Cellular Microbiology (2009) 11(2), 208–216
x
Microreview
Cystic fibrosis and innate immunity: how chloride
channel mutations provoke lung disease
Gerd Döring1* and Erich Gulbins2**
1
Institute of Medical Microbiology and Hygiene,
Wilhelmstrasse 31, 72074 Tübingen, Germany.
2
Department of Molecular Biology and Center for Medical
Biology, Hufelandstrasse 55, 45122 Essen, Germany.
Summary
Innate immunity is essential for prevention of
infection in vertebrates and plants and dysfunction
of single components of innate immunity may
provoke severe disease. Here we describe how
mutations in the cystic fibrosis transmembrane
conductance regulator gene dysregulate a variety
of components of the innate immune system in
individuals suffering from the hereditary disease
cystic fibrosis. In the airways of these individuals,
functions of the mucociliary clearance system, cat-
ionic antimicrobial (poly)peptides and neutrophils
and macrophages are impaired and inflammatory
signal transduction pathways exaggerated. Conse-
quently, chronic airway colonization with opportu-
nistic bacterial pathogens develops and leads to
life-threatening lung disease.
Introduction
In contrast to adaptive immunity, innate immunity provides
rapid prevention of infection. This remarkable ability of
innate immunity is based on the rapid recruitment of
professional phagocytic cells, omnipresent germ-line-
encoded pattern-recognition receptors, effective signal
transduction systems, immediate production of reactive
oxygen species (ROS) and a large armatoire of preformed
antimicrobial molecules, produced by various epithelial
cells. Another component of innate immunity, the muco-
ciliary clearance (MCC) system, protecting the human
respiratory tract, is permanently active and does not need
Received 26 September, 2008; revised 17 November, 2008;
accepted 20 November, 2008. For correspondence. *E-mail
gerd.doering@med.uni-tuebingen.de; Tel. (+49) 7071 298 2069;
Fax (+49) 7071 29 3011; **E-mail: erich.gulbins@uni-due.de; Tel.
(+49) 201 723 3118; Fax (+49) 201 723 5974.
© 2008 Blackwell Publishing Ltd
doi:10.1111/j.1462-5822.2008.01271.
First published online 17 December 2008
activation by microbial challenges. Given the essential
role of the innate immune system in preventing infection,
it is conceivable that dysfunction of even one of its many
components can cause disease. This is demonstrated
by the fairly large number of primary immunodeficiency
diseases, in which genetic defects provoke a variety of
microbial infections (Notarangelo et al., 2006).
Although not regarded as a primary immunodeficiency
disease, a variety of innate immune functions have been
reported to be dysregulated in cystic fibrosis (CF), a
hereditary disease caused by mutations in a membrane-
bound chloride channel, named CF transmembrane con-
ductance regulator (CFTR) (Kerem et al., 1989; Riordan
et al., 1989; Rommens et al., 1989). These concern
the MCC system, cationic antimicrobial (poly)peptides
(CAMPs), neutrophils, macrophages and CFTR itself. A
new twist to the pathophysiology of CF was found only
recently, when the effect of mutated CFTR on ceramide
accumulation was detected. Dysregulation of these innate
immune functions causes chronic bacterial lung infections
which are mainly responsible for the reduced life expect-
ancy in CF patients (CF Foundation, 2005). CF provides
not only a fascinating insight into biological mechanisms
of various innate immune functions, but also into the inter-
play of host defence functions and survival strategies of
opportunistic bacterial pathogens.
Mucociliary clearance
The inability of mutated CFTR to effectively secrete chlo-
ride from respiratory epithelial cells into the airway surface
liquid causes excessive water absorption from the airway
surface liquid leading to impaired MCC in individuals with
CF (Matsui et al., 1998; Boucher, 2007) (Fig. 1). This in
turn facilitates the colonization of the viscous mucus layer
on the respiratory epithelium with bacterial pathogens
(Ulrich et al., 1998; Worlitzsch et al., 2002). Although
studies using respiratory epithelial cell cultures from CF
individuals and controls convincingly demonstrate a
highly impaired MCC velocity, in vivo data from CF
patients suggest MCC rates are heterogeneous (Boucher,
2007). The nature of compensatory mechanisms which
allow at least partial function of the MCC system in CF
cellular microbiology
 Cystic fibrosis and innate immunity 209

Fig. 1. Bacterial killing mechanism of innate immunity in the respiratory tract of healthy individuals and cystic fibrosis (CF) individuals.
A. In healthy individuals, bacteria, entering the mucus layer overlaying the respiratory epithelium, are effectively removed from the airways by
a functional mucociliary clearance system and killed by CAMPs derived from submucosal glands or epithelial cells, by functional neutrophils or
macrophages or within epithelial cells after uptake via functional CFTR. PMN, polymorphonuclear leukocytes.
B. In individuals with CF defective CFTR leads to a highly viscous mucus layer which impairs mucociliary clearance, the migration of CAMPs,
neutrophils and macrophages towards the bacterial targets, the uptake of bacteria by CFTR itself, induces a pH shift in epithelial lysosomes
which due to ceramide accumulation results in DNA deposits which in turn serve as adhesion matrices for bacteria. A similar pH shift in the
phagolysosomes of macrophages impairs bacterial killing. Ceramide accumulation also triggers cytokine release which induces further
neutrophil influx.

patients needs to be studied. Ideally, this should be
studied in uninfected airways of CF individuals which are
rather difficult to find. Studies in CF mice, on the other
hand, may be less informative, since the MCC system
differs anatomically from that in humans.
Preformed antimicrobial molecules
Viscous secretions may obstruct submucosal gland ducts
in CF due to mutated CFTR (Engelhardt et al., 1992).
Consequently, preformed bacteriolytic enzymes and
CAMPs such as b-defensins and cathelicidins (Lehrer and
Ganz, 2002; Selsted and Quellette, 2003), produced by
submucosal glands, may not be secreted onto the epithe-
lium (Verkman et al., 2003; Joo et al., 2004) (Fig. 1),
resulting in facilitated colonization and infection of the
respiratory epithelium with bacterial pathogens. Attempts
to determine the concentration of CAMPs or their traffick-
ing in airways of CF individuals are still missing. In con-
trast to the notion that defective CFTR would result in

© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216

 210 G. Döring and E. Gulbins
hypotonic salt concentrations which become isotonic after
volume depletion, others have hypothesized that hyper-
tonic salt concentrations are present on the respiratory
epithelium (Smith et al., 1996). Such concentrations
would inactivate salt-sensitive CAMPs, thereby leading to
bacterial multiplication and subsequent infection.
However, it has been difficult to prove that the airway
surface liquid in CF is indeed hypertonic (Boucher, 2007).
Ceramide accumulation
Biological membranes contain predominantly cholesterol,
phospholipids and sphingomyelin. The latter is hydrolysed
by the activities of acid, neutral and alkaline sphingomyeli-
nases to ceramide. The acid sphingomyelinase (Asm),
residing within pre-lysosomes, lysosomes and secretory
lysosomes and on the outer leaflet of the cell membrane
(Grassmé et al., 2001), is involved in cell death and is
upregulated upon infection (Grassmé et al., 1997; 2003).
Recently, an abnormal age-dependent accumulation of
ceramide in the lungs of cftr-deficient mice and in epithelial
cells from CF patients has been detected (Teichgräber
et al., 2008). As a consequence, the rate of cell death
increased in respiratory epithelial cells of uninfected CF
mice, resulting in the formation of DNA deposits on the res-
piratory epithelium, which facilitated bacterial adherence.
The link between accumulated ceramide, cell death and
infection with Pseudomonas aeruginosa had been demon-
strated by treating CF mice with amitriptyline which blocked
ceramide accumulation, reduced cell death and normal-
ized the susceptibility of CF mice to P. aeruginosa infection
(Teichgräber et al., 2008). Further evidence for this notion
was provided by experiments in Cftr-/-/Asm+/- mice. In
these mice ceramide accumulation, cell death rates and
susceptibility to P. aeruginosa infection were normalized.
Ceramide accumulation also provoked a pro-
inflammatory status in the respiratory tissue of mice with
CF, which preceded bacterial infection: an increased syn-
thesis and release of cytokines and a subsequent, age-
dependent, recruitment of macrophages and neutrophils
had been observed in lungs of uninfected CF mice (Fig. 1)
(Teichgräber et al., 2008). That these phenomena are trig-
gered by accumulated ceramide was demonstrated by
experiments revealing that partial Asm inhibition by ami-
triptyline or Asm heterozygosity abrogated effector cells
accumulation in CF mouse lung tissues. How the accu-
mulation of macrophages and neutrophils in lung tissues
of uninfected CF mice contributes to the hypersuscepti-
bility of CF mice to P. aeruginosa lung infections remains
to be defined. The results unify several previously pub-
lished observations in CF mouse strains with regard to
inflammation (Zahm et al., 1997), apoptosis (Cannon
et al., 2003) and hypersusceptibility to P. aeruginosa
infection (Pier, 2000; Coleman et al., 2003).
How mutated CFTR mediates ceramide accumulation is
still discussed. Acidification of intracellular vesicles medi-
ated by Cftr seems to be critical for the concerted regula-
tion of ceramide by the activities of Asm and acid
ceramidase (Fig. 2) (Teichgräber et al., 2008). According
to current models, sphingomyelin is constitutively meta-
bolized to ceramide by Asm and further degraded to
sphingosine by acid ceramidase (Kolesnick et al., 2000).
The alkalinization of Cftr-deficient vesicles in respiratory
cells of CF mice from pH 4.5 to a pH of 5.9 causes an
imbalance of the Asm and acid ceramidase activities,
resulting in a net accumulation of ceramide. Two previous
reports support the notion of defective acidification as a
result of diminished Cl- conductance in other CF cell types.
These studies demonstrated defective acidification in the
Golgi/trans-Golgi network, pre-lysosomes and endosomes
of CF cell lines, leading to abnormal glycosylation of mem-
brane proteins (Barasch et al., 1991), and showed the
same phenomenon in phagolysosomes of alveolar macro-
phages from cftr-null mice, leading to impaired bactericidal
activity (Di et al., 2006). However, other studies that inves-
tigated the pH in phagosomes in wild type and cftr-deficient
cells employing zymosan conjugates containing fluores-
cein and tetramethylrhodamine did not report a failure of
these vesicles in CF cells to acidify, suggesting that a
distinct expression pattern of ion channels including Cftr
determines the pH in different vesicle populations (see
below).
Epithelial cells
Epithelial cells use CFTR as a receptor for internalization of
P. aeruginosa via endocytosis and subsequent removal
of bacteria from the airway. Thus, CFTR itself can be
regarded as a substantial part of the innate immune system
and mutant or missing CFTR has been linked to the
initiation of P. aeruginosa infection (Pier, 2000), since the
CFTR–P. aeruginosa interaction does not occur, allowing
for increased bacterial loads in the lungs. Binding occurs
between the outer core of the bacterial lipopolysaccharide
and amino acids 108–117 in the first predicted extracellular
domain of CFTR. In experimentally infected mice, inhibiting
CFTR-mediated endocytosis of P. aeruginosa by inclusion
in the bacterial inoculum of either free bacterial lipopoly-
saccharide or CFTR peptide 108–117 resulted in increased
bacterial counts in the lungs suggesting that internalization
contributes – via still unknown mechanisms – to the clear-
ance of the bacteria in the lung (Fig. 1) (Pier, 2000).
Professional phagocytes
Cell migration
The high viscosity of CF secretions negatively affects
migration of neutrophils (Matsui et al., 2005) (Fig. 1). The
© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216

Fig. 2. Consequences of defective acidification in lysosomes of
epithelial cells and the phagolysosome of macrophages for
bacterial infection.
A. In lysosomes and phagosomes of normal respiratory epithelial
cells and macrophages, respectively, sphingomyelin is constitutively
metabolized to ceramide by acid sphingomyelinase (Asm) and
further degraded to sphingosine by acid ceramidase (Ac). Due to
its counterion effect of Cl- in promoting lumenal H+ accumulation by
the lysosomal V-type H+-ATPase, CFTR ensures acidification of
lysosomes and late phagosomes to pH < 5. In macrophages this
pH provides optimal bactericidal activities for degradative enzymes
when lysosomes fuse with phagosomes during the maturation
process of these organelles.
B. In individuals with CF defective CFTR leads to an imbalance of
Asm and Ac activities which leads in turn to ceramide
accumulation. As a consequence of the alkalinization, increased
cytokine release, cell death and decreased bacterial killing are
observed leading to fibrosis and bacterial infection.
© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216
Cystic fibrosis and innate immunity 211
failure of neutrophils to rapidly encounter the pathogens in
the viscous mucus may allow bacterial multiplication and
profound changes on the bacterial phenotypes which
facilitate chronic infection (see below). The unaccom-
plished mission to kill mucus-encased bacteria will result
in increased endobronchial DNA deposition, further
increasing mucus viscosity, bacterial adhesion and biofilm
formation (Whitchurch et al., 2002; Worlitzsch et al., 2002;
Matsui et al., 2006) in addition to ceramide-driven DNA
deposition, as well as the liberation of granule-stored pro-
teases and antimicrobial peptides which finally leads a
lung tissue remodelling (Nathan, 2006). Increasing mucus
viscoelasticity then also impairs antimicrobial compounds
such as lactoferrin to encounter their bacterial targets
(Matsui et al., 2006). Impaired neutrophil migration may
also be partly responsible for the reduced efficacy of
vaccines against P. aeruginosa in individuals with CF,
despite the development of high and long-lasting
P. aeruginosa-specific antibody titres after vaccination
(Döring et al., 2007).
Defective acidification
A recent study demonstrated that lysosomal CFTR par-
ticipates in phagosomal pH control (Fig. 2) (Di et al.,
2006). Due to its counterion effect of Cl- in promoting
lumenal H+ accumulation by the lysosomal V-type
H+-ATPase, CFTR ensures acidification of late phago-
somes to pH < 5 and provides optimal bactericidal activi-
ties for degradative enzymes when lysosomes fuse with
phagosomes during the maturation process of these
organelles. Alveolar macrophages from CFtr -/- mice
retained the ability to phagocytose and generate an oxi-
dative burst, but exhibited defective killing of internalized
bacteria (Di et al., 2006). The contribution of CFTR to the
regulation of pH in intracellular organelles, first described
in trans-Golgi vesicles (Barasch et al., 1991), has been
controversially discussed (Seksek et al., 1996; Haggie
and Verkman, 2007), possibly due to experimental diffi-
culties to ensure targeting of identical vesicle populations
by pH-sensitive fluorescent dyes.
Oxidative killing
In the bronchi, the highly viscous mucus generates a
microaerobic milieu which may become anaerobic by
a rapid oxygen consumption by pathogens such as
P. aeruginosa (Worlitzsch et al., 2002). Consequently, the
generation of ROS by neutrophils and other cells is abol-
ished and bacterial killing impaired (Fig. 3). The observa-
tion that anaerobic conditions impair apoptosis (Hannah
et al., 1995; Zhang et al., 2003) and NETosis (Brinkmann
and Zychlinsky, 2007) may explain why neutrophil necro-
sis is so dominant in infected airways of individuals with
 212 G. Döring and E. Gulbins

Fig. 3. Bacterial sensitivity and resistance to neutrophil-mediated killing in aerobic and anaerobic environments.
A. On the respiratory epithelium of healthy individuals, neutrophils effectively kill the CF-related bacterial pathogens P. aeruginosa, S. aureus
and B. cepacia by reactive oxygen species (ROS) and defensins.
B. Neutrophils in the anaerobic milieu in the mucus layer overlaying the respiratory epithelium in individuals with CF are devoid of ROS,
leading to intra-neutrophil survival of S. aureus and B. cepacia which are resistant to non-oxidative killing. The anaerobic milieu also triggers
biofilm formation of S. aureus and P. aeruginosa which prevents phagocytosis.

© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216


CF. Whether lack of ROS also impairs the non-oxidative
innate defence mechanisms (Segal, 2005) has been
controversially discussed (Nathan, 2006). Nevertheless,
microaerobic/anaerobic environmental conditions on the
respiratory epithelium allow microbial pathogens, intrins-
ingly resistant to non-oxidative killing or capable to
change their phenotype to become rapidly resistant to
non-oxidative killing, to persist in airways of individuals
with CF.
This notion is in line with the observed microbial species
which cause disease in individuals with CF: the
microaerobic/anaerobic environmental conditions on the
respiratory epithelium in CF provoke a switch from single
cells of P. aeruginosa which are sensitive to both oxidative
and non-oxidative killing by neutrophils (Sousa et al.,
2007) to a mucoid phenotype (Worlitzsch et al., 2002;
Matsui et al., 2006) now resistant to non-oxidative killing
by phagocytes. Additionally, structural changes of the
outer membrane occur during anaerobic growth of
P. aeruginosa (Sabra et al., 2003). This change provides
enhanced resistance to cationic antimicrobials, such as
aminoglycoside antibiotics, and potentially to naturally
produced cationic peptides due to inhibition of charge-
mediated uptake. Similarly, Staphylococcus aureus
changes its phenotype under microaerobic/anaerobic
environmental conditions forming polysaccharide-
encased cells which confer resistance to non-oxidative
killing (McKenney et al., 1999; Cramton et al., 2001;
Ulrich et al., 2007). Finally, strains of the Burkholderia
cepacia complex, intrinsingly resistant to non-oxidative
killing, cause disease in individuals with CF (Sousa
et al., 2007). Since the anaerobic environment is locally
restricted to the lung epithelium, bacterial sepsis is virtu-
ally not observed in CF and chronic microbial infections
remain localized to the endobronchial site. Interestingly,
S. aureus and B. cepacia complex strains but not
P. aeruginosa strains are pathogens in chronic granulo-
matous disease, caused by mutations in the NADPH
oxidase (Heyworth et al., 2003).
Besides P. aeruginosa, S. aureus and B. cepacia
complex strains, also other microbial species colonize the
airways of CF patients. The microaerobic/anaerobic envi-
ronmental conditions in the sputum let strict anaerobic
species grow to cell numbers comparable to those of
P. aeruginosa (Tunney et al., 2008). A large number of
different species have been detected and future studies
will provide information whether these bacteria contribute
to the pathophysiology of CF lung disease. Taken
together, only bacteria that are able to protect themselves
from non-oxidative killing by the production of toxins or
biofilm formation and that can sufficiently grow rapidly to
high cell numbers in this specific niche become dominant
pathogens in CF lungs. Apparently, enteric bacteria lack
some of these characteristics.
© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216
Cystic fibrosis and innate immunity 213
Limitation of innate and adaptive immune function
during chronic infection
The exaggerated inflammatory response to persistent
infection not only facilitates tissue destruction but also
induces tissue remodelling, in which a number of pro-
teases, growth factors and other compounds from various
cells are involved. During chronic inflammation, several
immune cell functions are impaired. For instance, human
neutrophil elastase cleaves immunoglobulins, comple-
ment components (Brozna et al., 1977; Döring et al.,
1986; Berger et al., 1989), and a variety of cell receptors
(Sommerhoff et al., 1990; Döring et al., 1995; Ferry et al.,
1997; Voynow et al., 1999; Döring et al., 2000; Walsh
et al., 2001; Henriksen et al., 2004; Caldwell et al., 2005;
Suzuki et al., 2005; Hartl et al., 2007) and surfactant pro-
teins (Griese et al., 2004; Hirche et al., 2004; Rubio et al.,
2004). Persistent levels of active elastase also contribute
to ineffective removal of apoptotic cells and continuous
low-grade inflammation by cleavage of the phosphati-
dylserine receptor (Vandivier et al., 2002). Further, human
neutrophil elastase downregulated the expression of
CD40, CD80 and CD86 on DCs and inhibits DC matura-
tion (Roghanian et al., 2006). Finally, neutrophil elastase
cleaves its own endogenous inhibitor, a-1-antitrypsin
when protease concentrations are higher than inhibitor
concentrations (Goldstein and Döring, 1986).
Conclusion
Cystic fibrosis is unique in its remarkable number of dys-
regulated innate immune functions which are conse-
quences of mutations in CFTR. Synergistically, they may
provoke lung disease in virtually all subjects caused by a
limited number of microbial pathogens adapted to this
specific environmental niche. Combined, defective innate
immunity leads to chronic disease in which a vicious cycle
still worsens the function of innate defence mechanisms.
For instance, accumulation of ceramide on the respiratory
epithelium may still increase after the onset of chronic
P. aeruginosa infection since the pathogen itself triggers
Asm activation (Grassmé et al., 2003), and, hence, may
further facilitate bacterial adhesion by increasing the depo-
sition of DNA, impair MCC by increasing the viscoelasticity
of the airway surface liquid, and augment tissue destruc-
tion and remodelling by increasing inflammation. Neu-
trophil necrosis will further enhance extracellular DNA
concentrations. The anaerobic niche, in which biofilm
bacteria surrounded by viscous mucus persist, will impair
apoptosis and induce necrosis of neutrophils, thus
enhancing the volume and the viscosity of mucus plugs.
Increased sputum production then will attract strict anaero-
bic bacteria which further complicate the pathophysiology
of lung disease in CF. Unravelling dysregulation of innate
 214 G. Döring and E. Gulbins
immunity in CF may allow developing drugs with the aim to
normalize innate immune functions.
Acknowledgements
The authors want to thank Peter Michael Weber, Children’s
Clinic, University of Tübingen, Germany, for excellent graphic art.
References
Barasch, J., Kiss, B., Prince, A., Saiman, L., Gruenert, D.,
and al-Awqati, Q. (1991) Defective acidification of intracel-
lular organelles in CF. Nature 352: 70–73.
Berger, M., Sorensen, R.U., Tosi, M.F., Dearborn. D.G., and
Döring, G. (1989) Complement receptor expression on
neutrophils at an inflammatory site, the Pseudomonas-
infected lung in CF. J Clin Invest 84: 1302–1313.
Boucher, R.C. (2007) Airway surface dehydration in CF:
pathogenesis and therapy. Annu Rev Med 58: 157–170.
Brinkmann, V., and Zychlinsky, A. (2007) Beneficial suicide:
why neutrophils die to make nets. Nat Rev Microbiol 5:
577–582.
Brozna, J.P., Senior, R.M., Kreutzer, D.L., and Ward, P.A.
(1977) Chemotactic factor inactivators of human granulo-
cytes. J Clin Invest 60: 1280–1288.
Caldwell, R.A., Boucher, R.C., and Stutts, M.J. (2005) Neu-
trophil elastase activates near-silent epithelial Na+ chan-
nels and increases airway epithelial Na+ transport. Am J
Physiol 288: L813–L819.
Cannon, C.L., Kowalski, M.P., Stopak, K.S., and Pier, G.B.
(2003) Pseudomonas aeruginosa-induced apoptosis is
defective in respiratory epithelial cells expressing mutant
CF transmembrane conductance regulator. Am J Respir
Cell Mol Biol 29: 188–197.
CF Foundation (2005) Patient Registry Annual Report
2004. Bethesda, MD [WWW document]. URL http://www.
cff.org/LivingWithCF/CareCenterNetwork/PatientRegistry/.
Coleman, F.T., Mueschenborn, S., Meluleni, G., Ray, C.,
Carey, V.J., Vargas, S.O., et al. (2003) Hypersusceptibility
of CF mice to chronic Pseudomonas aeruginosa oropha-
ryngeal colonization and lung infection. Proc Natl Acad Sci
USA 100: 1949–1954.
Cramton, S.E., Ulrich, M., Gotz, F., and Doring, G. (2001)
Anaerobic conditions induce expression of polysaccharide
intercellular adhesin in Staphylococcus aureus and Sta-
phylococcus epidermidis. Infect Immun 69: 4079–4085.
Di, A., Brown, M.E., Deriy, L.V., Li, C., Szeto, F.L., Chen, Y.,
et al. (2006) CFTR regulates phagosome acidification in
macrophages and alters bactericidal activity. Nat Cell Biol
8: 933–944.
Döring, G., Goldstein, W., Botzenhart, K., Kharazmi, A.,
Schiøtz, P.O., Høiby, N., and Dasgupta, M. (1986)
Elastase from polymorphonuclear leukocytes – a regula-
tory enzyme in immune complex disease. Clin Exp
Immunol 64: 597–605.
Döring, G., Frank, F., Boudier, C., Herbert, S., Fleischer, B.,
and Bellon, G. (1995) Cleavage of lymphocyte surface
antigens CD2, CD4, and CD8 by polymorphonuclear leu-
kocyte elastase and cathepsin G in patients with CF.
J Immunol 154: 4842–4850.
Döring, G., Knight, R., and Bellon, G. (2000) Immunology of
CF. In Cystic Fibrosis. Hodson, M.E., and Geddes, D.
(eds). London: Arnold, pp. 109–140.
Döring, G., Meisner, C., Stern, M., for the Flagella Vaccine
Trial Study Group (2007) A double-blind randomized
placebo-controlled phase III study of a Pseudomonas
aeruginosa flagella vaccine in CF patients. Proc Natl Acad
Sci USA 104: 11020–11025.
Engelhardt, J.F., Yankaskas, J.R., Ernst, S.A., Yang, Y.,
Marino, C.R., Boucher, R.C., et al. (1992) Submucosal
glands are the predominant site of CFTR expression in the
human bronchus. Nat Genet 2: 240–248.
Ferry, G., Lonchampt, M., Pennel, L., de Nanteuil, G., Canet,
E., and Tucker, G.C. (1997) Activation of MMP-9 by neu-
trophil elastase in an in vivo model of acute lung injury.
FEBS Lett 402: 111–115.
Goldstein, W., and Döring, G. (1986) Lysosomal enzymes
from polymorphonuclear leukocytes and proteinase inhibi-
tors in patients with CF. Am Rev Respir Dis 134: 49–56.
Grassmé, H., Gulbins, E., Brenner, B., Ferlinz, K., Sandhoff,
K., Harzer, K., et al. (1997) Acidic sphingomyelinase medi-
ates entry of N. gonorrhoeae into nonphagocytic cells. Cell
91: 605–615.
Grassmé, H., Jekle, A., Riehle, A., Schwarz, H., Berger, J.,
Sandhoff, K., et al. (2001) CD95 signaling via ceramide-
rich membrane rafts. J Biol Chem 276: 20589–20596.
Grassmé, H., Jendrossek, V., Riehle, A., von Kurthy, G.,
Berger, J., Schwarz, H., et al. (2003) Host defense against
Pseudomonas aeruginosa requires ceramide-rich mem-
brane rafts. Nat Med 9: 322–330.
Griese, M., Essl, R., Schmidt, R., Rietschel, E., Ratjen, F.,
Ballmann, M., et al. (2004) Pulmonary surfactant, lung
function, and endobronchial inflammation in CF. Am J
Respir Crit Care Med 170: 1000–1005.
Haggie, P.M., and Verkman, A.S. (2007) CF transmembrane
conductance regulator-independent phagosomal acidifica-
tion in macrophages. J Biol Chem 282: 31422–31428.
Hannah, S., Mecklenburgh, K., Rahman, I., Bellingan, G.J.,
Greening, A., Haslett, C., and Chilvers, E.R. (1995)
Hypoxia prolongs neutrophil survival in vitro. FEBS Lett
372: 233–237.
Hartl, D., Latzin, P., Hordijk, P., Marcos, V., Rudolph, C.,
Woischnik, M., et al. (2007) Cleavage of CXCR1 on neu-
trophils disables bacterial killing in CF lung disease. Nat
Med 13: 1423–1430.
Henriksen, P.A., Hitt, M., Xing, Z., Wang, J., Haslett, C.,
Riemersma, R.A., et al. (2004) Adenoviral gene delivery of
elafin and secretory leukocyte protease inhibitor attenuates
NF-kappa B-dependent inflammatory responses of human
endothelial cells and macrophages to atherogenic stimuli.
J Immunol 172: 4535–4544.
Heyworth, P.G., Cross, A.R., and Cornutte, J.T. (2003)
Chronic granulomatous disease. Curr Opin Immunol 15:
578–584.
Hirche, T.O., Crouch, E.C., Espinola, M., Brokelman, T.J.,
Mecham, R.P., DeSilva, N., et al. (2004) Neutrophil serine
proteinases inactivate surfactant protein D by cleaving
within a conserved subregion of the carbohydrate recogni-
tion domain. J Biol Chem 279: 27688–27698.
Joo, N.S., Lee, D.J., Winges, K.M., Rustagi, A., and Wine,
J.J. (2004) Regulation of antiprotease and antimicrobial
© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216

protein secretion by airway submucosal gland serous cells.
J Biol Chem 279: 38854–38860.
Kerem, B., Rommens, J.M., Buchanan, J.A., Markiewicz, D.,
Cox, T.K., Chakravarti, A., et al. (1989) Identification of the
CF gene: genetic analysis. Science 245: 1073–1080.
Kolesnick, R.N., Goni, F.M., and Alonso, A. (2000) Compart-
mentalization of ceramide signaling: physical foundations
and biological effects. J Cell Physiol 184: 285–300.
Lehrer, R.I., and Ganz, T. (2002) Defensins of vertebrate
animals. Curr Opin Immunol 14: 96–102.
McKenney, D., Tibbetts, K.L., Wang, Y., Murthy, V., Ulrich,
M., Döring, G., et al. (1999) Broadly protective vaccine for
Staphylococcus aureus based on an in vivo-expressed
antigen. Science 284: 1523–1527.
Matsui, H., Grubb, B.R., Tarran, R., Randell, S.H., Gatzy,
J.T., Davis, C.W., and Boucher, R.C. (1998) Evidence for
periciliary liquid layer depletion, not abnormal ion compo-
sition, in the pathogenesis of CF airways disease. Cell 95:
1005–1015.
Matsui, H., Verghese, M.W., Kesimer, M., Schwab, U.E.,
Randell, S.H., Sheehan, J.K., et al. (2005) Reduced three-
dimensional motility in dehydrated airway mucus prevents
neutrophil capture and killing bacteria on airway epithelial
surfaces. J Immunol 175: 1090–1099.
Matsui, H., Wagner, V.E., Hill, D.B., Schwab, U.E.,
Rogers, T.D., Button, B., et al. (2006) A physical linkage
between CF airway surface dehydration and Pseudomo-
nas aeruginosa biofilms. Proc Natl Acad Sci USA 103:
18131–18136.
Nathan, C. (2006) Neutrophils and immunity: challenges and
opportunities. Nat Rev Immunol 6: 173–182.
Notarangelo, L., Casanova, J.L., Conley, M.E., Chapel, H.,
Fischer, A., Puck, J., et al. (2006) Primary immunodefi-
ciency diseases: an update from the International Union of
Immunological Societies Primary Immunodeficiency Dis-
eases Classification Committee meeting in Budapest 2005.
J Allergy Clin Immunol 117: 883–896.
Pier, G.B. (2000) Role of the CF transmembrane conduc-
tance regulator in innate immunity to Pseudomonas aerugi-
nosa infections. Proc Natl Acad Sci USA 97: 8822–8828.
Riordan, J.R., Rommens, J.M., Kerem, B., Alon, N., Rozma-
hel, R., Grzelczak, Z., et al. (1989) Identification of the CF
gene: cloning and characterization of complementary DNA.
Science 245: 1066–1073.
Roghanian, A., Williams, S.E., Sheldrake, T.A., Brown, T.I.,
Oberheim, K., Xing, Z., et al. (2006) The antimicrobial/
elastase inhibitor elafin regulates lung dendritic cells and
adaptive immunity. Am J Respir Cell Mol Biol 34: 634–642.
Rommens, J.M., Iannuzzi, M.C., Kerem, B., Drumm, M.L.,
Melmer, G., Dean, M., et al. (1989) Identification of the CF
gene: chromosome walking and jumping. Science 245:
1059–1065.
Rubio, F., Cooley, J., Accurso, F.J., and Remold-O’Donnell,
E. (2004) Linkage of neutrophil serine proteases and
decreased surfactant protein-A (SP-A) levels in inflamma-
tory lung disease. Thorax 59: 318–323.
Sabra, W., Lunsdorf, H., and Zeng, A.P. (2003) Alterations
in the formation of lipopolysaccharide and membrane
vesicles on the surface of Pseudomonas aeruginosa PAO1
under oxygen stress conditions. Microbiology 149: 2789–
2795.
© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216
Cystic fibrosis and innate immunity 215
Segal, A.J. (2005) How neutrophils kill microbes. Annu Rev
Immunol 23: 197–223.
Seksek, O., Biwersi, J., and Verkman, A.S. (1996) Evidence
against defective trans-Golgi acidification in CF. J Biol
Chem 271: 15542–15548.
Selsted, M.E., and Quellette, A.J. (2003) Mammalian
defensins in the antimicrobial immune response. Nat
Immunol 6: 551–557.
Smith, J.J., Travis, S.M., Greenberg, E.P., and Welsh, M.J.
(1996) Cystic fibrosis airway epithelia fail to kill bacteria
because of abnormal airway surface fluid. Cell 85: 229–236.
Sommerhoff, C.P., Nadel, J.A., Basbaum, C.B., and
Caughey, G.H. (1990) Neutrophil elastase and cathepsin G
stimulate secretion from cultured bovine airway gland
serous cells. J Clin Invest 85: 682–689.
Sousa, S., Ulrich, M., Bragonzi, A., Burke, M., Worlitzsch, D.,
Leitão, J.H., et al. (2007) Virulence of Burkholderia cepacia
complex strains in gp91phox-/- mice. Cell Microbiol 9: 2817–
2825.
Suzuki, T., Moraes, T.J., Vachon, E., Ginzberg, H.H., Huang,
T.-T., Matthay, M.A., et al. (2005) Proteinase-activated
receptor-1 mediates elastase-mediated apoptosis of
human lung epithelial cells. Am J Respir Cell Mol Biol 33:
231–247.
Teichgräber, V., Ulrich, M., Riethmüller, J., Grassme, H.,
Wilker, B., De Oliveira-Munding, C.C., et al. (2008) Cera-
mide accumulation mediates inflammation, cell death and
infection susceptibility in CF. Nat Med 14: 382–391.
Tunney, M.M., Field, T.R., Moriarty, T.F., Patrick, S., Döring,
G., Muhlebach, M.S., et al. (2008) Detection of anaerobic
bacteria in high numbers in sputum from patients with
cystic fibrosis. Am J Respir Crit Care Med 177: 995–1001.
Ulrich, M., Herbert, S., Berger, J., Bellon, G., Louis, D.,
Münker, G., and Döring, G. (1998) Localization of Staphy-
lococcus aureus in infected airways of patients with CF and
in a cell culture model of S. aureus adherence. Am J Respir
Crit Care Med 19: 83–91.
Ulrich, M., Bastian, M., Cramton, S.E., Ziegler, K., Pragman,
A.A., Bragonzi, A., et al. (2007) The staphylococcal respi-
ratory response regulator SrrAB induces ica gene tran-
scription and PIA expression, protecting Staphylococcus
aureus from neutrophil killing under anaerobic growth con-
ditions. Mol Microbiol 65: 1276–1287.
Vandivier, R.W., Fadok, V.A., Hoffmann, P.R., Bratton, D.L.,
Penvari, C., Brown, K.K., et al. (2002) Elastase-mediated
phosphatidylserine receptor cleavage impairs apoptotic
cell clearance in CF and bronchiectasis. J Clin Invest 109:
661–670.
Verkman, A.S., Song, Y., and Thiagarajah, J.R. (2003) Role
of airway surface liquid and submucosal glands in CF lung
disease. Am J Physiol Cell Physiol 284: C2–C15.
Voynow, J.A., Young, L.R., Wang, Y., Horger, T., Rose, M.C.,
and Fischer, B.M. (1999) Neutrophil elastase increases
MUC5AC mRNA and protein expression in respiratory epi-
thelial cells. Am J Physiol 276: L835–L843.
Walsh, D.E., Greene, C.M., Carroll, T.P., Taggart, C.C., Gal-
lagher, P.M., O’Neill, S.J., and McElvaney, N.G. (2001)
Interleukin-8 up-regulation by neutrophil elastase is medi-
ated by MyD88/IRAK/TRAF-6 in human bronchial epithe-
lium. J Biol Chem 276: 354–394.
Whitchurch, C.B., Tolker-Nielsen, T., Ragas, P.C., and
 216 G. Döring and E. Gulbins
Mattick, J.S. (2002) Extracellular DNA required for bacterial
biofilm formation. Science 295: 1487.
Worlitzsch, D., Tarran, R., Ulrich, M., Schwab, U., Cekici, A.,
Meyer, K.C., et al. (2002) Reduced oxygen concentrations
in airway mucus contribute to the early and late pathogen-
esis of Pseudomonas aeruginosa CF airways infection.
J Clin Invest 109: 317–325.
Zahm, J.M., Gaillard, D., Dupuit, F., Hinnrasky, J., Porteous,
D., Dorin, J.R., et al. (1997) Early alterations in airway
mucociliary clearance and inflammation of the lamina
propria in CF mice. Am J Physiol 272: C853–C859.
Zhang, B., Hirahashi, J., Cullere, X., and Mayadas, T.N.
(2003) Elucidation of molecular events leading to neutro-
phil apoptosis following phagocytosis: cross-talk between
caspase 8, reactive oxygen species, and MAPK/ERK acti-
vation. J Biol Chem 278: 28443–28454.
© 2008 Blackwell Publishing Ltd, Cellular Microbiology, 11, 208–216