsjournals.org
which are innervated by A␦ fibers, and unmyelinated c-fibers (4,
5). Unmyelinated sensory fibers have been shown using immunohistochemical techniques to contain calcitonin gene–related peptides and the tachykinins, substance P (SP) and neurokinin A
(6, 7). When stimulated by irritants such as capsaicin, c-fibers
antidromically release calcitonin gene–related peptides, SP and
neurokinin A (6, 8). Capsaicin is a vanilloid extract from hot
pepper, which acts specifically on c-fibers at low concentrations
but does not cause tachyphylaxis, thus allowing the construction
of dose–response curves (9–13). Animal studies have shown that
pretreatment with bradykinin increases this effect, suggesting that
in inflammatory conditions bradykinin may sensitize sensory afferents (14, 15). By contrast, the in vitro insensitivity of rapidly
adapting receptors to chemical irritants supports the notion that
such irritants have an indirect action on these fibers (4, 5). Furthermore, both SP and neurokinin A have been shown to stimulate
fibroblast proliferation, and neurokinin A increases fibroblast chemotaxis in vitro (16).
In other conditions characterized by cough such as asthma,
it has been reported that there is an increase in tachykinincontaining nerves within the larger airways (17, 18). There is
also evidence of increased levels of neurotrophic factors in the
airways in asthma (19, 20). Neurotrophic factors in turn may
modulate neuronal mechanisms that induce cough. Whether levels of neurotrophic factors are increased in IPF is not known.
In this context the aims of the present study were:
To examine the cough reflex response to capsaicin in a
defined group of patients with IPF, in whom all other
causes of cough had been excluded.
To determine the cough response to bradykinin and SP and
investigate whether these peptides modulate the cough
response to capsaicin.
To measure levels of the neurotrophic factors: nerve growth
factor (NGF), brain-derived neurotrophic factor (BDNF),
and glial cell line–derived neurotrophic factor in the
epithelial lining fluid of the larger airways by analyzing
induced sputum.
To assess the effect of oral corticosteroid therapy on the
cough response to capsaicin and SP in a smaller group
of patients with IPF who had severe cough.
Some of the results of these studies have been previously
reported in the form of abstracts (21–24).
METHODS
Patients
Thirteen patients fulfilling the American Thoracic Society criteria for
the diagnosis of IPF were recruited (25). All patients were elderly (mean
age 71.7 years) and had clinical, and physiologic features consistent
with a diagnosis of IPF. They also had characteristic changes on highresolution computed tomography scan, and therefore surgical lung biopsies were not performed. Thirteen healthy volunteers acted as control
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subjects. Exclusion criteria are shown in Table 1. Informed consent was
obtained from all subjects, and the Local Research Ethics Committee
approved the study.
All 13 patients studied had a metacholine challenge to exclude
airway hyperreactivity (26). No patients or control subjects with a history of gastroesophageal reflux (GER) were included. Patients but not
control subjects were given the proton pump inhibitor Omeprazole
(AstraZeneca Pharmaceuticals Ltd., Loughborough, UK) 20 mg/day
for 1 month before starting the study and continued this therapy for
its duration to exclude subclinical GER. No patients or control subjects
had smoked within 1 year of testing. All subjects were asked to grade
their cough severity from 0 (no cough) to 10 (disabling) using a 10-cm
visual analog scale (VAS). Measurement of pulmonary function was
performed using published guidelines (26, 27).
Cough Challenge
Ten patients and 10 control subjects underwent an initial capsaicin
cough challenge by compressed air–driven dosimeter (Mefar MB3, Bovezzo, Italy) as described elsewhere (16). Dosimeter driving pressure
1.8 ⫾ 0.1 bars, aerosol output of 9.7 mg/second, particle mass mean
diameter 3.5 ␮m, and fixed 1-second inhalation and 60-seconds pause
times were set.
Two further capsaicin challenges were performed. The first repeat
capsaicin challenge occurred at least 1 minute after SP inhalation, and
the second challenge occurred 1 hour later. Similar capsaicin challenges
before and after bradykinin inhalation were performed 1 week later.
One micrometer solutions of each of SP and bradykinin (Clinalfa,
Nottingham, UK) were prepared. Five separate inhalations of each
solution were received within 5 minutes of the initial capsaicin challenge
on each occasion, providing total doses of 0.50 ␮g bradykinin and 0.64
␮g SP, respectively. Inhalations of these substances were received at
60-second intervals.
Six additional patients with IPF, diagnosed using the same criteria
(25), who had disabling cough were treated with oral Prednisolone 40
to 60 mg/day for at least 4 weeks. Patient selection for this group was
based on severity of cough symptom (all had a VAS score greater than
5); however, there was no difference in lung function tests between
these patients and those not treated with steroids. Before starting steroid therapy they underwent a capsaicin cough challenge followed by
SP inhalation, and these tests were then repeated after 4 weeks of
treatment.
Coughs produced within 60 seconds of each inhalation were recorded. Cough threshold was defined as the concentration causing two
or more coughs (C2). The concentration causing five coughs (C5) was
also recorded. The challenge was terminated at C5 or when the maximum concentration of inhalant was reached.
Sputum Induction and Processing
Sputum was induced in 13 patients with IPF and 13 control subjects
by a standard technique (28) using an ultrasonic nebulizer (Sonix 2000;
Medix, Harlow, UK; 0.7 ml/minute output, 5.5 ␮m mean particle size).
Increasing concentrations of hypertonic saline (3, 4, 5%) were administered, each for 5 minutes. Expectorated sputum was collected on ice
and processed immediately at 4⬚C as described previously (29).
Sputum plugs were selected and divided into two parts. Briefly, one
part was treated with 4⫻ wt/vol 0.1% dithiothreitol (DTT) (Sigma, Poole,
UK) plus 4⫻ wt/vol Dulbecco’s phosphate-buffered saline (Sigma). The
suspension was filtered through 48 ␮m nylon gauze (Sefar Ltd, Bury,
UK) and centrifuged at 2,000 rpm for 10 minutes. The supernatant was
decanted and stored at ⫺80⬚C pending measurement of NGF and albumin
levels. The cellular portion was resuspended in Dulbecco’s phosphate
buffered saline and total cell counts were performed using a Neubauer
Haemocytometer (Fisher Scientific, Loughborough, UK). The remaining
selected sputum plugs were treated with equal wt/vol 1% protease
inhibitor cocktail (containing 4-(2-aminoethyl)benzenesulfonyl fluoride
hydrochloride (AEBSF), Aprotonin, Leupeptin, Bestatin, Pepstatin A
and E-64) (Sigma). The suspension was treated as described previously
and the supernatant stored at ⫺80⬚C pending measurement of BDNF
and glial cell line–derived neurotrophic factor levels.
Neurotrophic Factor ELISA Assays
NGF, glial cell line–derived neurotrophic factor (Promega, Southampton, UK), and BDNF (R&D Systems Europe Ltd, Abingdon, UK)
were measured in sputum samples using a quantitative “sandwich”
enzyme assay technique. Samples were run in duplicate and compared
with a standard curve. Results were corrected for sample dilution and
expressed as absolute values.
Statistical Analysis
Negative logarithmic transformation was applied to construct dose–
response curves. Mann–Whitney U test and Wilcoxon’s signed rank
test were used to compare nonparametric data. Fisher’s exact test was
used to compare response to bradykinin and SP. The unpaired t test was
used to compare baseline variables between groups and the Spearman’s
rank correlation was used to analyze association between variables (30).
p Values less than 0.05 was taken as significant (30). The software
package SPSS (SPSS, Inc., Chicago, IL) was used for statistical analysis.
RESULTS
Baseline Data
Demographic and pulmonary function data for patients and control subjects are shown in Table 2. Cough symptom severity as
assessed by VAS is shown in Table 3. Patients with IPF had
significantly greater cough symptom scores assessed by VAS
compared with normal subjects (p ⬍ 0.05). There was no correlation in the IPF group between cough symptom severity as assessed by the VAS score and any measurement of pulmonary
function (FEV1, FVC, total lung capacity, and lung diffusing
capacity for carbon monoxide [DlCO]).
Capsaicin Cough Challenge
In all subjects, a dose–response curve was constructed. No subject coughed during sodium chloride inhalation. After inhalation
of capsaicin, it was observed that the cough response occurred
immediately and was consistently completed within 15 seconds
for both patients and control subjects. No subject had prolonged
bouts of coughing. Figure 1A shows the concentration of capsaicin (⫺log10 ⫾ SEM) required to induce C2 in patients with IPF
(2.97 ⫾ 0.40) was significantly lower than in control subjects
(2.22 ⫾ 0.39; p ⬍ 0.01). The cumulative frequency plot in Figure
TABLE 2. BASELINE DATA FOR PATIENTS WITH IDIOPATHIC
PULMONARY FIBROSIS AND CONTROL SUBJECTS
TABLE 1. STUDY EXCLUSION CRITERIA
1.
2.
3.
4.
5.
6.
Evidence of respiratory tract infection within 1 mo
History of smoking within 1 yr
Postnasal drip, rhinitis, or catarrhal symptoms
Symptoms of GER
Asthma or other respiratory disease, other than IPF
Angiotensin-converting enzyme inhibitor, bronchodilator, or nonsteroidal
antiinflammatory drug therapy
7. Other major systemic illness
Definition of abbreviations: GER ⫽ gastroesophageal reflux; IPF ⫽ idiopathic
pulmonary fibrosis.
Mean age, yr
Sex, male:female
Mean FEV1, % predicted
FVC, % predicted
TLC, % predicted
DLCO, % predicted
Control Subjects
(n ⫽ 10 )
Patients with IPF
(n ⫽ 10 )
59.3
8:2
99.35
103.60
97.95
87.10
71.7*
9:1
84.68
77.43*
67.02*
42.44*
Definition of abbreviations: DLCO ⫽ lung diffusing capacity for carbon monoxide;
IPF ⫽ idiopathic pulmonary fibrosis; TLC ⫽ total lung capacity.
* p ⬍ 0.05 compared with control subjects.
Hope-Gill, Hilldrup, Davies, et al.: Cough in IPF
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TABLE 3. BASELINE COUGH SYMPTOM VISUAL ANALOG
SCALE DATA FOR INDIVIDUAL PATIENTS WITH IDIOPATHIC
PULMONARY FIBROSIS AND CONTROL SUBJECTS
Subject No.
1
2
3
5
6
7
8
9
10
Mean VAS
Control Subjects
Patients with IPF
0
0.5
0
0
3
0
2.5
1
1
0.8
5
0
1.5
3.5
1
4
5
5.5
8
4.0*
Definition of abbreviations: IPF ⫽ idiopathic pulmonary fibrosis; VAS ⫽ visual
analog scale.
* p ⬍ 0.05.
1B further illustrates the difference in C2 response between the
groups. Similar differences were seen for C5 values (IPF 2.70 ⫾
0.60 vs. control subjects 1.01 ⫾ 0.59; p ⬍ 0.01). The mean concentration of capsaicin causing C2 and C5 at the first visit and a
second visit 1 week later was reproducible (Figure 1C). There
was no significant difference in the two-cough response (p ⫽
0.87) or five-cough response (p ⫽ 0.32) to capsaicin in patients
with IPF or in control subjects (p ⫽ 0.44, p ⫽ 0.75, respectively)
between the two visits. In patients with IPF, cough sensitivity
to capsaicin did not correlate with lung function (FEV1, FVC,
total lung capacity, and DlCO), severity of disease on high-resolution computed tomography, or VAS grading of cough symptom
severity.
Effects of SP and Bradykinin Inhalation
There was no significant change in immediate or delayed C2 or
C5 cough response to capsaicin after SP inhalation (Figure 2).
Similar results were obtained after inhalation of bradykinin (results not shown).
Healthy control subjects showed no cough response to either
SP or bradykinin inhalation. However, there was a direct cough
response to inhaled SP in 7/10 patients with IPF (p ⬍ 0.002).
Nine out of 10 patients completed five inhalations of SP. One
patient tolerated one SP inhalation, which caused a prolonged
paroxysm of coughing. As a result, this patient withdrew from
further testing. In the other patients with IPF, cough response
to SP occurred within a few seconds of inhalation and did not
produce prolonged paroxysms of coughing. The total number
of coughs produced in response to five sequential SP inhalations,
each 1 minute apart, ranged between 2 and 56 coughs. However,
patients with IPF showed no consistent dose–response to either
sequential inhalations or increasing concentration (1.0 and 2.0
␮M) of SP.
Only two/nine patients coughed in response to inhaled bradykinin (p ⫽ 0.21). One patient with IPF who had a prolonged
paroxysm of coughing after SP inhalation declined further testing
with bradykinin due to discomfort caused by the previous cough
response.
Effects of Prednisolone Therapy
Figure 3A shows that in all six patients from the additional
group treated with corticosteroid therapy there was a significant
reduction in cough reflex sensitivity to capsaicin (p ⬍ 0.05).
Figure 3B shows that Prednisolone markedly abrogated a direct
cough response to SP inhalation in all patients (p ⬍ 0.03). This
was associated with a reduction in mean VAS score from 7.2 ⫾
Figure 1. The concentration of inhaled capsaicin (mean ⫺log10 ⫾ SEM)
required to induce both two and five coughs in patients with idiopathic
pulmonary fibrosis (IPF) (circles) is significantly lower than in healthy
control subjects (triangles); p value less than 0.01. The negative logarithmic transformation generates a higher numerical value for a lower dose
of capsaicin for a given response (A ). The cumulative frequency plot
in (B ) further illustrates the difference in two coughs (C2) response
to increasing concentration of inhaled capsaicin between the groups.
Reproducibility of C2 and five coughs (C5) response to inhaled capsaicin
on two separate visits is shown in (C ). There was no significant difference
in C2 or C5 cough response in either patients (circles) or control subjects
(triangles) between Visit 1 and Visit 2.
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Figure 2. There was no difference in C2 response to inhaled capsaicin
(mean –log10[capsaicin] ⫾ SEM) in either patients with IPF (triangles)
or control subjects (circles) after substance P (SP) inhalation immediately
or after 1 hour. Furthermore, there was no alteration in concentration
of capsaicin required to induce five coughs after SP inhalation in either
group (data not shown). Similarly, bradykinin inhalation had no effect
on C2 or C5 cough response in either group (data not shown).
0.8 to 2.2 ⫾ 2.5 in this group (p ⬍ 0.05) (Figure 3C). Although
five/six patients did show an improvement in gas transfer coefficient (Kco), there was no significant change in other lung function (FEV1, FVC, total lung capacity, DlCO) over the treatment
period (data not shown).
Sputum Induction
Sputum induction was well tolerated in all subjects without a
significant drop in FEV1. Sputum was induced in all 13 control
subjects; however, adequate supernatant for analyzing NGF levels was obtained in 10/13 patients with IPF and for BDNF analysis in 12/13 patients.
Sputum Cell Counts
Selected sputum was mucoid with a mean percentage squamous
cell count in both patients and control subjects of 8.8 ⫾ 7.6%
and nonsquamous viability of 47.2 ⫾ 17.7% indicating adequacy
of the selection process. The presence of significant numbers of
macrophages and bronchial epithelial cells confirmed origin from
the lower respiratory tract.
The mean total cell count (⫻ 106/g sputum ⫾ SEM) was 4.9 ⫾
4.8 in patients with IPF compared with 1.2 ⫾ 1.0 in control
subjects (p ⫽ 0.001). There were significantly more neutrophils
(2.8 ⫾ 3.9 vs. 0.29 ⫾ 0.31 ⫻ 106/g; p ⫽ 0.001) and lymphocytes
(0.09 ⫾ 0.08 vs. 0.02 ⫾ 0.02 ⫻ 106/g; p ⬍ 0.01) in patients compared with control subjects. There was no significant difference
in absolute count of macrophages (1.82 ⫾ 1.26, patients with
IPF vs. 0.75 ⫾ 0.74, control subjects), eosinophils (0.07 ⫾ 0.09,
IPF vs. 0.01 ⫾ 0.01, control subjects), or bronchial epithelial cell
counts (0.12 ⫾ 0.66, IPF vs. 0.08 ⫾ 0.11, control subjects) between
the groups.
The predominant cell type in patients with IPF was the neutrophil (54 ⫾ 14.5%), whereas macrophages predominated (65 ⫾
19.4%) in healthy control subjects (Figure 4). No subject had a
sputum eosinophilia greater than 3% (Figure 4).
Figure 3. (A ) Demonstrates a significant reduction in C2 response to
inhaled capsaicin (⫺log10[capsaicin]) after steroid therapy in all six patients with IPF studied (p ⬍ 0.05). Steroid therapy also caused abrogation of the direct cough response to inhaled SP (B; p ⬍ 0.03). The total
number of coughs in (B ) represents a cumulative cough score produced
in response to five sequential inhalations of 1.0 ␮M SP solution. (C )
Shows a significant reduction in cough symptom severity (p ⬍ 0.05)
assessed by visual analog scale (VAS) in five/six patients with IPF after
4 weeks of steroid therapy. One patient was unable to reliably indicate
cough symptom severity using the VAS.
There was no difference in serum albumin levels between the
groups.
Neurotrophins in Induced Sputum
Albumin Measurements
Sputum albumin levels (mg/L ⫾ SEM) were 547 ⫾ 359 in patients
with IPF compared with 246 ⫾ 172 in control subjects (p ⬍ 0.01).
Serial dilutions of sputum showed good linearity with each of
the assays used. BDNF was undetectable in samples processed
with DTT, and there was poor average recovery from spiking
Hope-Gill, Hilldrup, Davies, et al.: Cough in IPF
999
Figure 4. Neutrophils were the predominant cell type in
sputa from patients with IPF (patients with IPF are represented by solid circles and control subjects by open circles).
studies in these samples (7.9%). Therefore, measurements of
BDNF were obtained from sputum portions processed separately without DTT, which showed good recovery from spiked
samples (80.8%). Excellent recovery of NGF spiking was obtained from both DTT-processed (91.3%) and neat sputum
(96.1%). However, preliminary studies indicated that higher
levels of NGF were measured in samples processed with DTT.
Therefore, DTT-processed samples were used for measuring
NGF.
Figure 5 shows that levels of NGF and BDNF were higher
in sputum from patients compared with control subjects. Median
values of NGF (ng/ml) were 23.13 in patients and 10.49 in control
subjects (p ⬍ 0.01). Median values of BDNF (pg/ml) were 40.98
in patients and 17.87 in control subjects (p ⬍ 0.01). The glial
cell line–derived neurotrophic factor was undetectable in induced sputa from either group.
DISCUSSION
The initial observations in this study confirm previous findings
that patients with IPF have greater cough reflex sensitivity to
inhaled capsaicin than healthy control subjects and that this
cough response is highly reproducible (3). The careful exclusion
of confounding factors such as bronchial hyperreactivity and
GER strongly suggests that this enhanced cough is part of the
underlying disease process. In addition we show, for the first
time, that most patients with IPF cough in direct response to
inhaled SP but not to bradykinin. Interestingly, neither inflammatory mediator altered cough reflex sensitivity to subsequent
capsaicin challenge. We also demonstrate that oral corticosteroid
therapy reduced cough reflex sensitivity to inhaled capsaicin and
SP in all patients who received treatment. Furthermore, patients
reported a clear reduction in cough symptoms as assessed by
VAS after therapy.
The induced sputum studies demonstrate that patients with
IPF have higher levels of the neurotrophins NGF and BDNF in
their bronchial epithelial lining fluid than healthy control subjects.
Sputum neutrophilia and increased albumin levels reflect wellestablished findings from bronchoalveolar lavage studies in this
condition (31, 32). However, a recent study has demonstrated that
induced sputum samples central rather than peripheral airways
and alveoli (33). It is therefore possible that our findings suggest
inflammation within the more proximal bronchial epithelium
where sensory innervation is greatest. Our observation that there
was no correlation between cough reflex sensitivity, baseline lung
Figure 5. There were higher levels of nerve growth factor
(NGF) (p ⬍ 0.01) and brain-derived neurotrophic factor
(BDNF) (p ⬍ 0.01) within induced sputa from patients with
IPF (solid circles) compared with control subjects (open circles). There was inadequate supernatant obtained from
three patients with IPF to allow NGF measurement and one
patient for BDNF measurement.
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function and severity of cough symptoms in patients with IPF
provides indirect evidence for abnormalities affecting the central
airways. By contrast, lung function tests reflect the disease process
within the lung interstitium.
Interestingly, prior inhalation of either bradykinin or SP did
not result in an increase in cough reflex sensitivity to capsaicin.
This was the case both for those who coughed in response to
SP or bradykinin and those who did not. One reason for this
may be that repeated inhalation of high doses of capsaicin causes
a plateaux in cough response as has been observed previously
in healthy subjects (34). This phenomenon occurred in seven of
our control subjects and two patients with IPF who had a C2
response but did not cough five times even in response to the
maximum concentration of capsaicin tested. However, consistent
dose–response curves were obtained at lower doses despite sequential testing. It appears therefore that in some patients and
most control subjects a maximum cough response is obtained
beyond which no further increase in stimulation produces any
additional effect. Alternatively, it is possible that individual subjects may exhibit idiosyncratic patterns of coughing, for example,
some may control themselves to produce a few coughs only,
whereas others may produce more prolonged paroxysms of
coughing. We therefore conclude that the C2 threshold is a more
reliable measure of cough reflex sensitivity than C5.
Only two previous studies have investigated cough reflex sensitivity in fibrozing alveolitis. Doherty and coworkers found no
association between cough reflex sensitivity to capsaicin and either
cough symptom severity or lung function (3). Lalloo and coworkers made similar observations in patients with pulmonary fibrosis
associated with systemic sclerosis (35). However, neither of these
studies excluded the possible confounders of GER or bronchial
hyperreactivity. Acid reflux is a recognized cause of chronic dry
cough even in the absence of dyspeptic symptoms, and it enhances
cough reflex sensitivity in patients without cough (36–39). It has
also been proposed that GER may be an etiologic factor in IPF
(39). Furthermore, GER is common in patients with systemic
sclerosis, many of whom have esophageal dysmotility (40). For
this reason we were careful to exclude subjects with symptoms
of GER and gave empirical therapy with proton pump inhibitors
to minimize any subclinical effect of this confounding factor (41).
It is possible that the increased cough reflex sensitivity to capsaicin
and SP observed in patients with IPF is caused by pretreatment
with Omeprazole, as control subjects were not pretreated before
testing. However, this seems unlikely given that Omeprazole therapy has been shown to improve cough symptoms (42–44) and
reduce cough response to inhaled capsaicin (44) in patients with
GER. All patients with IPF also had a negative metacholine
challenge, thereby excluding bronchial hyperreactivity. In addition, the absence of sputum eosinophilia in these patients provides
further evidence that an asthmatic component is not responsible
for cough in IPF.
Neurotrophins regulate the development and survival of distinct
subsets of sensory neurones (45). They also act as mediators of
inflammatory hyperalgesia (46, 47). NGF and BDNF levels are
known to be elevated in both inflamed tissues and neurones innervating sites of inflammation (48–50) and they have been shown
to induce synthesis of tachykinins such as SP within noiciceptive
sensory neurones (48–53). NGF also increases nerve conductance
and sensitivity (54). After allergen challenge of the airways of
individuals with asthma, NGF induces phenotypic transformation
of non–tachykinin-containing neurones into tachykinin-containing
nociceptors (48). In addition, NGF and BDNF specifically cause
increased capsaicin sensitivity in sensory neurones (46, 48). In vitro
BDNF but not NGF directly regulates capsaicin sensitivity in
vagal sensory afferents (55), whereas NGF does so primarily in
dorsal root ganglia (56). However, NGF stimulates BDNF gene
transcription in sensory nerves in response to tissue inflammation
(57), thereby suggesting an indirect effect on capsaicin sensitivity
in vivo. Neurotrophins are therefore likely candidates to mediate
the respiratory tract neural hypersensitivity we have observed
in IPF both directly and via increased tachykinin synthesis. It is
also noteworthy that increased levels of SP have previously been
measured in bronchoalveolar lavage fluid in patients with IPF
(58) and that SP is a key mediator of neurogenic inflammation,
which is characterized by microvascular leakage (59).
It is not clear from this study whether the observed increase
in sputum neurotrophin levels represents increased production
in the lungs or increased vascular permeability. However, our
demonstration of higher levels of neurotrophins in sputum of
patients with IPF indicates that these mediators are found in a
location where they could influence the reactivity, differentiation, and proliferation of sensory nerves thereby enhancing the
cough reflex. It is interesting to note that intense NGF-immunoreactivity within the bronchial epithelium and submucosa has
recently been demonstrated in individuals with asthma (60).
Higher neutrophil counts in bronchoalveolar lavage fluid are
known to be related to increased disease severity detected on
computed tomography scanning (61), and it is believed that
neutrophils originate from the cystic spaces associated with pulmonary fibrosis. Therefore, one possible explanation for our
findings is that cells and mediators generated peripherally at
these sites pass proximally on the mucocilary escalator to exert
effects on the epithelium and sensory afferents in larger airways.
Alternatively, our findings may represent direct epithelial infiltration by neutrophils more centrally.
There are several possible mechanisms that may explain the
pathogenesis of cough in IPF. Increased proximal airway deposition of capsaicin seems unlikely given the previous study by
Doherty and coworkers (3). Furthermore, in a study using radiolabeled aerosol particles a greater cough response to inhaled
capsaicin was observed when small (3.5 ␮m mass mean diameter)
particles were used compared with large (5.5 ␮m mass mean
diameter) particles, resulting in more peripheral airways deposition (62).
The observation that cough reflex sensitivity to capsaicin is
enhanced compared with control subjects suggests functional
upregulation of sensory fibers within the respiratory tract. In
this context, it is noteworthy that a significant number of our
patients with IPF coughed in direct response to inhalation of
SP. This phenomenon has been recognized in patients with upper
respiratory tract infections and asthma, both conditions in which
disruption of the respiratory epithelium facilitates access to epithelial sensory afferents. Furthermore, epithelial disruption
causes reduced levels of neutral endopeptidase and angiotensinconverting enzyme, which rapidly metabolize SP and bradykinin.
Therefore, this process may enhance the effect of these mediators on sensory nerves. IPF is characteristically a disease affecting
the interstitium with epithelial disruption, inflammatory cell infiltration, and interstitial edema most evident in the alveoli.
However, our findings of a neutrophilic infiltrate and evidence
for microvascular leakage within induced sputa of patients with
IPF raises the possibility that abnormalities within the proximal
bronchial epithelium may also occur.
In conclusion, this study confirms enhanced cough reflex sensitivity in patients with IPF and demonstrates that most patients
have a direct cough response to SP. The increased response to
capsaicin, which is c-fiber–specific, supports the hypothesis that
there is functional upregulation of respiratory tract sensory
nerves in IPF. The direct cough response to SP in some patients,
the abrogation of this effect by steroid therapy, and the presence
of neurotrophins in airway epithelial lining fluid support the
notion that an inflammatory process can affect more proximal
Hope-Gill, Hilldrup, Davies, et al.: Cough in IPF
airways. The demonstration that cough in IPF is amenable to
therapeutic intervention is encouraging and should promote further investigation into this phenomenon.
1001
23.
Conflict of Interest Statement : B.D.M.H.-G. has no declared conflict of interest;
S.H. has no declared conflict of interest; C.D. has no declared conflict of interest;
R.P.N. has no declared conflict of interest; N.K.H. has no declared conflict of
interest.
24.
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