# The Importance of Fever as a Predictive Symptom  for  the Potency of Host's Monocytes to Release Pro- and Anti-Inflammatory Mediators

Authors: Magdalini Kyriakopoulou; Anastasia Antonopoulou; Maria Raftogiannis; Fotini Baziaka; Thomas Tsaganos; Kyriaki Kanellakopoulou; Evangelos J. Giamarellos-Bourboulis
Journal: Mediators of Inflammation
 (2008-01-22)

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## Abstract

Objective. To clarify whether time lapsing from advent of fever as a first sign of sepsis may be indicative of the potency of monocytes for the release of pro- and anti-inflammatory mediators.
Methods. Monocytes were isolated from blood of 51 septic patients and 9 healthy donors. Monocytes were incubated in the absence and presence of patients' serum and concentrations of tumour necrosis factor-alpha (TNFα), interleukin (IL)-6, IL-10, and malondialdehyde (MDA) were estimated in supernatants. Patients were divided into three groups: group A: <12 hours; group B: 12–24 hours, and group C: >24 hours between initiation of fever and blood sampling.
Results. TNFα of supernatants of groups B and C was higher than controls, as also were IL-6 of A and C, IL-10 of A and B,  and MDA of A. IL-6 of group A was increased after addition of patients serum. A negative correlation was found between time from initiation of symptoms and IL-6 of monocyte supernatants incubated in the presence of patients serum. Median IL-6 of survivors was higher than nonsurvivors.
Conclusion. Monocytes are potent for the release of pro- and anti-inflammatory mediators within the first 24 hours upon advent of fever related to sepsis; serum stimulates further release of IL-6 within the first 12 hours.


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## Body

## 1. Introduction

Despite the
increase of knowledge on the mechanisms of pathogenesis of sepsis, its
mortality remains high making sepsis the ninth cause of death in Northern
Europe and in the United
States
[1]. The increased incidence of the
septic syndrome has focused attention on various strategies for its management.
Among these strategies, immunotherapies have been evolved. These comprised
antibodies targeted against tumour necrosis factor-alpha (TNFα) and endotoxins,
soluble receptors of TNFα, plasma hemodialysis, inhibitors of nitric oxide
synthase, and activated protein C. Whereas most of them were successful when
used in phase II clinical trials, they have either failed or proved problematic
when applied in phase III clinical trials [2, 3]. Various hypotheses have been
proposed to explain the failure of immunotherapies. The most important seems to
be the lack of in-depth knowledge of the pathogenesis of sepsis [4]. Current
theories of pathogenesis are based on the production of proinflammatory and
anti-inflammatory cytokines after stimulation of blood monocytes by bacterial products and
immunotherapies are often targeted against these mediators. However, in all
clinical trials, therapies were administered upon fulfilment of certain
criteria of enrolment without fully knowing the immunological status of the
host on the time of start of any investigational product [4].The importance of
administration of immunotherapy on the time period when proinflammatory
mediators reach their peak has been underscored by experimental studies [5]. To
achieve that in the clinical field, it is mandatory to be able to correlate the
presence of symptoms with the function of the immune system. Monocytes of septic patients are often deactivated, that is, they
produce limited amounts of proinflammatory mediators when stimulated by
bacterial products [6]. However, no data are available if monocytes of septic
patients are capable to produce proinflammatory and anti-inflammatory
mediators when transferred ex vivo. The present study attempted to provide such
information and to correlate the ex vivo release of mediators from monocytes
with the time lapsing from the initiation of fever as a first sign of sepsis.


## 2. Patients and Methods

### 2.1. Characteristics of Patients

Enrolment
took place over the period January-September 2003 when the 4th Department of
Internal Medicine was situated in “Sismanoglion” General Hospital of Athens.
The protocol was approved by the Ethics Committee of the Sismanoglion General
Hospital; written informed consent was taken from the patients or first-degree
relatives. All consecutive admissions to the emergency department were eligible
for the study.Inclusion criteria were (a) the
presence of fever defined as body temperature greater than 38°C
being the first symptom of the underlying infection, (b) presence of one of the
following underlying infections: lower respiratory tract infection,
intrabdominal infection, or acute pyelonephritis, and (c) sepsis, severe sepsis,
or septic shock. Exclusion criteria were: (a) HIV infection, (b) neutropenia
defined as less 1000 neutrophils/μL,
and (c) intake of corticosteroids at a dose greater than or equal to 1 mg/kg of
equivalent prednisone for more than one month.Lower
respiratory tract infection was defined as the presence of all of the following
[7]: (a) core temperature > 38°C, (b) new or increased cough, (c) new
or increased purulent sputum production; and (d) new infiltrate on chest X-ray.Intrabdominal
infection was defined as the presence of all of
the following [8]: (a) core temperature > 38°C, (b) pain on
deep palpation, (c) radiological findings compatible with intra-abdominal
infection, and (d) white blood cells
> 12,000/μL or
<4,000/μL or > 10% of band forms.Acute
pyelonephritis was defined as the presence of all of  the following [9]: (a) core temperature > 38°C,
(b) lumbar tenderness or radiological findings compatible with acute
pyelonephritis, and (c) more than 10 white
blood cells phf of centrifuged urine.Sepsis was defined as the presence of an
infection accompanied by at least two of the following [10]: (a) body
temperature greater than 38°C, (b) respiratory rate higher than 20
breaths/min or PCO2 < 32 mmH, (c) heart rate above 90 beats/min, and
(d) white blood cells > 12,000 or <4,000/μL or more than 10% bands.Severe sepsis was defined as sepsis
complicated by the acute dysfunction of at least one organ due to an underlying
infection, that is, the acute presentation of at least one of the following [10]:(i)
acute respiratory
distress syndrome
(ARDS):
pO2/ FiO2 below
200 with diffuse
bilateral shadows in lung X-ray;(ii)
acute renal failure: urine production less than
0.5 mL/Kg body weight/h for at least two hours provided that the negative
fluid balance of the patient was restored;(iii)
metabolic acidosis:
pH < 7.30 or any base deficit
greater than
5 mEq/L and serum
lactate at least more than 2x normal value;(iv)
acute coagulopathy:
platelet count < 100.000/μL or INR > 1.5.Septic
shock was defined as sepsis complicated with systolic blood pressure below
90 mmHg for more than one hour requiring the administration of vasopressors
provided that the negative fluid balance of the patient is corrected [10].Twenty
five mL of whole blood were sampled from each patient after puncture of a
peripheral vein under sterile conditions before initiation of any antimicrobial
therapy; 20 mL were collected into a heparin-coated and sterile tube (Becton
Dickinson, Coskeysville, Md, USA) and the remaining into a sterile tube. The
latter was centrifuged and serum was kept at –70°C
until assayed.For each patient the following were recorded: age, sex, time interval
between initiation of fever and blood sampling, as well as APACHE II score and
outcome. The exact time of presentation of fever was provided by the patient’s
history. Nonreliable patients were not considered eligible.


### 2.2. Laboratory Techniques

For the isolation of peripheral blood mononuclear cells (PBMCs), heparinized
venous blood was layered over Ficoll Hypaque (Biochrom, Berlin, Germany) and
centrifuged. The separated mononuclear cells were washed three times with PBS
(pH 7.2) and resuspended in RPMI 1640 supplemented with 2 mM of glutamine
(Biochrom) in the presence of 100 U/mL of penicillin G and 0.1 mg/mL of
streptomycin (Sigma-Aldrich, Miss, USA). After incubation for 1 hour at 37°C
in 5% CO2, nonadherent cells
were removed while adherent monocytes were washed three times with Hank's
solution. Cells were then harvested by 0.25% trypsin/0.02% EDTA (Biochrom) and
counted in a Neubauer plate after trypan blue exclusion. Their purity in
monocytes was more than 95% as defined after incubation with the anti-CD14
mononuclear antibody at the fluorocolour FITC (Immunotech, Marseille, France)
and analysis by the EPICS/XL flow cytometer (Beckman Coulter Inc., Miami, Fl,
USA) using IgG-FITC-stained cells as negative controls.Half of cells were treated with an ice-cold cell lysis buffer (50 mM HEPES,
0.1% CHAPS, 5 mM DTT, 0.1 mM EDTA, pH 7.4). After centrifugation for ten minutes
at 10,000 g under 4°C, activity of
caspase-3 was estimated in the cytosolic extract by an enzymatic chromogenic
assay (BIOMOL Research Laboratories, Plymouth, Pa, USA). It was based on the
rate of hydrolysis at 37°C of a substrate
releasing p-nitroaniline overtime, as assessed by sequential photometry at
410 nm. The assay was also performed in the presence of a caspase-3 inhibitor.
The activity of caspase-3 in cell extracts was expressed as pmol/min⋅104 cells.The remaining half of monocytes were distributed into two wells of a
12-well plate at a
volume of 2.4 mL per well with RPMI 1640 supplemented with 2 mM of glutamine.
They were incubated in the absence or presence of 100μL of the patient’ serum
so that added serum represented 4.1% of the total well volume. After 24 hours
of incubation at 37°C under 5% CO2, the plate was
centrifuged and the supernatants were kept at –70°C
until assayed.  Monocytes isolated from nine
healthy donors and incubated only in the absence of serum were applied as
controls.Concentrations ofTNFα, interleukin (IL)-6, and IL-10 were estimated in
serum and supernatants by an enzyme immunoabsorbent assay (Diaclone, Paris,
France). The lower detection limits of the assay were 1.5 pg/mL for TNFα, 6.25 pg/mL for IL-6, and 12.5 pg/mL for IL-10. Their concentrations in monocyte
supernatants were expressed as pg/104 cells.Lipid peroxidation was estimated in serum and supernatants by the
concentration of MDA, as already described [11].
Briefly, a 0.1 mL aliquot of each sample was mixed to 0.9 mL of trichloroacetic
acid 20% (Merck) and centrifuged at 12,000 g and 4°C for 10 minutes. The supernatant was removed
and incubated with 2 mL of thiobarbituric acid 0.2% (Merck) for 60 minutes at 90°C. After centrifugation, a volume of 10 μL of
the supernatant was injected into a high-performance liquid chromatography
system (HPLC, Agilent 1100 Series, Waldbronn, Germany) with the following
characteristics of elution: Zorbax Eclipse XDB-C18 (4.6 × 150 mm, 5 μm) column
under 37°C; mobile phase
consisting by a 50 mM K3PO4 (pH: 6.8) buffer and methanol 99% at a
60/40 ratio with a flow rate of 1 mL/min; fluorometric detection with signals of
excitation at 515 nm and emission at 535 nm. The retention time of MDA was 3.5
minutes and it was estimated by a standard curve created with 1, 1, 3,
3-tetramethoxy-propane (Merck). All determinations were performed in duplicate.
Concentrations were expressed as μmol/mL of serum and μmol/104 cells
of supernatant.


## 2.1. Characteristics of Patients

Enrolment
took place over the period January-September 2003 when the 4th Department of
Internal Medicine was situated in “Sismanoglion” General Hospital of Athens.
The protocol was approved by the Ethics Committee of the Sismanoglion General
Hospital; written informed consent was taken from the patients or first-degree
relatives. All consecutive admissions to the emergency department were eligible
for the study.Inclusion criteria were (a) the
presence of fever defined as body temperature greater than 38°C
being the first symptom of the underlying infection, (b) presence of one of the
following underlying infections: lower respiratory tract infection,
intrabdominal infection, or acute pyelonephritis, and (c) sepsis, severe sepsis,
or septic shock. Exclusion criteria were: (a) HIV infection, (b) neutropenia
defined as less 1000 neutrophils/μL,
and (c) intake of corticosteroids at a dose greater than or equal to 1 mg/kg of
equivalent prednisone for more than one month.Lower
respiratory tract infection was defined as the presence of all of the following
[7]: (a) core temperature > 38°C, (b) new or increased cough, (c) new
or increased purulent sputum production; and (d) new infiltrate on chest X-ray.Intrabdominal
infection was defined as the presence of all of
the following [8]: (a) core temperature > 38°C, (b) pain on
deep palpation, (c) radiological findings compatible with intra-abdominal
infection, and (d) white blood cells
> 12,000/μL or
<4,000/μL or > 10% of band forms.Acute
pyelonephritis was defined as the presence of all of  the following [9]: (a) core temperature > 38°C,
(b) lumbar tenderness or radiological findings compatible with acute
pyelonephritis, and (c) more than 10 white
blood cells phf of centrifuged urine.Sepsis was defined as the presence of an
infection accompanied by at least two of the following [10]: (a) body
temperature greater than 38°C, (b) respiratory rate higher than 20
breaths/min or PCO2 < 32 mmH, (c) heart rate above 90 beats/min, and
(d) white blood cells > 12,000 or <4,000/μL or more than 10% bands.Severe sepsis was defined as sepsis
complicated by the acute dysfunction of at least one organ due to an underlying
infection, that is, the acute presentation of at least one of the following [10]:(i)
acute respiratory
distress syndrome
(ARDS):
pO2/ FiO2 below
200 with diffuse
bilateral shadows in lung X-ray;(ii)
acute renal failure: urine production less than
0.5 mL/Kg body weight/h for at least two hours provided that the negative
fluid balance of the patient was restored;(iii)
metabolic acidosis:
pH < 7.30 or any base deficit
greater than
5 mEq/L and serum
lactate at least more than 2x normal value;(iv)
acute coagulopathy:
platelet count < 100.000/μL or INR > 1.5.Septic
shock was defined as sepsis complicated with systolic blood pressure below
90 mmHg for more than one hour requiring the administration of vasopressors
provided that the negative fluid balance of the patient is corrected [10].Twenty
five mL of whole blood were sampled from each patient after puncture of a
peripheral vein under sterile conditions before initiation of any antimicrobial
therapy; 20 mL were collected into a heparin-coated and sterile tube (Becton
Dickinson, Coskeysville, Md, USA) and the remaining into a sterile tube. The
latter was centrifuged and serum was kept at –70°C
until assayed.For each patient the following were recorded: age, sex, time interval
between initiation of fever and blood sampling, as well as APACHE II score and
outcome. The exact time of presentation of fever was provided by the patient’s
history. Nonreliable patients were not considered eligible.


## 2.2. Laboratory Techniques

For the isolation of peripheral blood mononuclear cells (PBMCs), heparinized
venous blood was layered over Ficoll Hypaque (Biochrom, Berlin, Germany) and
centrifuged. The separated mononuclear cells were washed three times with PBS
(pH 7.2) and resuspended in RPMI 1640 supplemented with 2 mM of glutamine
(Biochrom) in the presence of 100 U/mL of penicillin G and 0.1 mg/mL of
streptomycin (Sigma-Aldrich, Miss, USA). After incubation for 1 hour at 37°C
in 5% CO2, nonadherent cells
were removed while adherent monocytes were washed three times with Hank's
solution. Cells were then harvested by 0.25% trypsin/0.02% EDTA (Biochrom) and
counted in a Neubauer plate after trypan blue exclusion. Their purity in
monocytes was more than 95% as defined after incubation with the anti-CD14
mononuclear antibody at the fluorocolour FITC (Immunotech, Marseille, France)
and analysis by the EPICS/XL flow cytometer (Beckman Coulter Inc., Miami, Fl,
USA) using IgG-FITC-stained cells as negative controls.Half of cells were treated with an ice-cold cell lysis buffer (50 mM HEPES,
0.1% CHAPS, 5 mM DTT, 0.1 mM EDTA, pH 7.4). After centrifugation for ten minutes
at 10,000 g under 4°C, activity of
caspase-3 was estimated in the cytosolic extract by an enzymatic chromogenic
assay (BIOMOL Research Laboratories, Plymouth, Pa, USA). It was based on the
rate of hydrolysis at 37°C of a substrate
releasing p-nitroaniline overtime, as assessed by sequential photometry at
410 nm. The assay was also performed in the presence of a caspase-3 inhibitor.
The activity of caspase-3 in cell extracts was expressed as pmol/min⋅104 cells.The remaining half of monocytes were distributed into two wells of a
12-well plate at a
volume of 2.4 mL per well with RPMI 1640 supplemented with 2 mM of glutamine.
They were incubated in the absence or presence of 100μL of the patient’ serum
so that added serum represented 4.1% of the total well volume. After 24 hours
of incubation at 37°C under 5% CO2, the plate was
centrifuged and the supernatants were kept at –70°C
until assayed.  Monocytes isolated from nine
healthy donors and incubated only in the absence of serum were applied as
controls.Concentrations ofTNFα, interleukin (IL)-6, and IL-10 were estimated in
serum and supernatants by an enzyme immunoabsorbent assay (Diaclone, Paris,
France). The lower detection limits of the assay were 1.5 pg/mL for TNFα, 6.25 pg/mL for IL-6, and 12.5 pg/mL for IL-10. Their concentrations in monocyte
supernatants were expressed as pg/104 cells.Lipid peroxidation was estimated in serum and supernatants by the
concentration of MDA, as already described [11].
Briefly, a 0.1 mL aliquot of each sample was mixed to 0.9 mL of trichloroacetic
acid 20% (Merck) and centrifuged at 12,000 g and 4°C for 10 minutes. The supernatant was removed
and incubated with 2 mL of thiobarbituric acid 0.2% (Merck) for 60 minutes at 90°C. After centrifugation, a volume of 10 μL of
the supernatant was injected into a high-performance liquid chromatography
system (HPLC, Agilent 1100 Series, Waldbronn, Germany) with the following
characteristics of elution: Zorbax Eclipse XDB-C18 (4.6 × 150 mm, 5 μm) column
under 37°C; mobile phase
consisting by a 50 mM K3PO4 (pH: 6.8) buffer and methanol 99% at a
60/40 ratio with a flow rate of 1 mL/min; fluorometric detection with signals of
excitation at 515 nm and emission at 535 nm. The retention time of MDA was 3.5
minutes and it was estimated by a standard curve created with 1, 1, 3,
3-tetramethoxy-propane (Merck). All determinations were performed in duplicate.
Concentrations were expressed as μmol/mL of serum and μmol/104 cells
of supernatant.


## 2.3. Statistical Analysis

Results
were expressed as median and 95% confidence intervals (CI) or as interquartile
range (IQR). For the purposes of analysis, patients were divided into three
groups according to the time interval between initiation of fever and blood
sampling: group A: <12 hours, group B: 12 to 24 hours, and group C, >24
hours from start of fever. Comparisons between groups were done by Mann-Whitney
U test with a Bonferroni correction; comparisons of yielded concentrations
between absence and presence of serum by Wilcoxon’s test. Correlations between
time interval from sampling and concentrations of cytokines were performed
according to Spearman’s rank of order. Any value ofP below .05 was
considered as significant.


## 3. Results

Over
the period of enrolment, 54 patients were eligible. Fifty one were finally
enrolled because only in them fever was the first symptom for underlying
infection. Demographic characteristics of these patients are shown in Table1. Sputum
cultures failed to disclose any pathogen.Table 1
Demographic characteristics of 51 patients
with septic syndrome enrolled in the study.
SepsisSevere sepsisSeptic shockNumber38103Male/Female15/233/70/3Age (mean ± SD)59.8±24.873.6±12.478.0±7.0APACHE
II score (mean ± SD)4.7±4.612.9±6.317.8±7.5White
blood cells
(/μl, mean ± SD)12800.9±5366.516367.0±4734.313856.7±2511.7Underlying infection [no. (%)]Lower
respiratory tract infection8
(21.1)4
(40.0)0Intrabdominal12 (31.6)2 (20.0)2 (66.7)Acute pyelonephritis18 (47.4)4 (40.0)1 (33.3)Bacteremia
[no. (% all
enrolled patients)]Providencia
stuartii—2 (20.0)—Escherichia
coli4 (10.5)——Positive urine cultures
(>105 cfu/ml) [no. (%)]Escherichia
coli16 (42.1)3 (30.0)1 (33.3)Pseudomonas
aeruginosa2 (5.3)1 (10.0)—Providencia
stuartii—1 (10.0)—Administered antimicrobials [no. (%)]2nd generation cephalosporin11 (28.9)1 (10.0)—2nd generation cephalosporin + metronidazole10 (10.5)2 (20.0)—Ceftriaxone + macrolide8
(21.1)4
(40.0)—Piperacillin/tazobactam + vancomycin03 (30.0)3 (100)Amplicilln/sulbactam9 (23.7)——Death (%)2 (5.3)1 (10.0)0 (0)Thirteen patients
belonged to group A, 16 patients to group B, and 22 patients to group C.
Concentrations ofTNFα, IL-6, IL-10, and MDA of serum in relation to that
time interval are shown in Table 2. No differences were found between groups of
time interval.Table 2
Serum concentrations of tumour necrosis
factor-alpha (TNFα),
IL-6,  IL-10, and malondialdehyde (MDA)
of 51 septic patients in relation to the time of blood sampling from advent of
fever.
Group A (n = 13) <12 hoursGroup B (n = 16) 12–24 hoursGroup C (n = 22) >24 hoursMedian (IQR)TNFα (pg/mL)9.88 (9.40)7.23 (7.54)7.64 (4.11)IL-6 (pg/mL)94.3 (225.1)65.1 (179.2)55.4 (120.5)IL-10 (pg/mL)<12.5<12.5<12.5MDA (μmol/mL)0.80 (11.30)2.87 (9.40)4.75 (10.05)Concentrations ofTNFα, IL-6, IL-10, and MDA of monocyte
supernatants without presence of
patients’ serum in relation to the time interval of initiation of fever from
sampling compared to controls are shown in Figure 1. TNFα of groups B and C was higher than controls, IL-6 of
groups A and C higher than controls, IL-10 of groups A and B higher than
controls, and MDA of group A higher than controls.Ex vivo release of tumour necrosis
factor-alpha (TNFα), IL-6, IL-10, and malondialdehyde (MDA) by monocytes
of 51 patients with septic syndrome and nine healthy donors. Patients were
divided into three groups depending on the time lapsing between blood sampling and advent of fever: group
A: <12 hours group B: 12–24 hours, and group C: >24 hours. Asterisks
denote outliers and circles denote extremes. P values refer to
comparisons with controls.
(a)(b)(c)(d)The effect of
serum on mediator release is shown in Figure2. IL-6 of supernatants of
monocytes isolated from patients of group A was increased after addition of
patients’ serum (P = .018). That was the case for MDA of groups B (P =
.049) and C (P = .048). IL-6 of supernatants of monocytes grown in the
presence of patients’ serum was higher for group A compared to groups B (P = .037) and C (P = .048).Effect of serum on ex vivo release of tumour necrosis factor-alpha (TNFα), IL-6, IL-10, and
malondialdehyde (MDA) by monocytes of 51 patients with septic syndrome.
Patients were divided into three groups depending of the time lapsing between
blood sampling and advent of fever: group A: <12 hours, group B: 12–24
hours, and group C: >24 hours. Circles denote extremes.
(a)(b)(c)(d)Median (IQR)
activity of caspase-3 was 53.5 (295.5) pmol/min⋅104 cells for group A, 200 (2020) pmol/min⋅104 cells for group
B, and 99.6 (1642) pmol/min⋅104 cells for group C (pNS between groups of time).A negative
correlation was found between time from initiation of symptoms and IL-6 of
monocyte supernatants incubated in the presence of patients’ serum (rs:−0.353, P = .041, Figure 3). No other significant correlation was found.Figure 3
Correlation between time lapsing from advent of fever to blood sampling and
concentrations of interleukin-6 (IL-6) released from monocytes of 51 septic
patients incubated in the presence of patients’ serum.Median
(IQR) IL-6 of supernatants of monocytes of survivors was 41.9 (149.2) pg/104 cells compared
to 1.9 (67.8) pg/104 cells of nonsurvivors
(P = .048). Respective values in the presence of serum were 51.6 (96.0) and 37.8 (30.0) pg/104 (pNS). No
differences were found in concentrations of TNFα, IL-10, and MDA of monocyte supernatants between
survivors and nonsurvivors.


## 4. Discussion

The
immunological function of the septic patient admitted to the emergencies cannot
yet be predicted with specificity despite the information provided by the existing
serum markers. This is of prime importance since data derived from animal
models underscore the need to know which immunological reaction takes place at
which time so as to administer the appropriate type of immunotherapy [5, 12].
Although various studies exist providing in-depth knowledge about the sequence
of events of the septic cascade in animals [4], there are various drawbacks for
the human situation. The present study attempted to correlate the potency of
monocytes of the septic host for the ex vivo release of pro- and
anti-inflammatory mediators at various time intervals from the advent of fever
in cases where fever was the initial symptom of the underlying septic process.
To our knowledge, a similar attempt has never been described before.Results
revealed that monocytes of septic patients were most potent for the ex vivo
release ofTNFα, IL-6, IL-10, and MDA compared to
healthy donors within the first 24 hours upon advent of fever (Figure 1). These
findings signify that monocytes of septic hosts are still able to release a
variety of mediators when put in an ex vivo milieu. Monocytes are probably
primed in their septic environment for that production.The pattern of
release of mediators by monocytes is not ubiquitous since MDA is mainly
secreted early,TNFα late, and IL-6 both early and late
in relation to start of fever. Perhaps safe conclusions can be drawn only for
IL-6 for two main reasons: (a) monocyte stimulation with patients’ serum yielded
further release of IL-6 only for monocytes isolated within the first 12 hours from
advent of fever (Figure 2), and (b) there is a negative correlation between time
lapsing from start of fever and serum-stimulated release of IL-6 by monocytes
(Figure 3).What may be the
underlying mechanism of induction of IL-6 by patients’ serum is not known and
only hypothesis can be done. Serum cytokines do not seem responsible for that,
since their levels do not differ between sera sampled at different time
intervals from advent of fever (Table2). Furthermore, caspase-3 activity of
monocytes was similar in correlation to time lapsing from presence of fever, so
it might not be hypothesized that endogenous inertia of monocytes was a
culprit. Whatever might be the underlying mechanism, the clinical significance
of that observation is that within the first 12 hours of fever, the monocytes
of the septic host are embedded in an environment promoting the release of IL-6
(Figure 2). This environment seems to be a major determinant of the outcome of
the septic patient. Despite the low number of deaths among the enrolled
patients, ex vivo release of IL-6 was greater among survivors compared to nonsurvivors.
That difference ceased to exist upon stimulation showing that the presence of
serum elicited significant release of IL-6 from monocytes of nonsurvivors. This
latter finding becomes of particular importance when considering data of animal
studies. In experimental peritonitis in mice, prompt administration of imipenem
could significantly decrease mortality with the sole exception of animals with
very high serum concentrations of IL-6 [13].The
findings of the present study are in general agreement with those described in
animal models of sepsis. Early sepsis in mice is accompanied by high serum
concentrations of both pro-inflammatory and anti-inflammatory cytokines as
opposed to low serum concentrations found in late sepsis [14]. On the same
context, ex vivo release of TNFα, IL-6, and IL-10 by monocytes of the septic host was
greater within the first 24 hours upon advent of fever.In
conclusion, our data revealed that fever might constitute a symptom predictive
of the immune function of the septic host. Monocytes are potent for the release
of pro- and anti-inflammatory mediators within the first 24 hours upon advent
of fever; serum of patients stimulates further release of IL-6 within the first
12 hours. These findings are in general agreement with both experimental and
clinical results that, as antibiotics, should be administered as early as
possible in the septic host [15, 16], the same may be necessary for
immunomodulatory treatment.



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*Source: 450196-2008-03-12.xml*