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Regular Articles |
From the Department of Pathology,*
University of
Michigan Medical School, Ann Arbor, Michigan; and the Department of
Pulmonary and Critical Care Medicine,
University of California, Los Angeles, California
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, and interferon-
. Strategies aimed at regulating these
inflammatory cytokines during clinical sepsis have been fraught with
limited or no success. One cellular
target of recent significance is the mast cell. Previous studies have
suggested that mast cells exert a beneficial role during experimental
septic peritonitis partly because of the ability of this cell to
quickly release preformed TNF-.3
TNF- is an
important component of the innate host defense system that manages the
lethal effects of intestinal bacteria invading the peritoneal
cavity.4
However, mast cells are also a source of many
other preformed (ie, histamine) and newly formed mediators such as
arachidonic acid metabolites and chemokines that may also positively
affect the outcome of sepsis.5
Stem cell factor (SCF) is a hematopoietic cytokine that triggers its
biological effects by binding to the c-kit
receptor.6
It is a primary cytokine involved in mast cell
activation7-10
and chemotaxis.11
SCF is
produced by stromal cells, notably the embryonic and adult
liver.12,13
It has multiple effects beyond mast cell
activation and these include acute erythroid expansion,
spermatogenesis, melanocyte development, gut motility, and response to
intestinal helminth infection. More recently, the chronic subcutaneous
administration of SCF to mice was shown to induce systemic mastocytosis
and, more importantly, enhance innate immunity and mouse survival after
cecal ligation and puncture (CLP) surgery.14
Although
increased mast cell numbers and TNF- release were critical for mouse
survival after CLP surgery, this previous study alluded to other
beneficial effects of SCF in this model.
Thus, the aim of the present study was to elucidate the effects of a
single intraperitoneal SCF treatment in the context of experimental
sepsis and CLP. To this end, we examined the effect of a single SCF
treatment on the response of mice to CLP surgery. Although a single SCF
pretreatment did not induce mastocytosis nor markedly increase TNF-
levels, it significantly improved mouse survival, promoted the release
of the chemokine MCP-1 and the regulatory cytokine IL-10, and
accelerated the hepatic nuclear expression of signal transducer and
activator of transcription-3 (STAT-3) after CLP surgery.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Specific pathogen-free CD-1 mice (6- to12-week-old females; Charles River Breeding Laboratories, Wilmington, MA) were used in the majority of the experiments. All mice were housed in specific pathogen-free conditions within the animal care facility at the University of Michigan (ULAM) until they were used in experiments. Specific pathogen-free C57BL/6J-Mgf SL-d/+ mast cell-deficient mice and appropriate controls (6-to 12-week-old females) were purchased from the Jackson Laboratories (Bar Harbor, ME) immediately before their use in separate experiments.
CLP Model
CLP has been used extensively in our laboratory.15 Briefly, all mice were anesthetized with an intraperitonal (i.p.) injection of 3 to 3.5 mg of ketamine HCl (Ketaset; Fort Dodge Laboratories, Overland Park, KS) followed by inhaled methoxyflurane (Metafane; Pitman-Moore Inc., Mundelein, IL) as needed. After swabbing the abdomen with 70% ethyl alcohol (EtOH), a 1- to 2-cm longitudinal incision was then applied to the lower right quadrant of the abdomen. The cecum was removed from the peritoneal cavity and the distal one-third was occluded with 3-0 silk suture, and punctured through and through with a 21-gauge needle. The cecum was then restored to the peritoneal cavity and the peritoneal incision was closed with surgical staples. All mice then were administered 1 ml of sterile saline via a subcutaneous injection to facilitate fluid resuscitation after CLP surgery. SCF, at a dose of 4 or 20 µg/kg (Preprotech, Rocky Hill, NJ), was injected into the peritoneal cavity at 2 hours before CLP.
In additional survival experiments, 0.5 ml of anti-MCP-1 antiserum was
injected i.p. into each mouse simultaneously with the SCF treatment.
Control mice received 0.5 ml of rabbit serum via an i.p. injection
concomitant with the SCF treatment. The biological half-life of the
anti-MCP-1 antiserum used in these experiments was 
36 hours.
Polyclonal anti-murine MCP-1 antiserum was raised by multiple-site
immunizations in New Zealand White rabbits with murine rMCP-1 (R&D
Systems, Minneapolis, MN), and this antiserum did not cross-react with
other CC and CXC chemokines in an enzyme-linked immunosorbent assay
(ELISA) and demonstrated neutralizing activity against murine rMCP-1 in
a chemotaxis assay.16
Murine Cytokine and Chemokine ELISAs
Murine IL-10, MCP-1, interferon-, IL-12, MIP-1, KC, MIP-2,
and SCF were quantified using a modification of a double-ligand method
as previously described.17
Briefly, flat-bottomed 96-well
microtiter plates (Nunc Immuno-Plate I 96-F; Roskilde, Denmark)
were coated with 50 µl/well of anti-mouse cytokine antibody (1
µg/ml in 0.6 mol/L NaCl, 0.26 mol/L
H3BO4, and 0.08 mol/L NaOH,
pH 9.6) for 16 hours at 4°C and then washed with wash buffer
containing phosphate-buffered saline (PBS), pH 7.5, and 0.05%
Tween-20. Nonspecific binding sites in each plate were blocked with 2%
bovine serum albumin (BSA) in PBS and incubated for 90 minutes at
37°C. Plates were rinsed four times with wash buffer and diluted
(neat and 1:10) peritoneal lavage fluids, serum, or cell-free
supernatants (50 µl) in duplicate were added to each plate and
incubated for 1 hour at 37°C. Plates were washed four times, followed
by the addition of 50 µl/well biotinylated rabbit antibodies against
the specific cytokines (3.5 µg/ml in PBS, pH 7.5, 0.05% Tween-20,
and 2% fetal calf serum), and plates incubated for 30 minutes at
37°C. After washing, streptavidin-peroxidase conjugate (Bio-Rad
Laboratories, Richmond, CA) was added and the plates were incubated for
30 minutes at 37°C. After washing again, chromagen substrate (Bio-Rad
Laboratories) was added. The plates were incubated at room temperature
to the desired extinction and the reaction terminated with 50 µl/well
of 3 mol/L H2SO4 solution.
Plates were read at 490 nm in an ELISA reader. Standards were one-half
log dilutions of lipopolysaccharide (LPS)-free recombinant murine
cytokines (R&D Systems or Preprotech) from 1 pg/ml to 100 ng/ml.
This ELISA method consistently detected murine cytokine concentrations
>25 pg/ml and ELISA specificity was confirmed for each cytokine and
chemokine measured. Because many of the mice were hypotensive and
dehydrated after CLP, protein concentrations were measured in serum
using a Bradford assay, and cytokine levels were normalized for the
amount of protein that was present in the serum.
Culture and Isolation of Mast Cells
Mast cells were derived from femur bone marrow of specific pathogen-free CBA/J mice. The mast cells obtained from cultured bone marrow cells were c-kit-positive but were negative for CD3, CD4, CD8, CD23, B220, and F480. Mast cells were plated in 6-well plates and exposed to 5, 10, and 50 ng/ml of SCF or medium alone 2 hours before being exposed to LPS at 500 µg/ml. After 24 hours, supernatants were taken for ELISA analysis.
Histology and Immunohistochemistry
Livers from SCF- and saline-treated mice were removed at 4, 8, 24,
and 48 hours after CLP and immediately fixed in 4% paraformaldehyde
for a minimum of 12 hours. Fixed samples were subsequently embedded in
paraffin, thin-sectioned, and placed on L-lysine-coated
slides. Slides were deparaffinized by sequential treatment with xylene,
100% EtOH, 90% EtOH, 70% EtOH, 50% EtOH, distilled water, and
Tris-buffered saline (TBS; pH 7.68). Slides were microwaved for 20
minutes in 10 mmol/L citric acid buffer and allowed to cool. Tissue
sections were blocked in 1:50 dilution normal goat serum (blocking
solution) for 2 hours. Tissue sections were treated with one of the
polyclonal anti-mouse SCF antiserum, monoclonal anti-mouse IL-10
antibody (PharMingen, San Diego, CA), or the appropriate control
antibody. All were diluted at 1:100 with TBS containing blocking
solution (1:1) and the tissue sections were incubated overnight at
4°C in a humidified chamber. After incubation, slides were washed
twice for 5 minutes in TBS. A 1:35 dilution of biotinylated goat
anti-rabbit AB (BioGenex, San Ramon, CA) was placed on the slides for 2
hours at 37°C in a humidified chamber. Slides were again washed twice
in TBS. Slides were incubated with a 1:35 dilution of streptavidin
conjugated to horseradish peroxidase (BioGenex) for 45 minutes,
followed by two washes in TBS with 50 mmol/L levamisole. Fast red
chromagen (BioGenex) was placed on each slide and staining was
visualized at low power until color development was complete. The
staining reaction was terminated in sterile water and each slide was
counterstained with Mayer’s hematoxylin (0.1%; Sigma Chemical Co.,
St. Louis, MO).
Cytospins
Red blood cells were lysed in peritoneal washes, and red blood cell-free samples were spun for 5 minutes at 5,000 revolutions per minute in a Cytospin (Shandon, Pittsburgh, PA). Mast cells were stained with 0.5% toluidine blue O (Sigma Chemical Co.) and were counted. The mast cell number in each cytospin was expressed per 1 x 105 of the total cell number present in the peritoneum. The total cell numbers from each treatment group were not significantly different from each other.
Nuclear Extraction and Direct Lysis of Nuclei
Preparation of nuclear extracts from liver was conducted as follows. Briefly, liver samples were rapidly homogenized in PBS containing CompleteTB protease inhibitor (10 mg/ml; Boehringer Mannheim, Mannheim, Germany) and washed with fresh PBS. Homogenates were then suspended in buffer A (10 mmol/L Hepes, 10 mmol/L KCl, 0.5 mmol/L dithiothreitol, 1% Nonidet P-40) for 10 minutes and centrifuged for 10 minutes at 14,000 x g and the cytoplasmic supernatant was removed. The cell nuclei (found in the pellet) were suspended in buffer C (20 mmol/L Hepes, 20% glycerol, 500 mmol/L KCl, 0.2 mmol/L ethylenediaminetetraacetic acid, 0.5 mmol/L phenylmethyl sulfonyl fluoride, 0.5 mmol/L dithiothreitol, 1.5 mmol/L MgCl2) for 15 minutes and centrifuged at 7,000 x g for 10 minutes. The supernatant containing the nuclear proteins was removed for Western blot analysis.
Western Blot Analysis
Nuclear protein was measured using a Bradford assay (Bio-Rad) and 50 µg of liver nuclear extracts were electrophoresed on a 12% polyacrylamide gel and then transferred to polyvinylidene difluoride membranes (Bio-Rad). Equal protein loading was confirmed by Coomassie blue staining of the gel after transfer. Membranes were blocked for 2 hours at room temperature in 5% dry milk. STAT3 antibodies (Santa Cruz Biotechnology, Inc., Santa Cruz, CA) were diluted 1:1,000 and incubated overnight at 4°C. Horseradish peroxidase-linked secondary antibody (Pierce, Rockford, IL) was then added at 1:3,000 for 2 hours at room temperature and bands were detected by chemiluminescence (Bio-Rad).
Statistical Analysis
Survival curves were generated with Prism computer software
(Graphpad Software, Inc., San Diego, CA), and survival was examined
using the Fisher’s exact test. For all other analyses, a Student’s
t-test was used to test for significance. A P
value of 
0.05 was considered statistically significant.
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Although it has previously been shown that multiple treatments
with SCF induced mastocytosis and subsequently protected mice from
CLP-induced mortality,14
we examined whether a single SCF
treatment also reduced mortality in this model of sepsis. In the
present study, mice were given an i.p. injection of a range of SCF
doses at 2 hours before CLP surgery. In our pilot studies, we
discovered that the SCF pretreatment was necessary because a SCF
injection given at the time of or after CLP surgery did not improve
survival (data not shown). As illustrated in Figure 1A
, SCF pretreatment was associated with
a dose-dependent increase in survival after CLP. A dose of 4 µg/kg
significantly increased survival above the control rate (29%
versus 43%), but a dose of 20 µg/kg of SCF produced the
greatest increase in survival (29% versus 56%,
P 0.05). At a dose of 20 µg/kg, the protective
effect of SCF was observed after the first day with the survival of the
SCF pretreatment group at 88% (ie, 35 of 41 animals),
compared with the control group in which only 44% were alive (ie, 19
of 34 animals). By the fourth day, only 10 of the original 34 control
animals had survived (29%), whereas 23 of the original 41 mice
pretreated with 20 µg/kg SCF were alive (56%). Because a dose of SCF
at 20 µg/kg produced the greatest beneficial effect on mouse
survival, subsequent in vivo experiments used this dose of
SCF.
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in the Peritoneal Wash Are Not Increased by
SCF Pretreatment
Mast cells are a necessary component in the host response to the
bacteria introduced into the peritoneum after CLP
surgery.3
Likewise, SCF is an important cytokine involved
in mast cell activation,7-10
survival,18-20
and chemotaxis.11
Thus, we determined whether the enhanced
survival after a single SCF pretreatment was because of its effect on
mast cell numbers and/or the release of preformed mediators such as
TNF- in the peritoneal wash. First, the number of granulated mast
cells that stained with toluidine blue O were counted in peritoneal
washes removed at 5, 10, and 15 minutes after SCF treatment or saline
treatment. Other samples were removed from both treatment groups at 15
minutes, and 4, 6, and 8 hours after CLP surgery. Cytospins of
peritoneal washes revealed no significant differences in the number of
mast cells present between the two treatment groups at all time points
examined (Table 1)
. Nevertheless, it is
notable that stained mast cell were decreased in both treatment groups
after CLP suggesting that peritoneal mast cells released their granular
contents in response to the peritonitis. Furthermore, TNF- levels
were not significantly different between the two treatment groups at
any time point measured before and after CLP surgery (Table 2)
.
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We next examined whether a SCF pretreatment prevented mortality in
Sld/+mice. These mice are
characterized by diminished levels of transmembrane SCF and markedly
lower mast cell numbers, but express the c-kit receptor. SCF
pretreatment failed to prevent mortality in these mice
(n = 5), and by day 4 after CLP all of the
Sld/+ mice were dead (Figure 1B)
.
SCF Treatment Augments Monocyte Chemoattractant Protein-1 Levels in the Septic Peritoneum
Although peritoneal washes did not exhibit altered TNF- levels
after a single SCF injection, we nevertheless had convincing data from
the survival study using Sld/+ mice
that mature mast cells were required for the protective effect of SCF.
To further explore the relative contribution of mast cells to the
protective effects of SCF in our CLP model, we next examined cytokine
and chemokine levels in the peritoneal wash. The peritoneum represents
the local environment in which the mast cells presumably exert part of
their beneficial effect during CLP. Of the C-C and C-X-C chemokines
screened in the peritoneal wash (MCP-1, MIP-2, and MIP-1), only the
pattern of expression for MCP-1 was markedly different between the two
CLP groups. MCP-1 has been shown to be critical to the peritoneal host
defense response and elicits the recruitment of mononuclear cells and
neutrophils into the peritoneal cavity during CLP.16
Before CLP (time 0), MCP-1 was only present in the peritoneal wash of
SCF-pretreated mice (Figure 2A)
. At 4
hours, both CLP groups exhibited marked levels of MCP-1 in the
peritoneal wash, but the saline-treated CLP group contained twofold
more MCP-1. MCP-1 levels were similar in both groups at 8 hours after
CLP, but levels of this CC chemokine remained significantly elevated at
24 (P 0.0005) and 48
(P 0.0005) hours in the SCF-treated group
compared with the saline-treated group after CLP surgery (Figure 2A)
.
To determine whether MCP-1 was a mast cell-derived chemokine that
released on SCF stimulation, cultured bone marrow-derived murine mast
cells were exposed to medium alone or medium containing 5, 10, or 50
ng/ml of SCF. SCF dose-dependently increased the levels of MCP-1 in
these cultures from below ELISA detection levels (untreated) to
12.8 ± 6.7 ng/ml (50 ng/ml SCF treatment) (Figure 2B)
.
Interestingly, SCF does not promote the release of MCP-1 from untreated
or LPS-stimulated peritoneal macrophages (N. W. Lukacs,
personal communication). Thus, taken together these data suggested that
mast cells were the primary source of MCP-1 in the peritoneal cavity.
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The importance of MCP-1 in the SCF protective effect during CLP is
illustrated in Figure 3
. When mice
received recombinant SCF and anti-MCP-1 antiserum 2 hours before CLP,
the protective effect of the SCF treatment was abolished (Figure 3)
. At
2 days after CLP, 81% of SCF-treated control mice were alive (13 of
16 mice), whereas the group that received SCF and anti-MCP-1 antiserum
contained only 50% of the starting number of mice (eight of 16 mice).
At day 7 after CLP, 68% of the SCF-treated mice were alive, in
contrast to the other group that only contained 31% of the mice
subjected to CLP (P = 0.03). No additional
protective effect is seen when SCF is given in conjunction with rabbit
serum because of the beneficial effect that rabbit serum has after
CLP.15
Thus, these findings demonstrated that SCF
pretreatment augments MCP-1 levels within the peritoneal cavity before
and after CLP surgery and these changes are absolutely required for the
protective effect of SCF.
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Not only did SCF have an effect in the local peritoneal
environment, there was also evidence for a beneficial systemic response
to SCF treatment. Endogenous circulating levels of SCF were markedly
affected by the single SCF treatment before CLP. As shown in Figure 4A
, endogenous SCF was significantly
increased in the serum of SCF-pretreated mice at 4
(P = 0.044), 8 (P
0.0005), and 48 (P = 0.025) hours after CLP.
Given that stromal cells in the embryonic and adult liver produce
SCF,12,13
we next ascertained whether there was also an
effect in the liver. Immunohistochemical analysis of the liver is
summarized in Figure 5
. At 4 and 8 hours
after CLP surgery in saline-treated mice, SCF was expressed in the
cytoplasm and nucleus of hepatocytes localized around hepatic central
veins (Figure 5, A and C)
. Conversely, in SCF-pretreated mice at
similar times after CLP, dramatically less SCF staining was present in
the cytoplasm and nucleus in hepatocytes surrounding hepatic central
veins (Figure 5, B and D)
. Taken together, this data suggests exogenous
treatment with SCF is having dramatic effects systemically on both the
levels of endogenous SCF and the regulation of SCF within the liver.
This may represent a beneficial positive feedback mechanism in which
SCF pretreatment causes the release of endogenous SCF from the liver to
maintain the levels of SCF systemically.
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The local release of cytokines is critical during the host defense
response, yet when these mediators spill over into the systemic
circulation, a systemic inflammatory reaction can result. IL-10 is a
modulatory cytokine that enhances survival in bacterial toxin-induced
shock models by inhibiting the synthesis of proinflammatory
cytokines.21-23
IL-10 levels were measured to determine
whether SCF treatment modulated levels of this cytokine. Indeed, the
dramatic changes in circulating levels of IL-10 were observed in
SCF-treated mice that underwent CLP surgery. IL-10 was significantly
increased at 4 (P = 0.045), 8
(P = 0.041), and 24 (P =
0.004) hours after CLP surgery in the SCF-pretreated group compared
with the saline-treated group (Figure 4B)
.
We next looked to potential cellular sources of IL-10 during the CLP
response. Kupffer cells are the resident macrophages of the liver that
generate IL-1024,25
and play an important role in the
release of cytokines during a septic episode. Eight hours after CLP
surgery in control mice, immunoreactive IL-10 was detected in cells
that morphologically resemble Kupffer cells (Figure 6A)
. In contrast, in mice that received a
pretreatment of SCF, IL-10 staining was completely absent in these
cells and hepatocytes exhibited weak IL-10 staining (Figure 6B)
.
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STAT-3 is responsible for the activation of acute phase
proteins26
and has been positively correlated with
survival from septic shock.27
Therefore, we next examined
whether SCF treatment had any effect on the nuclear expression of
STAT-3 in the liver at 4, 8, and 24 hours after CLP using Western blot
analysis. As shown in Figure 7, 4
hours
after CLP, saline-treated mice exhibited very low levels of STAT-3
nuclear protein in liver nuclear extracts, whereas nuclear protein
expression of STAT-3 was present in all SCF-treated mice at higher
levels. Likewise, 8 hours after CLP, two of three saline-treated mice
exhibited low levels of STAT-3, whereas all four of the SCF-treated
mice exhibited strong STAT-3 expression. By 24 hours, all of the
surviving mice in both the saline- and SCF-treated groups exhibited
similar nuclear expression of STAT-3. Thus, these findings suggest that
SCF can either directly through the c-kit receptor on
hepatocytes13
or indirectly cause earlier nuclear
translocation of STAT-3 in the liver .
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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to clear bacteria introduced into the peritoneum from the
punctured cecum. As a result, these mice showed a significantly
increased mortality rate compared to mice that have a normal complement
of mast cells.3
Recently, SCF, a mast-cell growth and
activating factor, has also been shown to improve the survival of mice
subjected to CLP in a mast cell-dependent manner. In the present study,
we elucidated the protective mechanism of a single SCF treatment in the
absence of a mastocytosis. A single i.p. injection of SCF was not
associated with increased mast cell numbers in the peritoneal wash, nor
did this treatment markedly affect TNF- levels. However, after CLP,
SCF treatment was associated with markedly augmented mouse survival,
enhanced MCP-1 in the peritoneal wash, an increase in systemic levels
of both endogenous SCF and IL-10, and resulted in earlier expression of
STAT3 in the liver. Thus, a single SCF treatment was sufficient to
inhibit or abolish many of the deleterious outcomes that followed CLP
surgery in mice.
Several lines of evidence in the present study suggested that SCF was
protective in mice subjected to CLP through its direct effect on mast
cells. First and most convincing, the protective effect of SCF was
abolished in animals with decreased numbers of mature mast cells
subjected to CLP, indicating that SCF did not exert protective effects
independent of mast cell involvement. Next, although mast cells and
TNF- did not change in the peritoneal wash of SCF-pretreated mice
before and after CLP, peritoneal levels of MCP-1 were markedly altered.
Although the source of MCP-1 in SCF-treated mice was not conclusively
identified, our in vitro studies showed that mast cells
exposed to SCF for 24 hours secreted MCP-1 whereas untreated mast cells
did not. Furthermore, peritoneal macrophages did not produce MCP-1 in
response to SCF and LPS (data not shown). This is in accordance with
other studies that have found that depletion of mast cells reduces the
production of MCP-1 in the peritoneal lavage fluids after exposure to
zymosan.28
The MCP-1 generation within the peritoneal
cavity was absolutely essential for the beneficial effect of SCF
because animals that were given SCF and antiserum to MCP-1 failed to
survive. Furthermore, MCP-1 generation in the peritoneal cavity has
been shown to be critical for mouse survival after infection with
Pseudomonas aeruginosa and Salmonella
typhimurium29
and its neutralization is detrimental
in the context of CLP because of its actions on peritoneal macrophages
to produce LTB4 a neutrophil chemoattractant.16
This study
and others suggest that MCP-1 is the primary activator of the host
response in the peritoneal cavity after infection.30
The present study also suggested that there were other effects of SCF that may have contributed to its benefit after CLP. At present, it is not known whether these additional effects are downstream of the mast cell or they are independent of the mast cell. Nevertheless, SCF-pretreated mice exhibited dramatic increases in serum levels of IL-10. IL-10 is a pivotal cytokine during immune and inflammatory responses because it inhibits gene transcription thereby down-regulating a broad range of cytokines and chemokines.31,32 A number of studies have shown that IL-10 enhances survival during experimental toxin-induced shock models and CLP.32 Furthermore, histological findings from the present study suggested that SCF promoted the release of IL-10 from Kupffer cells. It is not likely that hepatocytes were producing IL-10 in the context of SCF treatment because isolated hepatocytes and hepatocyte cell lines fail to produce IL-10 after exposure to LPS and SCF (data not shown). Thus, these ELISA and histological findings suggest that SCF treatment before CLP promotes the rapid and prolonged release of the IL-10 from hepatic Kupffer cells, and the increase in IL-10 may also account for the protective effect of SCF in this model. These findings are consistent with previous experiments showing that IL-10 expression is autoregulated at the transcriptional level in human and murine Kupffer cells and IL-10 mRNA is dramatically increased after endotoxin exposure.24
One manifestation of sepsis is liver dysfunction in which there is
decreased transcription of key hepatic enzymes necessary for the
survival of the animal. STAT-3 is a key transcription factor that is
responsible for the transcription of many of these enzymes. Many
cytokines can activate one or more STATS.33
Specifically,
MCP-134
and IL-1035
can result in
translocation of STAT-3 to the nucleus in both immune and nonimmune
cells. The present study suggested that the SCF treatment promotes an
earlier and marked nuclear translocation of STAT-3. This event has been
previously associated with increased survival after CLP.27
In a sublethal sepsis model, hepatic STAT-3 activation was detected at
3 hours and persisted for 3 days. In contrast, in a lethal model of
sepsis, hepatic STAT-3 activation was only present for 3 to 16 hours
after CLP, and at dramatically lower levels.27
STAT-3
activation in SCF-treated mice could also be responsible for
transcription of acute phase proteins such as C-reactive
protein,26
2-macroglobulin,27
and
lipopolysaccharide-binding protein,36
all of which are
important during the host response to pathogens. Thus, the beneficial
effect of a single SCF treatment may be related to the direct effect
this factor had on the synthetic capacity of the liver during CLP.
Thus, a single SCF treatment before CLP surgery exerts a number of dramatic beneficial effects in the local peritoneal environment, on the systemic levels of cytokines, and in the liver. Further studies are warranted to explore the clinical feasibility of using SCF in the treatment and prevention of bacterial peritonitis.
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Footnotes |
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This study was supported by National Institutes of Health grants HL60289 and HL31237.
Accepted for publication July 20, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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/cachetin. Nature 1990, 346:274-276[Medline]
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