Download This article appeared in a journal published by - Mi Portal

This article appeared in a journal published by Elsevier. The attached
copy is furnished to the author for internal non-commercial research
and education use, including for instruction at the authors institution
and sharing with colleagues.
Other uses, including reproduction and distribution, or selling or
licensing copies, or posting to personal, institutional or third party
websites are prohibited.
In most cases authors are permitted to post their version of the
article (e.g. in Word or Tex form) to their personal website or
institutional repository. Authors requiring further information
regarding Elsevier’s archiving and manuscript policies are
encouraged to visit:
http://www.elsevier.com/copyright
Author's personal copy
European Journal of Internal Medicine 24 (2013) 132–138
Contents lists available at SciVerse ScienceDirect
European Journal of Internal Medicine
journal homepage: www.elsevier.com/locate/ejim
Original article
Circulating sICAM-1 and sE-Selectin as biomarker of infection and prognosis in patients
with systemic inflammatory response syndrome☆
Raúl de Pablo a, b, 1, Jorge Monserrat b, 1, Eduardo Reyes b, David Díaz b, Manuel Rodríguez-Zapata b, c,
Antonio de la Hera b, d, Alfredo Prieto b, Melchor Álvarez-Mon b, c, d, e,⁎
a
Intensive Care Unit, Hospital Universitario Príncipe de Asturias, Department of Medicine, University of Alcalá, Alcalá de Henares, Madrid, Spain
Laboratory of Immune System Diseases and Oncology, Department of Medicine, University of Alcalá, Madrid, Spain
Internal Medicine Service, Hospital Universitario de Guadalajara, Department of Medicine, University of Alcalá, Guadalajara, Spain
d
Molecular Medicine Institute, National Research Council (IMMPA-CSIC), University of Alcalá, Madrid, Spain
e
Immune System Diseases and Oncology Service, Hospital Universitario Príncipe de Asturias, Department of Medicine, University of Alcalá, Alcalá de Henares, Madrid, Spain
b
c
a r t i c l e
i n f o
Article history:
Received 18 June 2012
Received in revised form 29 August 2012
Accepted 28 October 2012
Available online 23 January 2013
Keywords:
Soluble adhesion molecules
Systemic inflammatory response syndrome
(SIRS)
Sepsis
Vascular endothelium
Fatal outcome
a b s t r a c t
Background: Vascular endothelium activation is a key pathogenic step in systemic inflammatory response
syndrome (SIRS) that can be triggered by both microbial and sterile proinflammatory stimuli. The relevance
of soluble adhesion molecules as clinical biomarkers to discriminate between infectious and non-infectious
SIRS, and the individual patient prognosis, has not been established.
Methods: We prospectively measured by sandwich ELISA, serum levels of soluble E-Selectin (sE-Selectin),
soluble vascular cell adhesion molecule-1 (sVCAM-1), soluble intercellular adhesion molecule-1 (sICAM-1)
and soluble intercellular adhesion molecule-2 (sICAM-2) at ICU admission and at days 3, 7, 14 and 28 in
patients with sepsis and at days 3 and 7 in patients with non-infectious SIRS.
Results: At ICU admission, sE-Selectin, sVCAM-1 and sICAM-1 in patients with infectious SIRS were significantly higher than those found in patients with non-infectious SIRS. ROC analysis revealed that the AUC for
infection identification was best for sICAM-1 (0.900 ± 0.041; 95% CI 0.819-0.981; p b 0.0001). Moreover, multivariate analysis showed that 4 variables were significantly and independently associated with mortality at
28 days: male gender (OR 15.90; 95% CI, 2.54–99.32), MODS score (OR 5.60; 95% CI, 1.67–18.74), circulating
sE-Selectin levels (OR 4.81; 95% CI, 1.34–17.19) and sVCAM-1 concentrations (OR 4.80; 95% CI, 1.34–17.14).
Conclusions: Patients with SIRS secondary to infectious or non-infectious etiology show distinctive patterns of
disturbance in serum soluble adhesion molecules. Serum ICAM-1 is a reliable biomarker for classifying
patients with infectious SIRS from those with non-infectious SIRS. In addition, soluble E-Selectin is a prognostic biomarker with higher levels in patients with SIRS and fatal outcome.
© 2012 European Federation of Internal Medicine. Published by Elsevier B.V. All rights reserved.
1. Introduction
Vascular endothelium plays a critical role in the regulation of the systemic leukocyte migration and homing [1]. Both the innate and adaptive
immune responses depend on the migration of leukocytes across endothelial cells. This interaction between the blood circulating leukocytes
and the endothelial cells is a complex process involving several cell surface molecules whose expression is directly related to the leukocyte and
endothelium activation stage [2]. Activation of endothelial cells induces
☆ Grant support: this study was supported by grant S-BIO-0189/2006 MITIC/TIMEDIC
from Comunidad de Madrid Spain, Fondo de Investigación Sanitaria FIS 98/1431,
SAF2004-8138, and by a clinical research prize awarded by the Fundación Lilly.
⁎ Corresponding author at: Departamento Medicina, Universidad de Alcalá, Carretera
Madrid-Barcelona, Km 33,600, E-28871 Alcalá de Henares, Madrid, Spain. Tel.: +34 91
8818700x2205; fax: +34 91 8833430.
E-mail addresses: [email protected] (R. de Pablo), [email protected]
(M. Álvarez-Mon).
1
R. de Pablo and J. Monserrat are joint first authors.
the up-regulation and expression of different adhesion molecules provoking the subsequent impact on the adhesive intercellular interactions
with circulating leukocytes. Briefly, the interaction of the microvilli that
bear L-Selectin (or CD62L) on the leukocyte membrane with vascular
cell adhesion molecule (VCAM)-1 on endothelial luminal surface constitutes the first step of the migration process. On the following steps of
rolling, activation and adhesion of the migrating cell to the endothelium
several adhesion molecules are implicated, including E-Selectin (or
CD-62E), intercellular adhesion molecule (ICAM) 1 and 2, and VCAM 1
on the endothelial cell membrane and the leukocyte integrins. Finally,
leukocyte diapedesis and basal membrane cross occur [3]. Interestingly,
these membrane bound adhesion molecules may suffer a proteolytic
cleavage of the cytoplasmic domain and soluble forms are generated
that can be measured in the blood [4].
Multiorgan universal endothelium injury is a key pathogenic step
in many severe systemic inflammatory diseases including the systemic inflammatory response syndrome (SIRS) [5]. SIRS has a common
endothelium damage pathway which is the result of an inappropriate
0953-6205/$ – see front matter © 2012 European Federation of Internal Medicine. Published by Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.ejim.2012.10.009
Author's personal copy
R. de Pablo et al. / European Journal of Internal Medicine 24 (2013) 132–138
generalized inflammatory response of the host to a variety of acute
insults [5]. Sepsis, one of the major causes of mortality in critically
ill patients, is defined as SIRS due to infection [6]. Moreover, precipitating insult of non-infectious nature such as severe pancreatitis,
burns or trauma can also be the cause of SIRS [6]. This wide number
of causes of SIRS is explained by the different and diverse biological defense mechanism and inflammation mediators such as
cytokines, complement activation, coagulation and fibrinolysis systems, kinin generation, which are involved in the induction of
anomalous activation and injury of the endothelial cells [7]. Furthermore,
bacterial products can also directly interact and activate endothelial
cells [7].
From a clinical point of view, monitoring soluble adhesion molecule levels may be not only of interest to understanding the pathophysiology of SIRS, but might also be a reliable diagnostic laboratory
test to distinguish between infection and non-infectious states in critically ill patients with clinical signs of SIRS. On the other hand, the
measurement of soluble adhesion molecules in serum have uncovered individualized prognostic value in critical ill patients with SIRS
[8–10].
Thus, we have centered our interest on the study of the levels and
the kinetics of variation of adhesion molecules in serum obtained
from SIRS patients caused by infectious and non-infectious etiologies
whose clinical status requires admission at an Intensive Care Unit
(ICU). We hypothesized that the endothelium damage induced by
infectious and non-infectious pathogenic mechanisms might be associated with different inflammation patterns of damage and subsequent
release of endothelial adhesion surface molecules to the serum. A prediction of the hypothesis was that circulating levels of soluble adhesion
molecules might also have a distinctive survival prognostic value in
patients with SIRS.
2. Materials and methods
2.1. Patient population and healthy controls
Approval from Hospital Príncipe de Asturias Ethics Committee was
obtained for the study protocol and written informed consent was
obtained from the next of kin and from healthy controls. The study
was performed over a period of 36 months.
In this study, we included patients 18 years or older admitted at
the ICU with diagnostic criteria of SIRS, which includes two or more
of the following conditions: (1) temperature > 38 °C or b36 °C; (2)
heart rate>90 beats per minute; (3) respiratory rate>20 breaths per
minute or PaCO2 b 32 mm Hg; and (4) white blood cell count>12,000/
mm3, b 4000 mm3, or >10% immature (band) forms [6]. Patients diagnosed with sepsis had SIRS and clinical evidence of infection, defined
as the presence of a known source of infection and/or a positive blood
culture and had been started on parenteral antimicrobial therapy [6].
We will use sepsis and infectious SIRS indistinctly, like synonymous.
Non-infectious SIRS was established by the absence of microbiological
documentation or radiological evidence of foci within 7 days after
admission in ICU. Exclusion criteria included: a) subjects with immunodeficiency or who were under corticosteroids or any other form of
immunomodulation therapy; b) autoimmune or hypersensitivity diseases; c) disseminated malignancy; d) renal failure requiring hemodialysis or hemofiltration; e) participation in another research study. All the
patients received conventional intensive care and included patients were treated by physicians who are not involved in this
study. Patients treated with hydrocortisone for refractory hypotension were withdrawn from the study. No patient was treated
with activated protein C. We prospectively designated a control
group to set normal adhesion molecule values. This group was
formed by healthy volunteers age- and sex-matched with the
patients.
133
2.2. Measurements and protocol
In patients with sepsis, blood samples were collected using an
non-heparinized arterial catheter at admission to the ICU and after
informed consent was obtained as well as 48 h and on days 7, 14
and 28. In non-infectious SIRS group, blood samples were taken at
ICU admission, at 48 h and on day 7. Immediately after blood samples
were taken, the blood was put into sterilized, silicone-coated glass
tubes. Samples were centrifuged at 2000 rpm for 20 min, and the
serum was stored at − 70 °C until assay. The etiology of SIRS was
categorized as sepsis or as non-infectious SIRS. The reviewers were
blinded to circulating soluble adhesion molecule results. We measured serum levels of soluble E-Selectin (sE-Selectin), soluble VCAM-1
(sVCAM-1), soluble ICAM-1 (sICAM-1) and soluble ICAM-2 (sICAM-2)
by a modification of the enzyme-linked immunosorbent assay procedure (ELISA) known as a sandwich ELISA, using commercially available kits from Diaclone Research (Connecticut, USA). For sE-Selectin,
sVCAM-1, sICAM-1 and sICAM-2, the lower detection limits of the
assay were 0.50, 0.60, 8.00 and 0.0002 ng/ml, respectively. In the 36
healthy controls, the samples serve as a control of the accuracy of our
laboratory procedures and for establishing the normal range of these
immuno-inflammatory parameters.
2.3. Statistical analysis
All measurements are expressed as mean ± S.E. mean. First, the
Kolmogorov–Smirnov test was used for testing data against the
normal distribution. All the variables always fulfilled the normality
hypothesis, thus, differences between groups were analyzed using
the value of t-Test for the difference between the means of two independent samples. Bonferroni correction for multiple tests was used.
The reliability of the use of serum soluble molecule adhesion concentrations to predict death due to SIRS or for diagnostic accuracy to evaluate
infection was calculated by plotting receiver-operating characteristic
(ROC) curves and the respective areas under the curves (AUC).
A Cox proportional-hazards regression analysis with backward
stepwise selection was performed to estimate the variables associated with mortality at 28 days. Predetermined variables, or those significantly associated with mortality at 28 days in the univariate
model (p b 0.10) were included in the multivariate model, including:
age, gender, APACHE II score, SOFA score, MODS score, acute renal
failure, mean arterial pressure, sE-Selectin, sICAM-1 and sVCAM-1
concentrations. A p b 0.05 was considered statistically significant.
Statistical analyses were performed with SPSS Statistics 19.0 statistical software (IBM Inc., Armonk, NY).
3. Results
3.1. Study patient characteristics
Of 132 SIRS patients who underwent screening for eligibility, we
excluded 40 (35%) because they had a previous underlying condition
associated with recognized alterations in immunological patient status and 92 were enrolled in this study. We included 52 patients
with sepsis and forty patients who had non-infectious SIRS. Also, 36
healthy volunteers were included (Fig. 1). The source of infection in
patients with sepsis was: intra-abdominal sepsis [n = 22], pneumonia
[n = 16], primary bacteremia [n = 4], urinary tract infection [n = 4],
soft-tissue infection [n=3], surgical site infection [n=2] or mediastinitis
[n=1]. The etiologies of non-infectious SIRS were non-infected severe
acute pancreatitis [n=19], resuscitated cardiac arrest [n=10], intracranial hemorrhage [n =5], hemorrhagic shock [n=5] and acute ischemic
stroke [n =1]. The baseline and outcome characteristics of the study
participants are summarized in Table 1.
Infectious and non-infectious SIRS patients show different patterns of circulating soluble adhesion molecules. First, we investigated
Author's personal copy
134
R. de Pablo et al. / European Journal of Internal Medicine 24 (2013) 132–138
Fig. 1. Screening and enrollment.
the serum levels of sE-Selectin, sVCAM-1, sICAM-1 and sICAM-2 in
infectious and in non-infectious SIRS patients at admission, at 48 h
and on day 7 of follow up in the ICU. In septic patients, we continued
the kinetic study of the serum levels of these soluble molecules at
days 14 and 28 (Fig. 2). We also studied in parallel healthy donors
as controls of the normal levels of the adhesion surface molecules.
As shown in Fig. 2 (panels A, B and C), sVCAM-1, sICAM-1 and
sICAM-2 serum levels were significantly higher in patients with
sepsis than in the group of healthy volunteers at ICU admission. The
serum levels of these three molecules remained significantly increased along the 28 days of follow-up with respect to those found
in healthy donors. In contrast, sE-Selectin concentration was only
increased in sepsis patients at ICU admission and at 48 h, decreasing
to normal levels afterward (Fig. 1, panel D). Non-infectious SIRS
patients showed a different pattern of soluble surface adhesion molecules at admission and first week of follow-up. In non-infectious SIRS
patients, the serum levels of sVCAM-1 and sICAM-1 levels at ICU
admission and along the seven days of follow-up were similar to
those found in healthy donors (Fig. 2, panel A and B). At ICU admission, circulating sICAM-2 concentrations were significantly increased
Table 1
Clinical characteristics of studied patients.
Age (years)
Gender (male/female)
APACHE II score
SOFA score
MODS score
Patients with shock
Patients that required mechanical
ventilation
Medical/surgical
Outcome (survived/died)
Non-infectious
SIRS
Sepsis
p
value
62.5 ± 3.1
26/14
22.1 ± 4.3
9.14 ± 0.44
7.90 ± 0.42
9 (22.5%)
30/40 (75.0%)
61.2 ± 3.2
36/16
25.3 ± 1.4
8.88 ± 0.74
7.63 ± 0.69
52 (100%)
47/52 (90.3%)
0.635
0.416
0.501
0.553
0.490
b0.001
0.088
35/5
29/11
24/28
34/18
0.201
0.269
Categorical variables are expressed as number of patients and continuous variables are
expressed as mean ± SE mean.
in non-infectious SIRS patients, but returned to normality afterward
(Fig. 2, panel C). sE-Selectin concentration was significantly higher
in non-infectious SIRS than in healthy controls at ICU admission and
at 48 h (Fig. 2, panel D).
The comparative analysis of the serum levels of the soluble adhesion molecules in both groups of SIRS patients shows two distinctive
patterns of abnormalities. At ICU admission, the levels of sVCAM-1
and sICAM-1, sE-Selectin in patients with sepsis were significantly
higher than those in patients with non-infectious SIRS (Fig. 2).
However, serum soluble ICAM-2 levels were similar in both groups
of patients at ICU admission (Fig. 2, panel C). Persistent higher levels
of sVCAM-1 and sICAM-1 were found in patients with sepsis during
the first week of the follow-up (panel A and B, Fig. 2). The levels
of ICAM-2 in sepsis patients were markedly elevated compared
with those in the non-infectious SIRS patients at seven days of the
follow-up.
According to the working hypothesis, we next investigated the
discriminative value of the serum levels of the studied adhesion
molecules between both groups of SIRS patients. To investigate
whether serum soluble adhesion molecules were able to differentiate
the patients with infection, we calculated their ROC curves. The AUC
for infection identification was higher for sICAM-1 (0.900 ± 0.041;
95% confidence interval, 0.819–0.981; p b 0.0001), followed by
sVCAM-1 and sE-Selectin. A cutoff value of serum soluble ICAM-1 of
904 ng/ml had 74.3% sensitivity and 94.1% specificity for separating
patients with infection from those with non-infectious SIRS (Fig. 3).
Soluble E-Selectin levels show a strong prognostic value for SIRS
patients at admission in ICU. Finally, we focused our investigation
to address the prognostic value of fatal outcome of the circulating
soluble adhesion molecules in the population of all patients with
SIRS admitted at the ICU. Mortality was defined as death occurring
within 28 days after study enrolment. At ICU admission, serum soluble E-Selectin concentrations were at significantly higher levels in
the nonsurvival group (106.4 ± 24.1 ng/ml) than in survival group
(78.4 ± 30.3 ng/ml; p b 0.001) of SIRS patients. In contrast, we did
not find significant differences between survivors and nonsurvivors
in circulating soluble VCAM-1, ICAM-1and ICAM-2 at ICU admission
in SIRS patients with and without infection. Furthermore, over the
study period, there were no significant differences in serum levels
of circulating soluble adhesion molecule concentrations between
survival and nonsurvival patients in both groups of patients.
We further studied serum soluble E-Selectin levels at ICU admission
as a predictor of outcome in all patients with SIRS. The AUC at baseline
measurements was 0.735 ±0.59 (95% confidence interval, 0.619–0.851;
p = 0.001). Thus, a sE-Selectin concentration of 95 ng/ml at ICU admission was identified as the optimum threshold to distinguish survival
and nonsurvival patients with a sensitivity of 75% and specificity of
64% (Fig. 4). The prognostic value of sE-Selectin levels was also significantly different between nonsurvivors and survivors within both
etiology groups: for infectious SIRS patients (110.0 ± 8.0 ng/ml vs.
88.7± 4.9 ng/ml; p = 0.041) than for non-infectious SIRS patients
(88.2 ± 9.1 ng/ml vs. 52.1 ± 35.8 ng/ml; p = 0.004).
To assess the impact of immunological parameters on mortality, we
performed a Cox proportional-hazards regression analysis. Multivariate
analysis showed that 4 variables were significantly and independently
associated with mortality at 28 days: male gender, MODS score, sESelectin and sVCAM-1 concentrations (Table 2).
4. Discussion
In this prospective study, we have demonstrated that patients
with sepsis and with non-infectious SIRS patients show different patterns of disturbance of the serum levels of soluble adhesion molecules
E-Selectin, VCAM-1, ICAM-1, and ICAM-2 at ICU admission. Serum
soluble ICAM-1 is a good biomarker for discriminating between infectious and non-infectious causes of SIRS in patients at ICU admission.
Author's personal copy
R. de Pablo et al. / European Journal of Internal Medicine 24 (2013) 132–138
135
Fig. 2. Comparison of circulating soluble VCAM-1, ICAM-1, ICAM-2 and E-Selectin, levels between patients with sepsis and with non-infectious SIRS and healthy controls. Soluble
VCAM-1 and ICAM-1 levels were higher in sepsis patients at ICU admission and during the follow-up than in non-infectious SIRS patients or in healthy controls. At ICU admission,
circulating soluble ICAM-2 concentrations are higher in non-infectious SIRS patients than in healthy controls and on day 7, sepsis patients had higher levels of soluble ICAM-2 than
non-infectious SIRS. sE-selectin concentration was significantly higher in sepsis than in non-infectious SIRS at ICU admission. Both are elevated at ICU admission and at 48 h and in
relation to healthy controls. Data are shown as mean ± S.E. mean. * p b 0.05 for comparison of means between sepsis patients and non-infectious SIRS patients and pb 0.05 for
comparison of means between SIRS patients and healthy controls.
In addition, in SIRS patients soluble E-Selectin is a prognostic biomarker with higher levels in patients with fatal outcome.
We hypothesized that in patients with SIRS the intensity and profile of the endothelium activation and damage induced by infectious
or non-infectious insults may be different. To assess this hypothesis,
we comparatively studied the serum levels of soluble relevant adhesion molecules shed by activated endothelium in the two groups of
patients with fully established diagnosis of SIRS with or without
infectious etiology. To homogenize the diagnosis and avoid the interference of potential acquired nosocomial infections, we measured the
sE-Selectin, sVCAM-1, sICAM-1 and sICAM-2 serum levels in patients
with SIRS at admission and during the first week of follow-up at the
ICU. Our results clearly show different patterns of disturbance in
the levels of serum soluble adhesion molecules in both groups of
SIRS patients. The latter statement is supported by different findings
in both groups of SIRS patients during the seven days of ICU followup. The maximum serum levels of adhesion molecules are significantly higher in infectious SIRS patients. The levels of sVCAM-1 and
sICAM-1 are continuously enhanced in infectious SIRS, but normal
in non-infectious SIRS. The kinetics of variation of sE-Selectin and
sICAM-2 in both groups of patients is different and sICAM-2 levels
increase in infectious SIRS along the first week at ICU. These clear
differences in the serum levels of the measured adhesion molecules
support the notion that sepsis patients suffer more severe endothelial
dysfunction than non-infectious SIRS. It is not possible to establish if
this different pattern of adhesion molecules shedding is explained
by a more intense and/or extended endothelium damage or a distinctive mechanism of cellular injury.
Increased levels of soluble adhesion molecules have also been
reported in patients with non-infectious SIRS [9,11–14]. Thus, our results confirm and expand some previous reports. Cowley et al. [11]
found that sICAM-1, sVCAM-1 and most notably sE-Selectin were increased in concentration in patients with SIRS. Hynninen et al. reported
that sE-Selectin are released in acute severe pancreatitis, but the levels
seem to be comparable to those in patients with severe sepsis [13] and
Geppert et al. [9] also found sE-selectin levels higher in patients after
successful cardiopulmonary resuscitation than in non-critically ill control patients, but there was no difference between patients with and
without SIRS. Cummings et al. [12] found that serum sE-selectin was
higher in patients with culture-positive sepsis than in other critically
ill medical ICU patients. Recently, Schuetz et al. [14] reported a sepsisspecific activation of endothelium activation markers, particularly sESelectin and soluble fms-like tyrosine kinase 1, in emergency department patients with hypotension.
The differential pattern of circulating adhesion molecule expression between infectious and noninfectious SIRS groups observed in
Author's personal copy
136
R. de Pablo et al. / European Journal of Internal Medicine 24 (2013) 132–138
Table 2
Variables associated with mortality at 28 days in the multivariate analysis.
Gender (male)
MODS score
sE-Selectin
sVCAM-1
Fig. 3. Receiver-operating characteristic (ROC) analysis of serum ICAM-1 concentrations
to distinguish infectious from non-infectious systemic inflammatory response syndrome.
Circulating sICAM-1 levels at ICU admission of 904 ng/ml had 74.3% sensitivity and 94.1%
specificity for separating patients with sepsis from those with non-infectious SIRS. The
AUC for sepsis identification was higher for sICAM-1 (0.900±0.041; 95% CI, 0.819–0.981;
pb 0.0001).
our work, suggests a selective involvement of endothelium, and should
result in the genesis of differential, selective immune cell infiltration.
The mechanism provoking both patterns of serum levels of adhesion
molecules remains unknown. However, it is possible to suggest that
Fig. 4. Receiver-operating characteristic (ROC) analysis of serum sE-Selectin concentrations
for mortality. Circulating sE-Selectin levels at ICU admission of 95 ng/ml had 75% sensitivity
and 64% specificity for mortality at 28 days in all patients with SIRS. The AUC at baseline
measurements was 0.735±0.59 (95% confidence interval, 0.619–0.851; p=0.001).
Odds ratio
95% confidence interval
p value
15.90
5.60
4.81
4.80
2.54–99.32
1.67–18.74
1.34–17.19
1.34–17.10
p = 0.003
p = 0.005
p = 0.016
p = 0.016
the endothelial cell dysfunction induced by infections agents is different
to that provoked by systemic aseptic inflammation. This suggestion is
supported by the expression of pattern recognition receptors (PRRs)
such as Toll-like receptors (TLRs) and nucleotide oligomerization
domain (NOD)-like receptors by endothelial cells [15–17]. Gramnegative and Gram-positive bacteria causing septic shock have pathogen associated molecular patterns (PAMPs) such as lipopolysaccharide
(LPS), lipoteichoic acid or peptidoglycan that can interact with endothelium PRRs inducing the endothelial cell activation and the subsequent
expression of inflammation mediators (cytokines, chemokines, adhesion molecules etc.). Given that the endothelium represents the first
point of contact between vessel and blood borne bacteria, it has been
proposed that TLR4 and NOD1 receptor activity in endothelial cells is
likely to be critical in sensing pathogens [15]. Thus, patients with infectious and noninfectious SIRS may share mechanisms of endothelium
dysfunction such as those related to proinflammatory cytokines, but
the effect of bacterial PAMPs on PRRs endothelial cells is selective of
septic patients.
In the last years, the results of intense research have revealed the
need to find a reliable diagnostic laboratory test that indicates sepsis
because it is well known that prompt antibiotic, hemodynamic and
adjuvant therapy improves survival in patients with sepsis [18]. Our
data show that the serum levels of sICAM-1 at ICU admission may
have a relevant value as biomarker for infectious cause of the SIRS.
Serum soluble ICAM-1 levels over 900 ng/ml may be an excellent laboratory marker for discrimination between SIRS of non-infectious
cause and sepsis. Previously, Takakuwa et al. [19] reported that blood
sICAM-1 levels were higher in patients with sepsis than in patients
with trauma, who met the criteria for SIRS. However, they also found
these differences in the blood levels of other adhesion molecules such
as sE-Selectin and sVCAM-1, cytokines, nitrite/nitrate oxide as well as
type II phospholipase A2 and concluded that seems to be because they
reflected differences in severity of the patients and in outcome because
a significantly higher proportion of patients died in the infection group.
Similarly, Boldt et al. [20] found that trauma patients showed lower
plasma levels of circulating adhesion molecules sE-Selectin, sICAM-1
and sVCAM-1 than did sepsis patients. It is important to remark that
unlike these previous studies, in our study, mortality and APACHE II
score were similar in both groups of patients and it seems that infection
is the main responsible of the differences we found in sICAM-1 levels
and not the severity of the disease. To our knowledge our data are the
first demonstration of the value of serum soluble ICAM-1 levels as a biomarker for discriminating between infectious and non-infectious causes
of SIRS in patients at ICU admission. It has been recently described that
the serum levels of decoy receptor 3 (DcR3), a soluble decoy receptor of
the tumor necrosis alpha (TNF), have a sensitivity and specificity for the
diagnosis of infection as cause of SIRS similar to the values found for
sICAM-1 in this work [21]. It is possible to suggest that the combination
of sICAM-1 and DcR3 quantification may further improve the diagnosis
of infectious cause of SIRS. Furthermore, the research to find fast and
reliable assays to distinguish sepsis from SIRS caused by other noninfectious diseases is not only centered in the pattern of the host inflammatory response but also in the early identification of bacteria in the
blood. Promising results have been reported with the use of new microbiology molecular techniques [22].
Author's personal copy
R. de Pablo et al. / European Journal of Internal Medicine 24 (2013) 132–138
We further characterized the kinetics of the serum adhesion molecules in sepsis patients along the four weeks of follow-up. It is well
known that increased circulating levels of sE-Selectin, sICAM-1 and
sVCAM-1 occur in sepsis [10,11,23–27]. However, the time course of
these adhesion molecules along 28 days was not completely defined
until now. We have demonstrated that leukocyte and endothelial
activation extends throughout a very extensive time in the patients
with sepsis who survive.
On the other hand, to our knowledge there are no data available
regarding circulating sICAM-2 concentrations in sepsis. ICAM-2,
which is expressed on endothelial cell surface, is the requisite adhesion molecule involved in transendothelial migration of various T
cell subsets. Soluble ICAM-2 levels in serum of patients with infectious SIRS remained significantly higher than the control level
throughout the course of sepsis, and this might reflect the persistent
disorder of the cellular immune function [28]. It is remarkable that
serum ICAM-2 levels were significantly higher in the group with
sepsis than in the group with non-infectious SIRS on day seven. The
persistence of elevated concentrations of adhesion soluble molecules sVCAM-1, sICAM-1 and sICAM-2 during the follow-up could
indicate that these adhesion molecules are probably poorly sensitive
to the treatment and cannot be used to evaluate the response to
therapy in patients with septic shock.
We compared the behavior of the sE-Selectin, sVCAM-1, sICAM-1
and sICAM-2 levels in SIRS patients with fatal outcome with respect to
those that survive. Interestingly, there is a marked overlap between
the levels of sVCAM-1, sICAM-1 and sICAM-2 found in survivors and
nonsurvivors with SIRS. However, sE-Selectin serum levels are significantly higher in nonsurvivors than in survivors at ICU admission. This
finding is very important because the expression of E-selectin appears
to be largely restricted to activated endothelial cells, as this molecule
is almost absent from resting endothelium [29,30]. Newman et al. did
a pioneer report showing that soluble E-selectin is elevated in the
serum of patients with septic shock [26]. Later, other authors have
described similar findings [31]. However, the value of this molecule as
prognostic biomarker of the clinical outcome of patients has not been
fully established. Our results agree with those published by several authors. Knapp et al. reported that plasma concentrations of E-Selectin
were predicting the prognosis of patients with sepsis secondary to infection with Gram-positive bacteria [25]. Cowley et al. showed that
high plasma concentration of E-Selectin was closely associated with
multiple organ dysfunction and death in patients with SIRS [11]. Other
authors have found similar results [20,32]. However, it has also described that circulating sE-Selectin levels have no clinical prognostic
value in patients with bacterial meningitis [33], with septic shock [27]
or with SIRS [34].
To assess the impact of immunological parameters on mortality,
we performed a multivariate analysis, including age, gender, severity
scores, organ failures, hemodynamic, respiratory and immunological
variables at ICU admission. It is important to remark that sE-Selectin
was an independent variable on mortality. Circulatory soluble levels
higher than 95 ng/ml were identified as indicator of mortality not
only in ROC analysis, but also in the multivariate analysis. Other variables associated with mortality were male gender, MODS score and
sVCAM-1. Our hypothesis is that those patients with SIRS and more
endothelial damage appear to have higher incidence and severity of
multiple organ failure and therefore, they have a worse outcome.
Female gender appears to influence the susceptibility and development of sepsis [35,36]. The majority of the biomarkers in sepsis and
SIRS were assessed for the ability to differentiate patients likely to
survive from those likely to die. However, the complexity of the
host response to injury may not lend to the identification of a single
ideal marker. Recent articles propose certain cytokine profiles (IL6,
IL8, and IL10) as predictor of severity and fatal outcome in severe
sepsis with a very high reliability [37,38]. We have also described
that an early anti-inflammatory cytokine response (soluble TNF
137
receptor I, soluble TNF receptor II and IL-1 receptor agonist) is associated with an enhanced risk of fatal outcome in patients with septic
shock [39]. Probably, a multi-marker approach may prove useful for
prognosis [40]. A panel containing E-Selectin may allow for reaching
this purpose.
As a limitation of this study, we should mention the baseline time
of blood withdrawal. All blood samples were collected on ICU admission, but the time that the patients had previously spent in SIRS in the
emergency room or in the operating room is a potential source of
inaccuracy. We can, however, state that all blood samples were
obtained within 12 h of they entered the ICU. Besides this, other factors such as host genetic polymorphisms, the focus of infection or the
characteristics of the pathogen may have also introduced variability.
Nevertheless, our patient population was homogenous and representative of immunocompetent patients with systemic inflammatory
response admitted to intensive care. In non-infectious SIRS group,
samples were taken at ICU admission, 48 h and on day 7. After this
period of time, microbiological colonization or nosocomial infection
appeared in most of the studied patients and a non-infectious cause
of SIRS could not be warranted. In non-infectious SIRS group, 30
patients (75%) required mechanical ventilation. All of them received
selective decontamination of the digestive tract with parenteral ceftriaxone for three days as prophylaxis. We did not address more infection markers, because it is clear that there is no ‘gold standard’
biomarker for sepsis. It seems that procalcitonin could be the best
laboratory marker in the diagnosis of sepsis [18], but systematic
reviews and meta-analysis concerning its diagnostic and prognostic
utility are contradictory [18,41–44]. Moreover, in elderly patients,
like our patients, the diagnostic performance of procalcitonin seems
to be poor [18,45,46]. Another limitation of our study was not to
determine more biomarkers reflecting endothelial cell state of the
patients that might be useful in the tracking of sepsis [47]. For
example, high serum laminin concentrations have been described in
patients with Candida sepsis compared with non-infected controls
[48] or dramatic increases of endothelial cell-specific molecule-1
(endocan) have been reported in patients with septic shock [49].
Moreover, they found that endocan levels significantly varied between patients with SIRS vs. sepsis with a sensitivity from about
82%. Unfortunately, this study lacks kinetic data [49], but in agreement with our findings strengthens the idea of more intense endothelial damage in sepsis than in non-infectious SIRS. Furthermore, it
might be interesting to study other adhesion molecules as biomarkers
related to vascular endothelial damage in SIRS patients. But, in some
molecules, the contribution of different cells than endothelium to their
serum levels may be critical as for example for those of P-selectin the
role of activated platelets.
In conclusion, our findings show that patients with SIRS secondary
to infectious or non-infectious etiology show distinctive patterns of disturbance in serum soluble adhesion molecules. Serum sICAM-1 may be
a reliable biomarker of infection in SIRS. In addition, sE-Selectin levels
may be used as predictor of fatal outcome in patients with SIRS. Thus,
E-Selectin is a particularly attractive candidate as a target in the treatment of patients with SIRS, especially in septic shock. A pilot study
with murine monoclonal antibody to sE-Selectin in patients with septic
shock indicated that this antibody appeared to be safe and had promising results as a therapy of septic shock [50]. Further studies of larger
groups of patients with SIRS are necessary to clarify this issue.
Learning points
• Endothelium activation and damage induced by infectious or noninfectious insults are different in patients with Systemic Inflammatory
Response Syndrome (SIRS).
• In patients with sepsis, sVCAM-1, sICAM-1, sICAM-2 and sE-Selectin
serum levels are significantly higher than healthy population at the
first three days of ICU admission.
Author's personal copy
138
R. de Pablo et al. / European Journal of Internal Medicine 24 (2013) 132–138
• Serum sICAM-1 is a reliable biomarker for discriminating between
infectious and non-infectious causes of SIRS at ICU admission.
• In patients with SIRS, elevated serum levels of sE-Selectin are good
prognostic biomarkers of fatal outcome.
Conflict of interests
The authors have no direct or otherwise commercial association
that might lead to a conflict of interest.
Acknowledgments
The authors thank the valuable help of the nursing and medical
staff of our ICU, the Department of Medicine and the University of
Alcalá. We also would like to thank Ignacio Arribas and Consuelo
Pintado for their invaluable statistical analysis assistance.
References
[1] Harlan JM, Winn RK. Leukocyte-endothelial interactions: clinical trials of antiadhesion therapy. Crit Care Med 2002;30:S214-9.
[2] Muller WA. Leukocyte-endothelial-cell interactions in leukocyte transmigration
and the inflammatory response. Trends Immunol 2003;24:327-34.
[3] Muller WA. Leukocyte-endothelial cell interactions in the inflammatory response.
Lab Invest 2002;82:521-33.
[4] Pigott R, Dillon LP, Hemingway IH, Gearing AJ. Soluble forms of E-selectin, ICAM-1
and VCAM-1 are present in the supernatants of cytokine activated cultured endothelial cells. Biochem Biophys Res Commun 1992;187:584-9.
[5] Giddings JC. Soluble adhesion molecules in inflammatory and vascular diseases.
Biochem Soc Trans 2005;33:406-8.
[6] Bone RC, Balk RA, Cerra FB, Dellinger RP, Fein AM, Knaus WA, et al. Definitions for
sepsis and organ failure and guidelines for the use of innovative therapies in
sepsis. The ACCP/SCCM Consensus Conference Committee. American College of
Chest Physicians/Society of Critical Care Medicine. Chest 1992;101:1644-55.
[7] Peters K, Unger RE, Brunner J, Kirkpatrick CJ. Molecular basis of endothelial
dysfunction in sepsis. Cardiovasc Res 2003;60:49-57.
[8] Gando S, Kameue T, Matsuda N, Hayakawa M, Ishitani T, Morimoto Y, et al. Combined activation of coagulation and inflammation has an important role in multiple organ dysfunction and poor outcome after severe trauma. Thromb Haemost
2002;88:943-9.
[9] Geppert A, Zorn G, Karth GD, Haumer M, Gwechenberger M, Koller-Strametz J,
et al. Soluble selectins and the systemic inflammatory response syndrome after
successful cardiopulmonary resuscitation. Crit Care Med 2000;28:2360-5.
[10] Sessler CN, Windsor AC, Schwartz M, Watson L, Fisher BJ, Sugerman HJ, et al.
Circulating ICAM-1 is increased in septic shock. Am J Respir Crit Care Med
1995;151:1420-7.
[11] Cowley HC, Heney D, Gearing AJ, Hemingway I, Webster NR. Increased circulating
adhesion molecule concentrations in patients with the systemic inflammatory
response syndrome: a prospective cohort study. Crit Care Med 1994;22:651-7.
[12] Cummings CJ, Sessler CN, Beall LD, Fisher BJ, Best AM, Fowler III AA. Soluble
E-selectin levels in sepsis and critical illness. Correlation with infection and hemodynamic dysfunction. Am J Respir Crit Care Med 1997;156:431-7.
[13] Hynninen M, Valtonen M, Markkanen H, Vaara M, Kuusela P, Jousela I, et al. Interleukin 1 receptor antagonist and E-selectin concentrations: a comparison in patients
with severe acute pancreatitis and severe sepsis. J Crit Care 1999;14:63-8.
[14] Schuetz P, Jones AE, Aird WC, Shapiro NI. Endothelial cell activation in emergency
department patients with sepsis-related and non-sepsis-related hypotension. Shock
2011;36:104-8.
[15] Gatheral T, Reed DM, Moreno L, Gough PJ, Votta BJ, Sehon CA, et al. A key role for
the endothelium in NOD1 mediated vascular inflammation: comparison to TLR4
responses. PLoS One 2012;7:e42386.
[16] Wang W, Deng M, Liu X, Ai W, Tang Q, Hu J. TLR4 activation induces nontolerant
inflammatory response in endothelial cells. Inflammation 2011;34:509-18.
[17] Faure E, Thomas L, Xu H, Medvedev A, Equils O, Arditi M. Bacterial lipopolysaccharide and IFN-gamma induce Toll-like receptor 2 and Toll-like receptor 4 expression
in human endothelial cells: role of NF-kappa B activation. J Immunol 2001;166:
2018-24.
[18] Herzum I, Renz H. Inflammatory markers in SIRS, sepsis and septic shock. Curr
Med Chem 2008;15:581-7.
[19] Takakuwa T, Endo S, Inada K, Kasai T, Yamada Y, Ogawa M. Assessment of inflammatory cytokines, nitrate/nitrite, type II phospholipase A2, and soluble adhesion
molecules in systemic inflammatory response syndrome. Res Commun Mol
Pathol Pharmacol 1997;98:43-52.
[20] Boldt J, Muller M, Kuhn D, Linke LC, Hempelmann G. Circulating adhesion molecules
in the critically ill: a comparison between trauma and sepsis patients. Intensive Care
Med 1996;22:122-8.
[21] Hou YQ, Xu P, Zhang M, Han D, Peng L, Liang DY, et al. Serum decoy receptor 3, a
potential new biomarker for sepsis. Clin Chim Acta 2012;413:744-8.
[22] Fitting C, Parlato M, dib-Conquy M, Memain N, Philippart F, Misset B, et al.
DNAemia detection by multiplex PCR and biomarkers for infection in systemic
inflammatory response syndrome patients. PLoS One 2012;7:e38916.
[23] Gando S, Kameue T, Matsuda N, Hayakawa M, Hoshino H, Kato H. Serial changes
in neutrophil-endothelial activation markers during the course of sepsis associated with disseminated intravascular coagulation. Thromb Res 2005;116:91–100.
[24] Kayal S, Jais JP, Aguini N, Chaudiere J, Labrousse J. Elevated circulating E-selectin,
intercellular adhesion molecule 1, and von Willebrand factor in patients with
severe infection. Am J Respir Crit Care Med 1998;157:776-84.
[25] Knapp S, Thalhammer F, Locker GJ, Laczika K, Hollenstein U, Frass M, et al. Prognostic
value of MIP-1 alpha, TGF-beta 2, sELAM-1, and sVCAM-1 in patients with grampositive sepsis. Clin Immunol Immunopathol 1998;87:139-44.
[26] Newman W, Beall LD, Carson CW, Hunder GG, Graben N, Randhawa ZI, et al.
Soluble E-selectin is found in supernatants of activated endothelial cells and is
elevated in the serum of patients with septic shock. J Immunol 1993;150:644-54.
[27] Weigand MA, Schmidt H, Pourmahmoud M, Zhao Q, Martin E, Bardenheuer HJ.
Circulating intercellular adhesion molecule-1 as an early predictor of hepatic
failure in patients with septic shock. Crit Care Med 1999;27:2656-61.
[28] Qi BT, Wang P, Li J, Ren HX, Xie M. Levels of soluble vascular cell adhesion
molecule-1 and soluble intercellular adhesion molecule-2 in plasma of patients
with hemorrhagic fever with renal syndrome, and significance of the changes in
level. Viral Immunol 2006;19:565-9.
[29] Bevilacqua MP, Nelson RM. Selectins. J Clin Invest 1993;91:379-87.
[30] Jilma B, Blann A, Pernerstorfer T, Stohlawetz P, Eichler HG, Vondrovec B, et al.
Regulation of adhesion molecules during human endotoxemia. No acute effects
of aspirin. Am J Respir Crit Care Med 1999;159:857-63.
[31] Leone M, Garcin F, Chaabane W, Boutiere-Albanese B, Albanese J, Dignat-Georges
F, et al. Activation of adhesion molecules in patients with septic shock. Ann
Fr Anesth Reanim 2003;22:721-9.
[32] Endo S, Inada K, Kasai T, Takakuwa T, Yamada Y, Koike S, et al. Levels of soluble
adhesion molecules and cytokines in patients with septic multiple organ failure.
J Inflamm 1995;46:212-9.
[33] Megarbane B, Marchal P, Marfaing-Koka A, Belliard O, Jacobs F, Chary I, et al. Increased
diffusion of soluble adhesion molecules in meningitis, severe sepsis and systemic inflammatory response without neurological infection is associated with intrathecal
shedding in cases of meningitis. Intensive Care Med 2004.
[34] Stoiser B, Knapp S, Thalhammer F, Locker GJ, Kofler J, Hollenstein U, et al. Time
course of immunological markers in patients with the systemic inflammatory
response syndrome: evaluation of sCD14, sVCAM-1, sELAM-1, MIP-1 alpha and
TGF-beta 2. Eur J Clin Invest 1998;28:672-8.
[35] Schroder J, Kahlke V, Staubach KH, et al. Gender differences in human sepsis. Arch
Surg 1998;133:1200-5.
[36] Wichmann MW, Inthorn D, Andress HJ, Schildberg FW. Incidence and mortality of
severe sepsis in surgical intensive care patients: the influence of patient gender
on disease process and outcome. Intensive Care Med 2000;26:167-72.
[37] Andaluz-Ojeda D, Bobillo F, Iglesias V, et al. A combined score of pro- and antiinflammatory interleukins improves mortality prediction in severe sepsis. Cytokine 2012;57:332-6.
[38] Rodriguez-Gaspar M, Santolaria F, Jarque-Lopez A, et al. Prognostic value of cytokines in SIRS general medical patients. Cytokine 2001;15:232-6.
[39] De Pablo R, Monserrat J, Reyes E, et al. Mortality in patients with septic shock
correlates with anti-inflammatory but not proinflammatory immunomodulatory
molecules. J Intensive Care Med 2011;26:125-32.
[40] Ventetuolo CE, Levy MM. Biomarkers: diagnosis and risk assessment in sepsis.
Clin Chest Med 2008;29:591-603.
[41] Tang BM, Eslick GD, Craig JC, McLean AS. Accuracy of procalcitonin for sepsis diagnosis in critically ill patients: systematic review and meta-analysis. Lancet Infect
Dis 2007;7:210-7.
[42] Jones AE, Fiechtl JF, Brown MD, Ballew JJ, Kline JA. Procalcitonin test in the diagnosis of bacteremia: a meta-analysis. Ann Emerg Med 2007;50:34-41.
[43] Uzzan B, Cohen R, Nicolas P, Cucherat M, Perret GY. Procalcitonin as a diagnostic
test for sepsis in critically ill adults and after surgery or trauma: a systematic
review and meta-analysis. Crit Care Med 2006;34:1996-2003.
[44] Simon L, Gauvin F, Amre DK, Saint-Louis P, Lacroix J. Serum procalcitonin and
C-reactive protein levels as markers of bacterial infection: a systematic review
and meta-analysis. Clin Infect Dis 2004;39:206-17.
[45] Stucker F, Herrmann F, Graf JD, Michel JP, Krause KH, Gavazzi G. Procalcitonin and
infection in elderly patients. J Am Geriatr Soc 2005;53:1392-5.
[46] Dwolatzky T, Olshtain-Pops K, Yinnon AM, Raveh D, Rogowski O, Shapira I, et al.
Procalcitonin in the elderly: normal plasma concentrations and response to bacterial infections. Eur J Clin Microbiol Infect Dis 2005;24:763-5.
[47] Pierrakos C, Vincent JL. Sepsis biomarkers: a review. Crit Care 2010;14:R15-.
[48] Presterl E, Lassnigg A, Mueller-Uri P, Wenisch C, El-Menyawi I, Graninger W. High
serum laminin concentrations in patients with Candida sepsis. Eur J Clin Invest
1999;29:992-6.
[49] Scherpereel A, Depontieu F, Grigoriu B, Cavestri B, Tsicopoulos A, Gentina T, et al.
Endocan, a new endothelial marker in human sepsis. Crit Care Med 2006;34:532-7.
[50] Friedman G, Jankowski S, Shahla M, Goldman M, Rose RM, Kahn RJ, et al. Administration of an antibody to E-selectin in patients with septic shock. Crit Care Med
1996;24:229-33.