(Chest. 2000;117:386S-389S.)
© 2000
American College of Chest Physicians
Role of Neutrophil Elastase in Hypersecretion During COPD Exacerbations, and Proposed Therapies*
Jay A. Nadel, MD
*
From the Cardiovascular Research Institute, Departments of Medicine and Physiology, University of California, San Francisco, CA.
Correspondence to: Jay A. Nadel, MD, University of California, San Francisco, CVRI, 505 Parnassus Ave, M-1325, San Francisco, CA 94143-0130; e-mail: janadel{at}itsa.ucsf.edu
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Abstract
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A common feature of COPD and other chronic lung diseases is
hypersecretion of mucus into the airways, causing peripheral airway
plugging and further airflow obstruction. The mucus is secreted by
goblet cells, which are present in excessive numbers in COPD. This
review describes how neutrophils in the airways of COPD patients
stimulate the goblet cells to secrete their products. Recent findings
on the mechanisms of neutrophil stimulation of goblet cell
degranulation are discussed. These implicate the proteolytic enzyme
elastase and cell surface adhesion molecules, and provide a basis for
the investigation of potential novel therapies.
Key Words: CD11b/CD18 • hypersecretion • goblet cell degranulation • intercellular adhesion molecule-1 • neutrophil elastase
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Introduction
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Airway
hypersecretion is a characteristic feature in COPD and is a significant
contributor to exacerbations. Excess mucus in the airways causes severe
cough and discomfort, and can lead to further obstruction and
inflammation. The mechanism of hypersecretion is, however, poorly
understood. The airway epithelium undergoes metaplastic change in COPD,
resulting in an excess of goblet cells; these cells, along with the
submucosal glands, are responsible for the mucus production. In
designing a therapeutic strategy to combat hypersecretion, two
approaches might thus be considered: inhibiting the growth of the
goblet cells, or preventing their degranulation, ie, the
release of their secretory products. The mechanism of goblet cell
degranulation in COPD is currently not known. This review highlights
the possible role of neutrophils (which are present in inflammatory
sites in the airways) in stimulating goblet cell degranulation and
mucus hypersecretion in COPD and exacerbations.
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Neutrophils Stimulate Goblet Cell Degranulation
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Neutrophils are recruited into the airways in COPD and other
chronic respiratory diseases1
2
by chemoattractants that
are present in the airways3
4
5
; in vitro and
in vivo studies show that these neutrophils can stimulate
goblet cell degranulation.6
The guinea pig provides a
convenient model to study degranulation, since the airways normally
contain goblet cells.7
Direct tracheal injection of
neutrophil chemoattractants such as interleukin (IL)-8 or
N-formyl-methyl-leucyl-phenylalanine (fMLP) results in migration of
neutrophils into the lumen of the trachea. The tracheas of these
animals can then be removed and examined microscopically, both for
neutrophil infiltration (by myeloperoxidase staining) and for goblet
cell degranulation (by Alcian blue [AB]/periodic acid-Schiff [PAS]
staining of mucosubstances on the mucosal surface epithelium; Fig 1
). Degranulation is quantified by volume density of staining using a
semiautomatic imaging system (the public domain National Institutes of
Health Image program, available at zippy.nimh.gov, or on disk from the
National Technical Information Service, Springfield, VA; part number
PB95–500195GEI).
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Figure 1.. Photomicrographs of human bronchial epithelium
removed from patient with idiopathic pulmonary fibrosis at the time of
lung transplantation and stained with AB/PAS. In the
unstimulated condition (top left, a), AB-positive cells
are conspicuous in the bronchial epithelium. Bronchial segments that
are incubated with neutrophils alone (top right, b) or
with IL-8 alone (bottom left, c) do not show
degranulation. However, segments incubated with IL-8 plus neutrophils
show profound goblet cell degranulation (arrows; bottom right,
d). Reprinted with permission from Takeyama et
al.6
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Injection of IL-8 or fMLP was found, as expected, to result in
neutrophil infiltration of the guinea pig trachea, maximal at 1 to
2 h.6
It was also demonstrated that this infiltration
was followed by profound goblet cell degranulation (maximal at 4
h). Moreover, inhibition of neutrophil recruitment, using the leukocyte
migration inhibitor NPC 15669, also inhibited goblet cell
degranulation. These data, therefore, strongly suggest that
neutrophils, which infiltrate the airways in COPD, promote in the
degranulation of goblet cells and the resulting hypersecretion and
concomitant problems.
The Role of Neutrophil Elastase
Purified neutrophil elastase has been shown previously to be a
potent secretagogue for goblet cells in
vitro.8
9
However, it is not clear whether this
molecule is responsible for the effect of neutrophils on degranulation,
since the cells themselves release little or no elastase in
vitro, even after "activation" with a variety of molecules
including IL-8, fMLP, leukotriene-B4, and tumor
necrosis factor-
.10
Rather, activation of neutrophils
causes a translocation of elastase from the azurophilic granules to the
cell surface.11
In the guinea pig model, however, we have
shown that an elastase inhibitor, ICI 200355, inhibited the
neutrophil-dependent goblet cell degranulation seen after tracheal
instillation of neutrophil chemoattractants.7
These
results confirmed the role of neutrophil elastase in degranulation, but
the question still remained as to how elastase exerts its effect, since
neutrophil activators/chemoattractants do not appear to promote its
release. To investigate this question, isolated human neutrophils were
incubated with segments of guinea pig trachea, and the effects of a
variety of agents on degranulation were assessed. It was confirmed,
firstly, that activation of neutrophils with IL-8 and other
chemoattractants resulted in expression of cell surface elastase. It
was also shown that preactivated neutrophils, washed with sterile
phosphate-buffered saline solution to avoid further contamination with
IL-8, could still promote degranulation, while the cell-free
supernatant from activated neutrophils had no effect. This demonstrated
that it is cell-associated elastase, rather than the released enzyme,
which is involved in degranulation.
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The Role of Adhesion Molecules
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The role of cell-associated elastase in goblet cell degranulation
implies a direct interaction between the neutrophils and goblet cells.
The possible role of adhesion molecules in this process has therefore
been investigated. Human neutrophils and human tracheal segments,
obtained from patients undergoing lung transplantation, were chosen for
these experiments. These tracheae contained a high proportion of goblet
cells (> 20%) due to the pathologic condition involved (pulmonary
fibrosis, cystic fibrosis, COPD). The data from guinea pigs were
confirmed in the human system; IL-8-activated neutrophils stimulated
degranulation, and this was inhibited by elastase inhibitors.
Furthermore, when neutrophils were pretreated with monoclonal antibody
against cell surface adhesion molecules, antibodies to integrins of the
ß2 subfamily (CD11b [Mac-1] or CD18 [lymphocyte
function-associated antigen-1ß]) also inhibited IL-8 mediated
degranulation (Fig 2
). A similar inhibitory effect was obtained by pretreatment of
the airway tissue with antibody to intercellular adhesion molecule-1.
These results suggest that neutrophil-stimulated goblet cell
degranulation is mediated by a direct interaction between the
neutrophils and the goblet cells in the airway epithelium involving
intercellular adhesion molecule-1 and integrins of the ß2 subfamily.
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Figure 2.. Effects of neutrophils (106 cells/mL)
alone, IL-8 (10-7 M) alone, neutrophils plus IL-8, or
neutrophils "activated" by IL-8 on goblet cell degranulation
(expressed as percent area of epithelium stained with AB/PAS in human
bronchial segments in vitro) and effects of anti-CD18
antibody on goblet cell degranulation caused by "activated"
neutrophils. For explanation, see text. Responses are expressed as mean
± SEM; n = 4 for each group. *p < 0.01, significantly
different from control values.  p < 0.05, significantly different
from the response to "activated" neutrophils. M = molar.
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Discussion
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Metaplasia of the airway epithelium in COPD and other airway
diseases results in large numbers of goblet cells in the peripheral
airways. When these goblet cells are stimulated to release their
contents, the excess mucus in the airways can cause severe cough,
airflow obstruction, peripheral airway plugging, and may lead to
further inflammation and possibly infection. Neutrophils, which migrate
into the airways in COPD, and particularly into inflammatory sites
during exacerbations, stimulate goblet cell degranulation. These cells
thus play a very important role in hypersecretion.
The experiments discussed here indicate the likely mechanism by which
neutrophils promote hypersecretion, and thus offer a number of possible
avenues for design of therapeutic strategies. Firstly, prevention of
neutrophil migration into the airways is one obvious mechanism by which
hypersecretion could be reduced. This might also reduce inflammation,
but could also have unwanted side effects, including an impaired
ability to combat infection. Selective inhibition of neutrophil
elastase might, however, inhibit hypersecretion without other less
desirable inhibitory effects on neutrophil activity. Similarly,
interfering with the neutrophil/goblet cell interaction, either using
integrin/adhesion molecule antagonists or regulating cell surface
expression, offers another interesting possibility.
Neutrophils are undoubtedly crucial players in the pathogenesis of COPD
and its exacerbations, and we are now beginning to understand the
mechanisms by which these cells mediate hypersecretion. Armed with this
knowledge, it is now possible to consider ways in which the
debilitating consequences of hypersecretion can be alleviated.
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Footnotes
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Abbreviations: AB = Alcian blue;
fMLP = N-formyl-methionyl-leucyl-phenylalanine; IL = interleukin;
PAS = periodic acid-Schiff
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References
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TOP
Abstract
Introduction
