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Regular Articles |
From the Department of Trauma Surgery,*
University of
Freiburg Medical School, Freiburg/Breisgau, Germany, and the Department
of Pathology,
University of Michigan Medical
School, Ann Arbor, Michigan
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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) in bronchoalveolar lavage fluids, and increased
expression of lung vascular intercellular adhesion molecule-1 (ICAM-1).
Complement depletion or treatment with anti-C5a abolished all evidence
of enhanced lung injury in septic animals. When stimulated in
vitro, bronchoalveolar lavage macrophages from septic
animals had greatly enhanced CXC chemokine responses as compared with
macrophages from sham-operated animals or from septic animals that had
been complement depleted. These data indicate that the septic state
causes priming of lung macrophages and suggest that enhanced lung
injury in the septic state is complement dependent and related to
increased production of CXC chemokines.
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, or IL-6.1,2,8-10
This phenomenon has been termed the systemic inflammatory response
syndrome. This condition appears to place organs (liver, lung, and
kidneys) at risk of injury and failure. It has been postulated that
during sepsis the lung is especially susceptible to injury in the
presence of a direct intrapulmonary insult (the so-called second
hit),11
such as ischemia, blunt thoracic injury,
intrapulmonary presence of bacteria, or ventilator-induced pulmonary
injury, to name only a few examples.
Why the liver and kidneys also become targets of injury during sepsis
is poorly understood. Although a septic-like state has been induced experimentally by infusion of LPS or live bacteria, cecal ligation/puncture (CLP) in rodents seems to mimic many features of the septic state in humans. Animals develop progressive bacteremia, appearance of multiple cytokines and chemokines in plasma, hypermetabolism, fever, and other clinical features similar to those found in humans with sepsis.8 In the present study CLP was studied in rats. The experimental data indicate that during sepsis a direct intrapulmonary insult (induced by LPS or deposition of immune complexes) results in augmented accumulation of neutrophils, higher levels of CXC chemokines, and evidence of enhanced lung injury. The studies also indicate that these events are complement dependent and associated with a priming of lung macrophages.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Male Long-Evans specific-pathogen-free rats (275 to 300 mg;
Harlan, Indianapolis, IN) were used in all studies. Anesthesia was
induced by intraperitoneal administration of ketamine (20 mg/100 mg
body weight). After shaving the abdomen and application of a topical
disinfectant, a 2-cm midline incision was made, and the cecum was
identified and ligated below the ileocecal valve, with care being taken
not to occlude the bowel. The cecum was then subjected to a single
through-and-through perforation with a 23-gauge needle. After
repositioning the bowel, the abdominal incision was closed in
layers with plain gut surgical suture 4-0 (Ethicon, Somerville, NJ)
and metallic clips. Sham animals underwent the same procedure except
for ligation and puncture of the cecum. Before and after surgery,
animals had unlimited access to food and water. After an interval of up
to 36 hours, lung injury was induced either by intratracheal
instillation of 100 µg of bacterial LPS (Sigma Chemical Co., St.
Louis, MO) or by intrapulmonary deposition of IgG immune complexes as
described elsewhere.12,13
For the latter, 10 mg of bovine
serum albumin (BSA; Sigma Chemical Co.) were given intravenously (with
trace amounts, 0.5 µCi, of 125I-labeled BSA) after
intratracheal administration of 2.5 mg of polyclonal rabbit anti-BSA
IgG (Organon Teknika Corp., West Chester, PA) in a total volume of 300
µl. In the LPS model of lung injury, 125I-labeled BSA was
also given intravenously. Animals were sacrificed 6 hours after
instillation of LPS and 4 hours after IgG immune-complex-induced
alveolitis. These intervals selected for sacrifice represent times of
peak lung injury, as determined in previous
experiments.12,13
The pulmonary circulation was then
flushed with 10 ml of phosphate-buffered saline (PBS). Lung vascular
permeability indices were determined by the ratio of extravasated
125I-labeled BSA present in lung parenchyma to the amount
present in 1.0 ml of blood obtained from the posterior vena cava at the
time of sacrifice. In other sets of animals, bronchoalveolar lavage
(BAL) fluids were collected at the times indicated, using repetitive
(three times) instillation and withdrawal of 5 ml of saline via an
intratracheal cannula. After addition of a protease inhibitor cocktail
(1 µg/ml leupeptin, 1 µg/ml aprotinin, 10 µg/ml trypsin
inhibitor, and 1 µg/ml pepstatin), samples were centrifuged at 3000
rpm for 10 minutes, and supernatant fluids were subsequently used for
chemokine quantitation. Cell pellets from centrifuged BAL fluids were
assessed for differential cell counts.14
Animals receiving
antibody treatment were injected intravenously at the time of the CLP
procedure with 300 µg of goat anti-rat C5a antibody (purified and
characterized as described previously15
) or with 300 µg
of preimmune goat IgG, similar to an earlier protocol.15
For another set of experiments, anti-C5a was administered at 6, 12, 18,
or 24 hours after CLP. Complement depletion was performed by three
serial intraperitoneal injections of 25 U of purified cobra venom
factor (CVF) at 12-hour intervals as described
elsewhere.12
In some animals, a carotid artery catheter
(PE-50, Becton-Dickinson Co., Sparks, MD) was placed through an
anterior cervical incision and, after subcutaneous tunneling,
externalized at the posterior neck. The animals were allowed to recover
for 24 hours with free access to food and water and then subjected to
the CLP procedure. Blood samples were obtained in 12-hour intervals and
subsequently used for measuring serum hemolytic complement activity (CH
50 assay) as reported.15
For each group,
n 
4.
Measurement of Lung Vascular Intercellular Adhesion Molecule-1 and Quantitation of Cytokines
Quantitation of lung vascular intercellular adhesion molecule
(ICAM)-1 was determined by binding of an 125I-labeled
antibody to ICAM-1 (1A29, a gift from Dr. M. Miyasaka, Osaka
University, Japan) as described in previous studies.16
The
ELISA techniques for measurement of cytokine-induced neutrophil
chemoattractant (CINC) and macrophage inflammatory protein (MIP)-2 in
rat BAL fluids are described elsewhere.14
Cell culture
supernatants and BAL fluids were evaluated for TNF- activity using a
standard WEHI cell cytotoxicity assay as previously
reported.17
Isolation and In Vitro Stimulation of Alveolar Macrophages
Alveolar macrophages were isolated at the times indicated by BAL of lungs of anesthetized rats of CLP or sham-treated groups. After centrifugation of lavage fluids, cells were resuspended in culture medium and plated into 48 microtiter well plates (Corning, New York, NY) at a concentration of 1 x 106 cells/well. Immune complexes were formed by addition to anti-BSA of BSA at the point of antigen equivalence. Complexes were added to wells over a concentration range of 0.8 to 100 µg/ml. After an incubation period of 4 hours at 37°C, supernatant fluids from macrophages were collected and evaluated for chemokine content.
Statistical Analysis
All values were expressed as mean ± SEM. Significance was assigned where P < 0.05. Data sets were analyzed employing one-way analysis of variance, and individual group means were then compared with the Student-Newman-Keuls multiple comparison test. To calculate percentage change between groups, values obtained from negative controls were subtracted from each data point.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Animals subjected to CLP developed the typical clinical signs of
sepsis: decreased physical activity, piloerection, cessation in
grooming behavior, glazed eyes with crusting exudate, diarrhea,
lethargy, and occasionally death. The overall mortality rate 36 hours
after the CLP procedure under the conditions used (two punctures with a
23-gauge needle) was 12%. Bacteremia was regularly detectable in blood
samples drawn 36 hours after CLP (data not shown). To assess the extent
of systemic complement activation, serial blood samples were collected
in 12-hour intervals via an intra-arterial catheter and examined for
serum complement hemolytic activity (using sensitized sheep
erythrocytes). Absorbance values (541 nm) were determined and values
expressed as percentage of the hemolytic activity present in serum
obtained before CLP. At 12 hours after induction of sepsis, complement
hemolytic activity was reduced to 86.3 ± 4.4%
(P < 0.05) of that found in pre-sepsis serum,
with a further decline at 24 hours and 36 hours, to 69.5 ± 5.7%
and 68.8 ± 5.6% (for both, P < 0.05),
respectively (Figure 1)
. Thus, consumptive
depletion of complement occurred in animals undergoing CLP.
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The extent of lung injury was assessed in septic and nonseptic
animals after a direct intrapulmonary insult (4 hours after deposition
of IgG immune complexes or 6 hours after airway instillation of LPS).
Lung vascular permeability relative to time and experimental
manipulation is shown in Figure 2
. The
vascular permeability index, as determined by the ratio of
125I-labeled albumin in lung and blood, rose from a value
of 0.21 ± 0.02 in sham-operated animals to 0.46 ± 0.06 in
nonseptic rats with immune-complex-induced alveolitis. Compared with
the sham control groups, there was no detectable evidence of increased
lung permeability 36 hours after onset of CLP in the absence of a
primary intrapulmonary insult (CLP, 36 hours, Figure 2
), the
permeability index being 0.25 ± 0.02. Lung injury caused by a
direct intrapulmonary insult (immune complexes) 12 and 24 hours after
induction of sepsis was associated with permeability indices of
0.39 ± 0.03 and 0.41 ± 0.01, respectively, which
statistically were not different from the values in the nonseptic group
receiving immune complexes alone. However, in CLP animals at 36 hours,
immune-complex-induced lung injury resulted in a permeability index
increase by 106% to a value of 0.73 ± 0.1
(P < 0.05 when compared with injury caused by
immune complex deposition alone). Increased lung injury was also found
in septic (CLP) animals receiving airway instillation of LPS.
Intratracheal instillation of 100 µg of LPS into normal animals
resulted in a permeability index of 0.34 ± 0.02 (as compared with
the value obtained in sham lungs receiving PBS, 0.22 ± 0.02). At
36 hours after CLP in animals receiving LPS, the permeability index
rose by 105% (P < 0.05) to a value of
0.47 ± 0.02 (Figure 2)
. Thus, in animals receiving a direct
intrapulmonary insult, sepsis caused an increased lung permeability
index.
|
. In the presence of sepsis, the
direct intrapulmonary insult caused the vascular leakage index to rise
to 0.74 ± 0.06, but the value fell by 50%
(P < 0.05) to 0.47 ± 0.04 in
complement-depleted rats and by 57% (P < 0.05)
to 0.43 ± 0.08 in rats treated with anti-C5a (Figure 3)
. As the
vascular leakage indices of the complement-depleted and
anti-C5a-treated groups did not statistically differ from the values
obtained in the immune-complex-injured group in the absence of sepsis,
the complement-depleting or blocking procedures completely abolished
the injury-enhancing effects of sepsis in lung. To investigate when
during sepsis in vivo blockade of C5a (with antibody) would
be effective, treatment with anti-C5a was done 6, 12, 18, and 24 hours
after CLP. The results are shown in Figure 4
.
In this set of experiments the intra-alveolar deposition of IgG immune
complexes resulted in a lung vascular permeability index of 0.47
± 0.01, whereas the same direct intrapulmonary insult in septic
animals caused a value of 0.76 ± 0.05. As expected, intravenous
administration of anti-C5a at the time of CLP resulted in a complete
abrogation of the enhancing effect of sepsis on direct intrapulmonary
insult, with a vascular leakage index of 0.49 ± 0.07. Systemic
infusion of anti-C5a at 6 hours proved to be as effective in inhibiting
the priming effect of CLP on lung, with a value of 0.49 ± 0.02.
Administration of anti-C5a at 12 hours led to significant but somewhat
reduced protection from lung injury enhancement, with a permeability
index of 0.57 ± 0.02 (P < 0.05, when
compared with animals receiving both hits in the presence of
nonspecific IgG). Delaying treatment with anti-C5a to 18 hours and 24
hours after CLP did not demonstrate protective effects in lung injury,
with lung vascular leakage indices of 0.69 ± 0.02 and 0.7 ±
0.06 (for both, the P value was not significant when
compared with the values in the positive control groups treated with
preimmune IgG), respectively. Thus, C5a blockade was effective in
suppressing the inflammatory-enhancing effects of sepsis only in the
early stages of the sepsis model.
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As lung injury in the immune complex alveolitis model is
neutrophil dependent,12
we also investigated neutrophil
content in BAL fluids from animals with CLP alone or CLP together with
a direct lung injury induced by deposition of immune complexes. The
enhancement of lung vascular permeability in septic animals receiving a
direct intrapulmonary insult (described in Figures 2 and 3
) was also
reflected by increased numbers of neutrophils in BAL fluids (Figure 5)
. The neutrophil counts (expressed as
x 104/ml BAL fluid) were 24.3 ± 1.79/ml in
sham-operated animals and 17.7 ± 1.20/ml in CLP animals at 36
hours. Immune complex deposition in lungs of otherwise normal animals
caused BAL neutrophil values to rise to 40.7 ± 2.96/ml. However,
36 hours after CLP and 4 hours after immune complex deposition, there
was a 3.4-fold increase in neutrophil content in BAL fluids rising to a
value of 95.0 ± 16.5/ml (P < 0.05 when
compared with BAL neutrophils after immune complex deposition in
nonseptic animals), representing a 340% increase. In a companion set
of experiments, BAL neutrophils were assessed in animals receiving
intravenous administration of anti-C5a or preimmune goat IgG, each
being given as 300 µg before CLP and induction of lung injury.
Whereas the neutrophil content in BAL fluids from animals treated with
preimmune goat IgG followed by CLP and immune-complex-induced
alveolitis was 205 ± 22.6/ml, treatment of rats under the same
protocol with anti-C5a led to a dramatic drop to a value of 101 ±
23.5/ml (P < 0.05). There was no statistically
significant difference between BAL levels of neutrophils in nonseptic
animals with immune complex deposition and in anti-C5a-treated septic
animals with co-existent immune-complex-induced alveolitis
(P > 0.05). Thus, CLP-induced sepsis at 36
hours was associated with greatly increased accumulation of neutrophils
after exposure of lung to a direct insult. This, in turn, was followed
by a significant increase in lung vascular permeability, both effects
being completely suppressible by complement depletion or systemic
treatment with anti-C5a.
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Morphological evaluation of lungs was performed. In keeping
with vascular leakage indices and BAL neutrophil content (Figures 2 to 4)
, normal lungs and lungs from CLP animals at 36 hours had normal
morphological features (Figure 6, A and B
,
respectively). Lungs directly injured with immune complexes in
nonseptic animals had the expected evidence of hemorrhage and
neutrophil accumulation (Figure 6C)
, whereas in the co-presence of CLP,
the intensity of hemorrhage and neutrophil influx was accentuated
(Figure 6D)
.
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To explore the causes for enhanced lung injury and increased
neutrophil accumulation in septic animals subjected to a direct lung
insult (immune complex deposition or LPS instillation), BAL fluids were
assessed for content of the CXC chemokines MIP-2 and CINC. These CXC
chemokines were selected because, in the setting of lung injury induced
by LPS or deposition of IgG immune complexes, these chemokines have
been shown to play a major role in neutrophil recruitment and
development of lung injury.13,14
MIP-2 levels were
undetectable in BAL fluids from sham-operated animals and from animals
36 hours after CLP alone (Figure 7A)
. In BAL
fluids, direct lung injury induced by immune complex deposition
resulted in MIP-2 content of 301 ± 30.9 ng/ml. Lungs of animals
injured with immune complexes 36 hours after CLP had significantly
increased (38%, P < 0.05) levels of MIP-2 in BAL
fluids, 414.9 ± 12.5 ng/ml. This enhancement above and beyond
levels found in immune-complex-injured lung of nonseptic animals was
completely abolished when animals were pretreated with anti-C5a,
causing MIP-2 levels to fall to 294 ± 15.9 ng/ml. Expression of
CINC in BAL fluids was very low in sham-operated animals (12 ±
6.9 ng/ml) and undetectable in BAL fluids of CLP animals at 36 hours.
In lungs from nonseptic rats after IgG immune complex deposition, CINC
levels rose to 2060 ± 162 ng/ml. In animals 36 hours after CLP
and with immune-complex-induced lung injury, CINC levels in BAL fluids
were further increased (by 81%, P < 0.05) to
3726 ± 497 ng/ml (Figure 7B)
. In companion rats, treatment with
anti-C5a at the time of induction of sepsis also prevented the
increased levels of CINC, reducing the values to 2131.2 ± 190.4
ng/ml BAL fluid.
|
. When LPS was instilled
into otherwise normal lungs, MIP-2 and CINC levels rose to 139 ±
6.2 and 1906 ± 128 ng/ml, respectively. The combination of LPS
instillation into CLP animals treated intravenously with 300 µg of
preimmune IgG caused increases in MIP-2 and CINC levels by 25% and
88%, respectively. In animals treated with anti-C5a, these increases
in levels of CXC chemokines were diminished, such that the levels were
the same as those found in otherwise normal rats with airway
instillation of LPS. Thus, in CLP rats undergoing direct injury by LPS
instillation, there was a C5a-dependent increase in MIP-2 and CINC,
similar to findings in CLP animals undergoing direct lung injury by IgG
immune complex deposition (Figure 7)
.
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in BAL Fluids
TNF- is known to be expressed in immune-complex-induced lung
injury and to regulate the expression of vascular ICAM-1 in the
inflamed lung.16
TNF- levels were measured in BAL
fluids in the presence or absence of CLP-induced sepsis and in the
presence or absence of a direct lung insult (immune complexes). In
agreement with other reports,18,19
CLP alone did not cause
increased levels of TNF- in BAL fluids at 24 or 36 hours. The levels
of TNF- were similar to those in sham-operated animals, with values
consistently below 1.5 ng/ml. The presence or absence of sepsis did not
affect the levels of TNF- in BAL fluids after direct lung injury.
Immune-complex-induced lung injury in the presence or absence of sepsis
was associated with BAL levels of TNF- of 70.6 ± 4.7 and
60.7 ± 5.10 ng/ml, respectively (P value not
significant). Thus, the enhancement of lung injury in septic rats
cannot be linked to changes in BAL levels of TNF-.
Enhanced Expression of Lung Vascular ICAM-1 in Lungs of Septic Animals
As lung vascular ICAM-1 is known to be involved in intrapulmonary
recruitment of neutrophils, in a variety of conditions, we measured
this adhesion molecule by lung vascular fixation of
125I-labeled anti-ICAM-1 (1A29, a murine IgG1).
The binding index was calculated by the ratio of radioactivity in lung
to radioactivity in blood, adjusted by subtraction of nonspecific
binding values of irrelevant 125I-labeled control
IgG1 (MOPC-21, Sigma Chemical Co.). Results are shown in
Figure 7C
. Sham-operated and animals with CLP alone (at 36 hours)
demonstrated very small binding indices, 0.005 ± 0.002 and
0.004 ± 0.002, respectively (P value not
significant). After immune-complex-induced lung injury in otherwise
normal animals, the binding index rose nearly 10-fold to a value of
0.033 ± 0.007. In septic animals with lungs also subjected to the
immune complex insult, there was a further increase (by 118%,
P < 0.05) in the binding index for ICAM-1, rising to a
value of 0.066 ± 0.006. Thus, sepsis sets the stage for
accentuated up-regulation of lung vascular ICAM-1 in the presence of a
direct intrapulmonary insult, although this is not reflected in changes
in BAL levels of TNF-. Increased lung vascular ICAM-1 expression is
associated with intensified neutrophil recruitment and lung injury.
Generation of CXC Chemokines by Alveolar Macrophages from Septic and Nonseptic Rats
Alveolar macrophages were obtained from sham-operated and from
septic rats (36 hours after CLP) and placed into tissue culture wells,
with 1 x 106
cells/well. The resulting monolayers
were then stimulated for 4 hours with increasing doses of IgG immune
complexes (0.8 to 100 µg/ml). Cell culture supernatant fluids were
assessed for chemokine content. As expected, there was dose-dependent
expression of MIP-2 and CINC in cultures of alveolar macrophages
obtained from sham-operated animals (Figure 8)
. Macrophages from septic animals
consistently demonstrated significantly higher levels of MIP-2 and CINC
production when compared with macrophages from nonseptic animals,
indicating that priming of these cells had occurred in vivo
during sepsis (Figure 8, A and B)
. These data are consistent with the
results obtained in BAL fluids (Table 1
; Figure 7
). Macrophages from
septic animals that had been complement depleted failed to show
enhanced in vitro generation of CXC chemokines (Figure 8)
,
supporting the concept that complement is required for the in
vivo priming process. These results extend the in vivo
findings of enhanced pulmonary chemokine production in
complement-intact, septic animals that have received a direct
intrapulmonary insult. Complete abrogation of this enhancement occurred
with C5a blockade.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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. The reason for
enhanced levels of these CXC chemokines appears to be linked to the
fact that the septic state primes alveolar macrophages for exaggerated
chemokine responses. The in vivo priming effect of sepsis on
lung was time dependent, as the increase in lung permeability index and
in neutrophil accumulation were not detectable until 36 hours after
CLP. An accentuated lung inflammatory response continued beyond 36
hours, as enhancement of lung injury after CLP was also seen at 48
hours (data not shown). All of these events were complement dependent
and specifically required the role of C5a. In the face of complement
depletion or blockade of C5a, all increments in chemokines, in
neutrophil accumulation, and in lung vascular leakage of albumin in
septic animals subjected to a direct intrapulmonary insult were
abrogated. It is perhaps not too surprising that a role for C5a was
found, as C5a plays a role in vivo in lung vascular ICAM-1
up-regulation of rats undergoing acute inflammatory injury after
intrapulmonary deposition of IgG immune complexes.15,16
Furthermore, in acute lung injury after systemic activation of
complement, C5a has been shown to be critically required for
up-regulation of lung vascular P-selectin and subsequent recruitment of
neutrophils.20
It seems likely that, in CLP-induced
sepsis, C5a is produced locally (in the peritoneum) or systemically (in
the plasma compartment), as described by Nakae et al,21
and that it ultimately gains access to the distal airway compartment
where it reacts with C5a receptors to bring about macrophage priming.
These cells then become hyperresponsive to a subsequent stimulus. In
the IgG immune-complex-induced lung injury model, a compartmentalized
role for C5a has been demonstrated by the superior protective effects
of intratracheally administered anti-C5a when compared with intravenous
delivery of anti-C5a.15
In the current report we
hypothesize that C5a present in the plasma gains access to lung
macrophages. This is supported by the finding of progressive systemic
complement activation during sepsis (Figure 1)
and by the ability of
intravenously administered anti-C5a to block the priming phenomenon in
lung during the first 12 hours of sepsis (Figure 4)
.
There is precedent for the role of complement in the septic syndrome
and in the response to the presence of bacteria or LPS. Dogs
genetically deficient in C3 showed decreased clearance of LPS and
enhanced shock and organ damage after infusion of LPS.22
LPS-induced lethality was greatly increased in C4-deficient guinea
pigs,23
in C6-deficient rabbits,24
and in
mice that were made genetically deficient in C3 or C4.25
Also, C3-deficient mice had delayed clearance of group B streptococci
and poorer survival.26
Curiously, C5-deficient mice seem
resistant to the effects of LPS, a finding that was associated with a
diminished TNF- response.27
In septic humans,
nonsurvivors had significantly higher blood levels of
C5a,21
suggesting the failure to regulate generation of
C5a. In LPS-infused rats, C5a blockade by antibody attenuated changes
in hemodynamic parameters,28
protected against secondary
lung injury, and improved short-term survival.29,30
C5-deficient mice had improved survival times after CLP.31
Finally, in LPS-infused pigs, blockade of C5a reduced blood levels of
IL-6 by 75%, although it was not apparent that other parameters were
affected by this treatment.32
All of these data suggest
that, after LPS infusion and in some bacteremic and septic states,
complement activation products may be produced in excess (or may not be
adequately regulated), contributing to undesirable outcomes.
It was somewhat surprising to note that TNF- levels in BAL fluids
obtained from septic rats with immune-complex-injured lungs were the
same as in immune-complex-induced lungs from sham-operated rats. This
was in contrast to increased neutrophil accumulation occurring in
septic rats (Figure 5)
, as in earlier studies using the immune complex
model a linkage was established between BAL levels of TNF- and
up-regulation of lung vascular ICAM-1.16
However, recently
it has been shown that, when TNF- is present together with the
complement-derived membrane attack complex (C5b-9), synergistic
up-regulation in vitro of endothelial ICAM-1 and E-selectin
occurs.33
It is possible in the context of the in
vivo model of lung injury in septic rats that such an interaction
might be occurring, but this remains to be demonstrated. The in
vivo relationship between C5a, TNF-, and up-regulation of
vascular ICAM-1 in the lung15
might also suggest
synergistic interactions in vivo between these mediators and
lung vascular ICAM-1.
The priming of lung macrophages obtained from septic rats appears to be
linked to a requirement for C5a based on the data in Figure 8
. Whether
C5a can per se prime alveolar macrophages has not been
evaluated. An alternate possibility might be that in septic rats there
has been a C5a-dependent release in lung of cytokines such as TNF-
or IL-1, both of which are known to have the ability to prime
phagocytic cells for enhanced responses to other stimuli. In the
priming process biochemical events such as tyrosine phosphorylation and
NF-
B activation are thought to be involved.34
Again,
whether C5a can induce tyrosine phosphorylation of proteins or whether
it can cause NF-B activation in the lung remains to be determined.
The priming of lung macrophages in the CLP model in rats as well as
enhanced injury after a direct lung insult appear to be dependent on
C5a. The advantage of direct blockade of C5a over other blocking
strategies such as with anti-C5 is that anti-C5a may preserve the
protective roles of C5b-9 in bacterial lysis. Protection against lung
injury afforded by infusion of anti-C5a occurred even when infusion of
anti-C5a was delayed (Figure 4)
. This might be relevant in septic
humans, providing a longer therapeutic window.
|
Acknowledgements |
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|
Footnotes |
|---|
Supported by National Institutes of Health grants GM-29507 and HL-31963.
Accepted for publication January 8, 1999.
|
References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
regulates in vivo intrapulmonary expression of ICAM-1. Am J Pathol 1993, 142:1739-1749[Abstract]
, IL-1ß, and Ia (I-A) mRNA expression during peritonitis is site dependent. J Surg Res 1993, 54:426-430[Medline]
induced endothelial cell expression of E-selectin and ICAM-1. J Immunol 1995, 155:1434-1441[Abstract]
, and granulocyte/macrophage-colony stimulating factor: correlation with priming of the respiratory burst. Biochim. Bioophys. Acta 1997, 1355:343-352
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J. C. Parker and M. I. Townsley Evaluation of lung injury in rats and mice Am J Physiol Lung Cell Mol Physiol, February 1, 2004; 286(2): L231 - L246. [Abstract] [Full Text] [PDF]
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A. M. Chinnaiyan, M. Huber-Lang, C. Kumar-Sinha, T. R. Barrette, S. Shankar-Sinha, V. J. Sarma, |
