Sérový protein S100B jako molekulární marker závažnosti poranění mozku u dětí
Cíle: S100B je proteinový biomarker, který odráží poranění CNS. Cílem této studie bylo zjistit korelaci mezi počáteční hladinou sérového proteinu S100B a mortalitou a pomocí nálezů počítačové tomografie (CT) a dále prozkoumat, zda existuje spojení mezi S100B a Glasgowskou škálou výsledků (GOS) po šesti měsících.
Metody: Do této prospektivní studie bylo zařazeno 43 pacientů s poraněním mozku (TBI). TBI bylo ověřeno počítačovou tomografií podle Marshallovy klasifikace. Při přijetí a následně každých 24 hod po dobu maximálně šesti po sobě jdoucích dní byly odebírány vzorky žilní krve. Výsledky byly u všech pacientů posouzeny šest měsíců po TBI pomocí Glasgowské škály výsledků (GOS).
Výsledky: GOS byla považována za základní koncový bod pro všechny prediktivní analýzy. Prokázali jsme statisticky významný vztah mezi skupinou pacientů s GOS = 1 a GCS ≤ 8 (p = 0,046), Marshallovým klasifikačním hodnocením > II (p < 0,001) a zvýšeným výskytem určitého typu zranění – intrakraniálním krvácením (p = 0,045), subdurálním hematomem (p = 0,008), frakturou lebky (p = 0,024), edémem (p = 0,001). Poměr S100B druhý den/počáteční hodnota S100B významně statisticky diferencoval porovnávanou skupinu pacientů (s GOS = 1 a s GOS > 1; p = 0,03). Hladiny S100B byly zvýšeny u pacientů s určitým typem poranění, zejména intrakraniálním krvácením, subdurálním hematomem a edémem.
Závěr: Hladina S100B byla potvrzena jako klinicky cenný indikátor závažnosti zranění a byla navržena jako efektivní prediktor rizika výsledku (GOS = 1).
Klíčová slova: S100 protein – severe head injury – outcome – children
J. Žurek 1; L. Bartlová 2; L. Marek 1; M. Fedora 1
LF MU a FN Brno
Klinika dětské anesteziologie a resuscitace
1; LF MU a FN Brno
Klinika dětské neurologie
Cesk Slov Neurol N 2010; 73/106(1): 37-44
Objectives: S100B is a protein biomarker that reflects CNS injury. The aims of the current study were to investigate correlations between the initial level of serum S100B protein and mortality and computerized tomography (CT) findings, as well as to establish whether there is an association between S100B and Glasgow outcome scale (GOS) after six months.
Methods: This prospective study enrolled 43 patients with traumatic brain injury (TBI), verified by computerized tomography and categorized by Marshall classification. Venous blood samples were taken on admission and every 24 h for a maximum of six consecutive days. The outcome was evaluated six months after TBI using the Glasgow outcome scale (GOS) in all patients.
Results: GOS was taken as principal end‑point for all predictive analyses. We demonstrated statistically significant relationships between groups of patients with GOS = 1 and GCS ≤ 8 (p = 0.046), Marshall classification score > II (p < 0.001) and increased incidence of some types of injury – intracranial bleeding (p = 0.045), subdural haematoma (p = 0.008), skull fracture (p = 0.024), and oedema (p = 0.001). The ratio of S100B in 2nd day/initial S100B value significantly differentiated between the groups of patients compared (with GOS = 1 and with GOS > 1; p = 0.03). Levels of S100B were elevated in patients with some specific types of injury, namely intracranial bleeding, subdural haematoma and oedema.
Conclusion: The level of S100B was confirmed as a clinically valuable indicator of severity of injury and is proposed as an effective predictor of risk outcome (GOS = 1).
Key words: protein S100 – vážný úraz hlavy – výsledek – děti
are a major cause of morbidity and mortality in children aged a year
or more, and traumatic brain injury (TBI) is the injury most responsible for
deaths . Case series from multiple trauma centres report that 75–97% of
trauma deaths in children result from TBI .
Despite significant progress in cerebral
monitoring, it remains difficult to quantify the extent of primary trauma in
TBI, to monitor secondary changes, and to predict neurological outcome with the
available to us today. Standard methods of addressing prognoses of severity of
initial brain injury and anticipating the onset of secondary injury have
included neurological examination, neuro‑imaging
studies, intracranial pressure monitoring, electrodiagnostic testing, and
However, the use of protein biomarkers for
the detection of injury and prediction of outcomes has been attracting
increasing clinical interest. These markers have been investigated in serum,
cerebrospinal fluid (CSF) and even urine. Most of the protein biomarkers that
have been investigated to date are mediators of the injury response or
subsequent cellular damage or death .
The S100 proteins are a family of
dimeric cytosolic calcium‑binding
proteins made up of an alpha and a beta isomer. S100 proteins are
found in abundance in the astroglial and Schwann cells and have also been found
in certain tumours, such as schwannoma, glioma, melanoma, and neuroblastoma
[4,5]. Elevations in serum S100B protein levels have been reported to reflect
injury to the brain and increased permeability of the blood‑brain barrier .
believe that initial serum S100B levels (simple blood test) can provide
valuable diagnostic indications of the structural damage at an early stage of
severe TBI, especially in patients lacking reliable clinical (sedated and
intubated), imaging (normal or minimal damage on initial computerized
tomography) or neurophysiological evaluation, and may assist in eventual
prognosis. Serial measurements of S100B levels can also monitor secondary
events and contribute to better day‑to‑day assessment and treatment.
The aims of the current
study were to study correlations between the initial level of serum S100B
protein and both mortality and the computerized tomography (CT) findings that
indicate radiologically visible brain damage, as well as to investigate whether
there is an association between S100B and the Glasgow outcome scale (GOS).
This prospective cohort study consecutively enrolled
43 children of < 19 yrs of age with severe TBI brought to the
University Children’s Hospital, Brno, from June 2007 through November
2008. All the patients were monitored continuously in PICU and received
standard neuro‑intensive care, including intubation and mechanical
ventilation, haemodynamic monitoring, and intracranial pressure monitoring
based on TBI therapeutic protocol. Intracranial hypertension was treated
progressively by use of a standard step‑wise protocol that
included sedation, paralysis, mild hyperventilation (target paCO2 32–35 mmHg),
osmotherapy with mannitol, and use of barbiturates. Cerebral perfusion pressure
was maintained at 60 mmHg (50 mmHg in infants) by lowering
intracranial pressure to 20 mmHg (15 mmHg in infants) and by
maintaining mean arterial blood pressure at 80 mmHg (70 mmHg in
The Glasgow coma scale
 was based on evaluation of the emergency measures taken, although without consideration of pharmacological
interventions. Initial CT scans obtained on admission were analyzed after the
classification established by Marshall et al  into the following categories:
grade I, no visible intracranial pathology seen on CT; grade II, cisterns present
with midline shift 0–5mm, lesion densities present, but no high‑ or mixed‑density lesion larger than 25ml, may include bone fragments and foreign bodies; grade III, cisterns
compressed or absent with midline shift 0–5mm, no high‑ or mixed‑density lesion larger than 25ml; grade IV, midline shift more than 5mm, no high‑ or mixed‑density lesion larger than 25ml; grade V, any lesion surgically evacuated; and grade VI, high‑ or mixed‑density lesion larger than 25ml, not surgically evacuated.
Venous blood samples to
determine levels of S100 were always taken on admission, then every day
until a maximum of six days of hospitalization. Samples were centrifuged
and serum was frozen to –70°C for analysis. S100B was measured immuno‑luminometrically with a commercially available kit (Elecsys®
analyzer, Roche Diagnostics).
outcome was measured after six months using the GOS  with its ratings of 1 (death),
2 (vegetative state), 3 (severe disability), 4 (moderate
disability), and 5 (good outcome), based on electro‑encephalography and neurological
examination. The person
who carried out the assays and decided upon outcome was completely blinded to
the clinical information.
The study protocol and
informed consent approach were approved by the ethicscommittee of the University Hospital, Brno. Parents provided written
informed consent for their children to participate in this trial.
The standard methods of nonparametric statistics were
employed for the analysis. Categorical data were described by frequency
analysis of categories, continuous data using median and percentiles.
Relationships between categorical variables were analyzed by means of maximum
likelihood chisquare test; the Mann‑Whitney U test was
used for comparison of values of continuous variables between categories of
characteristic curves (ROC) were applied for the analysis of the predictive
power of injury characteristics, biochemical parameters and initial CT
diagnostics. GOS score = 1 was taken as principal study endpoint
for the purposes of predictive analyses.
Analyses were performed
using Statistica 8 (StatSoft, Inc.) and SPSS 17 (SPSS Inc.).
GOS was taken as principal end‑point for all predictive analyses, i.e. those analyses that have been
performed in order to assess the predictive potential of a set of
characteristics, both of patient and of injury.
total of 43 children (53.5% boys, 46.5% girls) with TBI were enrolled.
Table 1 shows summary
statistics of the initial characteristics of the injured children. Common
descriptors such as age, sex and weight are not related to the final GOS
values. Similarly, no significant correlation between age and S100B initial values
was observed (Figure 1).
Glasgow outcome scale
Table 1 shows some statistically significant
relationships between certain descriptors and a group of patients with
GOS = 1:
GCS ≤ 8 (all
patients with GOS = 1 belong to this category,
p = 0.046),
score > II (no patient with GOS = 1 has Marshall score
I or II, p < 0.001),
Increased incidence of
some type of injury, as diagnosed by CT scan:
multiple CT findings,
namely skull fracture in combination with some other injury type
(p = 0.012).
All the parameters that
significantly distinguished patients with GOS = 1 from the others (Table
1) serve as candidates for effective predictors.
Dynamics of S100B
Figure 2 shows a statistically significant
decrease in S100B values in the group of patients with final GOS > 1. This
significant decrease started during the second day of hospitalization and continued
for the whole follow‑up period of six days. Initially, 91.7% of patients
exceeded the cut‑off level of 0.105 µg/l, while by the end of the
follow‑up period this had become only 6.5% (p < 0.001). On
the other hand, the cohort with GOS = 1 did not exhibit
a statistically significant decrease in S100B values, although in median
value the level of S100B decreased within the period (from 1.6 µg/l to
0.27 µg/l). The level of S100B remained significantly higher in the risk
group (GOS = 1) in comparison with the GOS > 1 group for the
whole follow‑up period, including initial value on the first day
(Table 2). The decrease of S100B level in the risk group of patients was not
significant in comparison with the initial value (Figure 2). Based on time‑related profiles, we related the subsequent follow‑up values of S100B to the initial level in pair‑wise calculated ratios. Only S100B in second day/initial S100B ratio
significantly differentiated the two groups of patients compared (with
GOS = = 1 and with GOS >1;
p = 0.03) (Figure 3, Table 2).
Levels of S100B and CT findings
Levels of S100B were elevated in patients with some
specific types of injury, namely intracranial bleeding, subdural haematoma and
oedema (Table 3). Increased S100B levels (on both first and second days of
hospitalization) were found in patients with GCS ≤ 8;
p = 0.024 and p = 0.031 (Table 4).
Combined injury (skull
fracture plus some other type) is significantly associated with GCS and
Marshall classification score (Table 3). Apart from this relationship, the GCS
score appears to be rather independent of injury type, while increased Marshall
classification is also associated with skull fracture, subdural and epidural
haematoma (Table 3).
Increased initial values of S100B also contributed to
risk prediction, but not at the cut‑off level >
0.105 µg/l. In fact, this level appears to be too low to serve as an
effective cut‑off point. In first day of hospitalization it was
exceeded by 100% of the risk patients (GOS = 1) but also by 91.7% of
the others. Consequently, the initial level of S100B attained only a very
low specificity at this point (Table 5). Objective ROC analysis revealed level
first day > 1.28 µg/l, sensitivity 83.3 and specificity 81.2%;
AUC = 0.752; p = 0.032 and second day of
hospitalization >0.54 µg/l, sensitivity 83.3 and specificity 97.2;
AUC = 0.880; p = 0.003.
parameters were confirmed as significant
predictors of GOS = 1:Marshall classification s. > II (AUC = 0.919;
p = 0.001), subdural haematoma (AUC == 0.782; p = 0.028), oedema (AUC = 0.824; p = 0.012), skull
fracture if combined with some other type of injury (AUC = 0.768;
p = 0.037).
GCS scores and cranial CT are the contemporary standards
for considering severity and visualizing brain damage after TBI . Despite
the information from clinical and radiological examinations, estimating
prognosis of TBI patients remains unmanageable in some instances.
The aims of this study
were to investigate correlations between initial levels of serum S100B protein
and mortality and computerized tomography (CT) findings, as well as to
establish whether there is an association between S100B and Glasgow outcome
scale (GOS) after six months. We anticipated that the initial biomarker
concentrations would represent the severity of the primary injury and the
accompanying immediate cell death. Peak concentration may, however, represent
ongoing cell death occurring as part of secondary injury. We might expect the
severity of secondary brain injury to have a stronger relationship with
outcome than primary injury.
The use of serum
biomarkers to assist in the prediction of outcome after TBI of all severities
is an area of active study in adults. The paediatric literature is far less
extensive. A study by Spinella et al  evaluated 27 children with
TBI of varying severities, measured a single S100B concentration as soon
as possible after injury and assessed outcome by dichotomized paediatric cerebral
performance category (PCPC), a six point scale similar to GOS. In this
study, there was a significant difference in S100B concentrations between
subjects with good and poor outcomes six months post‑injury, although there were no subjects with mild or moderate TBI with
poor outcome. Beers et al  measured serial serum concentrations of neuron‑specific enolase (NSE), S100B and myelin basic protein (MBP) in children
with acute TBI. Outcome was assessed by GOS, Vineland adaptive behaviour scale
(VABS) and an intelligence quotient (IQ) measurement. Student’s t‑tests, analysis of covariance, and nonparametric tests assessed between‑group differences for GOS, IQ and biomarkers. For all biomarkers and
time points, higher biomarker concentrations were associated with worse
outcomes. Our findings show that S100B levels significantly decrease in
a group of patients with final GOS > 1. This decrease started in the
second day of hospitalization and continued for the whole follow‑up period of six days. On the other hand, the cohort with GOS
value = 1 did not exhibit a significant decrease in S100B
values. Compared to adults, Lavička et al  demonstrated a rapid
decrease in serum levels in the first 6–12 hours after admission in
patients with good outcome, Vajtr et al  recorded increased S100B serum
levels in patients who died.
Ingebrigtsen et al 
found a correlation between serum S100B and post‑concussion symptoms in 50 patients with minor head injury and normal
CT scan. The same authors reported increased serum levels in 4 of
24 patients with minor head injury and normal CT scans where magnetic
resonance imaging revealed contusions in two cases . In severe head injury,
they observed increased S100B values in all cases; however, there are no data
available on the diagnostic and prognostic value of these observations. Data in
children are not available in these terms. In our study, we compared
S100 values, different outcome groups at six months and the categories of
Marshall classification of initial CT findings. This classification was
developed to allow more accurate predictive statements at the time of initial
patient evaluation regarding the likelihood of a fatal or nonfatal
outcome. This is widely accepted as a classification that facilitates
grading of injury severity and improves outcome prediction. Comparable to the
findings in the original study , we were able to demonstrate
a statistically significant relationship between a group of patients
with GOS = 1 and Marshall classification score > II
(no patient with GOS = 1 had Marshall score I or II),
a statistically significant relationship between a group of patients
with GOS = 1 and some type of injury diagnosed using CT
examination: intracranial bleeding, subdural haematoma, skull fracture,
subarachnoid bleeding, oedema, and multiple CT findings, namely skull fracture
in combination with some other injury type. Levels of S100B were increased in
patients with some specific types of injury, namely intracranial bleeding,
subdural haematoma and oedema. On the other hand, S100B initial value and S100B
second‑day are not significantly related to Marshall
classification score. This may indicate, that serum S‑100B protein adds diagnostic information.
The diagnostic accuracy
of a biomarker is a measure of its ability to accurately identify
a disease state. The relationship between sensitivity and specificity may
be determined using a receiver‑operating
characteristic curve (ROC) . An ROC curve is a graph of (sensitivity)
versus (specificity) and the diagnostic accuracy of a biomarker may be
quantified by using the area under the ROC curve (AUC). Good markers have AUC
values between 0.8 and 0.9, and excellent markers have AUC values between
0.9 and 1.0. In our study, increased initial values of S100B also contributed
to the risk prediction, but not at the cut‑off level >
In fact, this level
seems to be too low to be an effective cut‑off point. In the first
day of hospitalization it was exceeded by 100% of the risk patients
(GOS = 1) but also by 91.7% of the other patients. Consequently, the
initial level of S100B attained only very low specificity at this point.
Objective ROC analysis proposed levels of > 1.28 µg/l on the first day
of hospitalization and > 0.54 µg/l on the second.
Despite the wealth of
data indicating the use of diagnostic and prognostic biomarkers for traumatic
brain injury, there are still many outstanding issues in this area of research,
especially in paediatric patients.
The level of S100B was confirmed as a clinically
valuable indicator of the severity of injury and was proposed as an effective
predictor of risk outcome (GOS = 1). The commonly recommended cut‑off level of > 0.105 µg/l appears to be too low in this study.
Alternative cut‑off levels for the first and second day of
hospitalization were proposed and verified in ROC analysis (both with
sensitivity and specificity > 80%).
S100B values reveal
significant time‑related dynamics; they decreased significantly in the
group of patients with GOS > 1. The most significant decrease and
at the same time the most specific differentiation of risk patients
(GOS = 1) was observed in the second day of hospitalization.
We would like to thank the nurses of the Department of Anaesthesia and Intensive Care, University Children’s Hospital, Brno, for their efficient care and precise documentation.
Přijato k recenzi: 9. 10. 2009
Přijato do tisku: 23. 11. 2009
MUDr. Jiří Žurek
Klinika dětské anesteziologie a resuscitace LF MU a
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