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THE S-100B SUBSTUDY OF THE GLUTAMICS-TRIAL: ELEVATION OF S-100B IS
LINKED TO AORTIC CALCIFICATION AND POSTOPERATIVE CONFUSION
Mårten Vidlund3 MD, Jonas Holm1 MD, Erik Håkanson2, MD PhD, Örjan Friberg3 MD PhD, Lena
Sunnermalm3 MD, Farkas Vanky1 MD PhD, Rolf Svedjeholm1, MD PhD
Departments of Cardiothoracic Surgery1 and Cardiothoracic Anesthesia2, Linköping Heart Center,
University Hospital, Linköping, Sweden. Department of Cardiothoracic Surgery and Anesthesiology3,
University Hospital Örebro, Sweden.
Running head: The S-100B substudy of the GLUTAMICS-trial
Key words: XX
Word count: XX
Corresponding author:
Dr Rolf Svedjeholm, Department of Cardiothoracic Surgery, University Hospital, SE-581 85
Linköping, Sweden. Phone: +4613224825 Fax: +4613100246
E-mail: [email protected]
ABSTRACT
Background: Although concerns have been raised there is no evidence for increased risk of clinically
evident neurological injury in cardiac surgical patients receiving glutamate by cardioplegia or
intravenous infusion. As an early safety measure in the GLUTAMICS trial a prespecified subgroup of
patients were analyzed with regard to postoperative S-100B levels to detect potential subclinical
neurological injury related to glutamate infusion. Furthermore, we wanted to assess the relationship
between S-100B and clinical signs of neurological injury and established risk factors for stroke.
Methods: 69 patients undergoing CABG for unstable coronary syndrome were randomized to
intravenous infusion of glutamate (n=35) or saline (n=34) perioperatively. All except four procedures
were done with cardiopulmonary bypass. Plasma levels of S-100B were obtained on the third
postoperative day. Clinical data were collected prospectively.
Results: There were no differences in postoperative S-100B levels between the glutamate group
(0.079 + 0.034 µg/L) and the control group (0.090 + 0.042 µg/L; p=0.245). There were no patients
with stroke and no mortality. Three patients in the control group and two in the glutamate group had
postoperative confusion. These patients had significantly elevated S-100B compared to those without
confusion (0.132 + 0.047 v 0.081 + 0.036; p=0.003). Overall 21 patients had elevated S-100B (≥ 0.10
ug/L) and these patients had significantly higher Euroscore, more calcifications in the ascending aorta
detected by epiaortic scanning and more often had postoperative confusion.
Conclusions: Postoperative elevation of S-100B was linked to calcification of the ascending aorta and
postoperative confusion but not to glutamate infusion.
INTRODUCTION
Glutamate has been claimed to protect the heart from ischemia and facilitate metabolic and
hemodynamic recovery after ischemic insults[1-3]Glutamate enhancement of cardioplegic solutions
has therefore been advocated[3]We have administered glutamate as an intravenous infusion as it
provides the opportunity to supply the heart with substrate during the preoperative and postoperative
phase where myocardial ischemia appears to be particularly detrimental in association with CABG[4]
With this approach encouraging results have been achieved both in study populations and in clinical
practice[5, 6]Based on this experience the GLUTAMICS-trial was initiated. The primary aim of this
ongoing trial (ClinicalTrials.gov Identifier: NCT00489827) is to determine the cardioprotective role of
intravenous glutamate infusion given perioperatively to patients operated for unstable coronary artery
disease.
A major concern with the use of glutamate is that it has been claimed to act as an excitotoxin under
certain conditions and to participate in events leading to neurological damage[7] Glutamate
administration has been shown to cause neurological injury in rodents but not in primates due to the
blood-brain barrier that prevents passage of exogenous glutamate to the brain[8-10]The role of
exogenously administered glutamate in patients undergoing cardiac surgery with cardiopulmonary
bypass and potential blood brain barrier dysfunction remains to be clarified. Available data have not
suggested any increase in clinically evident neurological injury when glutamate enhanced cardioplegic
solutions or intravenous infusions have been implemented in clinical practice[11, 12]
As an early safety measure in the GLUTAMICS trial a prespecified subgroup of patients were
analyzed with regard to postoperative S-100B levels to detect potential subclinical neurological injury
related to glutamate infusion. Furthermore, we wanted to assess the relationship between S-100B and
intraoperative epiaortic scanning of the ascending aorta, clinical signs of neurological injury and
established risk factors for stroke.
PATIENTS AND METHODS
The GLUTAMICS-trial is a prospective randomized controlled trial (ClinicalTrials.gov Identifier:
NCT00489827) evaluating metabolic intervention with intravenous glutamate infusion in association
with surgery for unstable coronary artery disease. The study is externally randomized and an external
professional statistician was provided with the randomization codes for the patients in the S-100B
substudy. Analysis was limited to prespecified variables reported in this article and blinded to the
investigators. A prespecified subgroup of 70 consecutive patients were enrolled in the S-100B
substudy of GLUTAMICS-trial at the University Hospital in Linköping. Sampling according to
protocol was done in 69 patients. Patients were randomized to blinded intravenous infusion of
glutamate 0.125M solution or saline at a rate of 1.65 ml/kg and hour commencing at the induction of
anesthesia. As the by cardioplegia arrested heart is reported to leak glutamate rather than extract
exogenous glutamate, the infusions were temporarily stopped during aortic cross-clamping[13, 14].
Infusions were resumed after declamping of the aorta and continued for a further 2 hours or up to a
maximum of 500 ml of study solution.
Clinical data were prospectively recorded in an institutional database. Epiaortic scanning of the
ascending aorta with ultrasound XX was done intraoperatively before cannulation for cardiopulmonary
bypass in all patients. Results were recorded according to a protocol. Aortic calcification was
classified by the operating surgeon as none, mild, moderate, severe or completely calcified. Sampling
for S-100B was done on the third postoperative day. S-100B was analyzed with commercially
available automated electrochemiluminescense immuno assay Elecsys® S-100, Roche Diagnostics.
Epiaortic scanning ( ekomaskin,probe osv……) ……. XX
Ethics
After written informed consent the patients were enrolled in the study. The study was performed
according to the Helsinki Declaration of Human Rights and was approved by the ethics committee for
medical research at the University Hospital of Linköping.
Clinical management
Anestesi Erik XX
The patients underwent surgery using standard techniques with cardiopulmonary bypass (CPB) and
aortic cross clamping. Four procedures were performed off pump. This decision was based on findings
from epiaortic scanning demonstrating severe calcifications. Ringer's acetate and mannitol was used
for priming the extracorporeal circuit. Moderate hemodilution (hematocrit 20 - 25%) and mild
hypothermia (33-36 C) were employed. Antegrade or combined ante- and retrograde delivery of a
cold crystalloid cardioplegic solution (PlegisolTM, Abbot, IL, US) supplemented with procaine
hydrochloride was used for myocardial protection. Weaning from CPB was started at a rectal
temperature of 35-36 C. Heparin was neutralized with protamine chloride. Shed mediastinal blood
was discarded and Ringer's acetate was used for volume substitution postoperatively.
Definitions
S-100B levels above 0.010 µg/L were defined as elevated.
Postoperative confusion was defined as XX
Aortic calcification XX
Stroke was defined as focal neurological deficit persisting for more than 24 hours or depression of
consciousness or confusion if associated with signs of cerebral injury on CT-scan. Patients with
clinical suspicion of neurological injury underwent CT-scan of the brain.
Statistical analysis
Statistical analysis done by an external professional statistician and analysis was blinded to the
investigators. Fisher’s exact test was used for comparison of dichotomous variables and Students t-test
or Mann-Whitney U test was used as appropriate for comparison of continuous variables. Statistical
significance was defined as p<0.05.
RESULTS
There was no statistically significant difference of S-100B levels between the patients who received
intravenous glutamate infusion and the patients who received placebo (0.079 ± 0.034 µg/L v 0.090 ±
0.042 µg/L; Figure 1). Preoperative and intraoperative data are given in Table 1. There was no
mortality and no patient suffered from stroke.
Five patients, two in the glutamate group and three in the placebo group had postoperative confusion
and these patients had significantly elevated S-100B compared to those without confusion (0.132 ±
0.047 µg/L v 0.081 ± 0.036 µg/L; p=0.003). Out of the five patients with postoperative confusion,
three patients had mild symptoms and recovered completely before discharge and two patients had
more persistent confusion and thus underwent a CT-scan of the brain which did not show any new
lesions. These two patients also recovered completely from the postoperative confusion XX.
Out of 69 patients overall, 21 showed elevated S-100B. These 21 patients had significantly higher
Euroscore, more calcification of the ascending aorta, and had more often postoperative confusion.
Pre-, intra-, and postoperative data in the group with elevated S-100B versus the group with normal S100B is shown in Table 2.
DISCUSSION
The main finding of this study was that elevation of plasma S-100B after surgery for unstable coronary
artery disease was linked to occurrence of postoperative confusion and intraoperative findings of aortic
calcification but not to intravenous glutamate infusion. Based on previous clinical experience and the
present study we did not find any reasons to discontinue the GLUTAMICS-trial because of safety
aspects related to glutamate infusion and potential neurological injury. However, surveillance of all
neurological events will remain a key safety issue throughout the trial.
Several markers for neurological injury have been introduced. S-100B, Neurone-specific Enolase
(NSE) and glial fibrillary acidic protein (GFAP) are those proteins investigated most often as surrogate
markers of brain damage[15]. S-100B, a calcium-binding glial protein, is the marker that has received
greatest attention in association with cardiac surgery[16]. Initial enthusiasm with this marker was
subdued because of conflicting results mainly related to contamination from extra-cerebral sources
such as fat, muscle, skin and bone marrow. In association with cardiac surgery shed mediastinal blood
was found to contain high concentrations of S-100B[16, 17]. However, due to the short biological
half-life of S-100B, reported to be only 25 minutes after cardiac surgery, the levels related to
contamination is of importance only during the first hours after cardiac surgery. Accordingly, the S100B concentrations obtained 24 hours or later after cardiac surgery show consistent correlation with
clinical neurological findings. S-100B sampled 2 days postoperatively correlates with the size of MRIdetected brain injury[16, 18]. Furthermore, it has been demonstrated that elevations of S-100B two
days after cardiac surgery is a powerful predictor of impaired late survival regardless of clinical signs
of stroke. In the setting of carotid artery surgery it has been reported that S-100B may be a useful
marker of subclinical neurological injury[19].
A normal S100B level reliably predicts the absence of significant CNS injury[20]. The samples in our
study were obtained on the third postoperative day and thus a number of patients with S-100B
elevations during the first two postoperative days may have been missed[19]. However, there was no
relation between the use of glutamate infusion and elevated postoperative S-100B levels suggesting
that administration of exogenous glutamate was not linked to subclinical neurological damage. In
contrast, we found a clear relationship between S-100B levels and aortic calcification. This may
indicate subclinical neurological injury, possibly due to microembolisation from the ascending aorta
which is known to occur during aortic manipulation such as cannulation or aortic cross-clamping[21].
Trans-cranial Doppler studies have demonstrated similar findings in association with carotid
endarterectomy. In this setting it has been shown that both perioperative micro-embolisation and
postoperative cognitive dysfunction, suggesting subtle cerebral injury, is associated with a rise in S100B [22, 23]. Cognitive dysfunction was not assessed with neuropsychometric tests in our study but
patients with postoperative confusion had significantly elevated S-100B levels. This finding supports
previous evidence that minor neurological injury can be detected by S-100B.
The role of S-100B in clinical practice remains to be established but the results obtained in our study
argue against a role for exogenously administered glutamate in the evolution of neurological injury in
association with cardiac surgery. This information is of importance not only for potential future
application of glutamate in cardiac surgery but also of more general interest as most commercially
available amino acid solutions contain glutamate. This issue has raised concerns particularly in
neurosurgical care but the limited data available did not provide any evidence of a deleterious effect of
such amino acid solutions[24] Dosage of glutamate is obviously an issue to consider. In the
neurosurgical study an amino-acid solution containing 3,75 g glutamate /L infused over 24 hours
doubled plasma levels of glutamate[24]. The dosage of glutamate in the GLUTAMICS-trial was based
on studies demonstrating that an infusion rate sufficient to increase arterial whole blood levels by twothree fold also was sufficient to meet the myocardial demands or to saturate myocardial capacity to
extract glutamate from the circulation[25]. Glutamate plays a key role in myocardial metabolism
during ischemia and early after coronary artery bypass surgery the myocardium extracts approximately
40-50% of circulating glutamate[26]. This is probably due to a relative substrate deficiency as it has
been shown that freely available glutamate in the cytosol of the cardiomyocytes is rapidly depleted
during ischemia and cardioplegic arrest[27]. Duration of exposure to elevated glutamate levels is also
an issue to consider and is mainly limited to the period of infusion as plasma levels rapidly normalize
due to high extraction from the heart and skeletal muscle[25].
To conclude, elevation of plasma S-100B after surgery for unstable coronary artery disease was linked
to occurrence of postoperative confusion and intraoperative findings of aortic calcification but not to
intravenous glutamate infusion.
REFERENCES
1. Rau EE, Shine KI, Gervais A, Douglas AM, Amos EC. Enhanced mechanical recovery of
anoxic and ischemic myocardium by amino acid perfusion. AmJPhysiol 1979;236:H873-H9.
2. Pisarenko OI. Mechanisms of myocardial protection by amino acids: facts and hypotheses.
[Review] [88 refs]. Clinical & Experimental Pharmacology & Physiology 1996;23:627-33.
3. Lazar HL, Buckberg GD, Manganaro AJ, Becker H. Myocardial energy replenishment and
reversal of ischemic damage by substrate enhancement of secondary blood cardioplegia with
amino acids during reperfusion. JThoracCardiovascSurg 1980;80:350-9.
4. Svedjeholm R, Hakanson E, Vanhanen I. Rationale for metabolic support with amino acids
and glucose-insulin-potassium (GIK) in cardiac surgery. Ann Thorac Surg 1995;59:S15-22.
5. Svedjeholm R, Huljebrant I, Hakanson E, Vanhanen I. Glutamate and high-dose glucoseinsulin-potassium (GIK) in the treatment of severe cardiac failure after cardiac operations.
Ann Thorac Surg 1995;59:S23-30.
6. Svedjeholm R, Vanhanen I, Hakanson E, Joachimsson PO, Jorfeldt L, Nilsson L. Metabolic
and hemodynamic effects of intravenous glutamate infusion early after coronary operations. J
Thorac Cardiovasc Surg 1996;112:1468-77.
7. Greenamyre JT, Porter RH. Anatomy and physiology of glutamate in the CNS. Neurology
1994;44:S7-13.
8. Sacks W, Sacks S, Brebbia DR, Fleischer A. Cerebral uptake of amino acids in human
subjects and rhesus monkeys in vivo. JNeurosciRes 1982;7:431-6.
9. Daabees TT, Finkelstein MW, Stegink LD, Applebaum AE. Correlation of glutamate plus
aspartate dose, plasma amino acid concentration and neuronal necrosis in infant mice. Food
ChemToxicol 1985;23:887-93.
10. Fernstrom JD. Pituitary hormone secretion in normal male humans: acute responses to a
large, oral dose of monosodium glutamate. J Nutr 2000;130:1053S-7S.
11. Loop FD, Higgins TL, Panda R, Pearce G, Estafanous FG. Myocardial protection during
cardiac operations. Decreased morbidity and lower cost with blood cardioplegia and coronary
sinus perfusion. J Thorac Cardiovasc Surg 1992;104:608-18.
12. Svedjeholm R, Hakanson E, Szabo Z, Vanky F. Neurological injury after surgery for
ischemic heart disease: risk factors, outcome and role of metabolic interventions. Eur J
Cardiothorac Surg 2001;19:611-8.
13. Kimose HH, Ravkilde J, Helligs” P, et al. Myocardial loss of glutamate after cold
chemical cardioplegia and storage in isolated blood-perfused pig hearts.
ThoracCardiovascSurg 1993;41:93-100.
14. Suleiman MS, Fernando HC, Dihmis WC, Hutter JA, Chapman, Ra. A loss of taurine and
other amino acids from ventricles of patients undergoing bypass surgery. British Heart Journal
1993;69:241-5.
15. Marchi N, Rasmussen P, Kapural M, et al. Peripheral markers of brain damage and bloodbrain barrier dysfunction. Restor Neurol Neurosci 2003;21:109-21.
16. Jonsson H. S100B and cardiac surgery: possibilities and limitations. Restor Neurol
Neurosci 2003;21:151-7.
17. Anderson RE, Hansson LO, Nilsson O, Liska J, Settergren G, Vaage J. Increase in serum
S100A1-B and S100BB during cardiac surgery arises from extracerebral sources. Ann Thorac
Surg 2001;71:1512-7.
18. Jonsson H, Johnsson P, Birch-Iensen M, Alling C, Westaby S, Blomquist S. S100B as a
predictor of size and outcome of stroke after cardiac surgery. Ann Thorac Surg 2001;71:14337.
19. Johnsson P, Backstrom M, Bergh C, Jonsson H, Luhrs C, Alling C. Increased S100B in
blood after cardiac surgery is a powerful predictor of late mortality. Ann Thorac Surg
2003;75:162-8.
20. Bloomfield SM, McKinney J, Smith L, Brisman J. Reliability of S100B in predicting
severity of central nervous system injury. Neurocrit Care 2007;6:121-38.
21. Lund C, Hol PK, Lundblad R, et al. Comparison of cerebral embolization during off-pump
and on-pump coronary artery bypass surgery. Ann Thorac Surg 2003;76:765-70; discussion
70.
22. Brightwell RE, Sherwood RA, Athanasiou T, Hamady M, Cheshire NJ. The neurological
morbidity of carotid revascularisation: using markers of cellular brain injury to compare CEA
and CAS. Eur J Vasc Endovasc Surg 2007;34:552-60. Epub 2007 Aug 24.
23. Connolly ES, Jr., Winfree CJ, Rampersad A, et al. Serum S100B protein levels are
correlated with subclinical neurocognitive declines after carotid endarterectomy.
Neurosurgery 2001;49:1076-82; discussion 82-3.
24. Stover JF, Kempski OS. Glutamate-containing parenteral nutrition doubles plasma
glutamate: a risk factor in neurosurgical patients with blood-brain barrier damage? Crit Care
Med 1999;27:2252-6.
25. Vanhanen I, Svedjeholm R, Hakanson E, et al. Assessment of myocardial glutamate
requirements early after coronary artery bypass surgery. Scand Cardiovasc J 1998;32:145-52.
26. Svedjeholm R, Ekroth R, Joachimsson PO, Ronquist G, Svensson S, Tyden H. Myocardial
uptake of amino acids and other substrates in relation to myocardial oxygen consumption four
hours after cardiac operations. J Thorac Cardiovasc Surg 1991;101:688-94.
27. Pisarenko OI, Oleynikov OD, Shulzhenko VS, Studneva IM, Ryff IM, Kapelko VI.
Association of myocardial glutamate and aspartate pool and functional recovery of
postischemic heart. BiochemMedMetabBiol 1989;42:105-17.
Table 1
Preoperative data
Age (years)
Females %)
Weight (kg)
Length (cm)
BMI (kg/m2 )
Active Smokers %
Blood haemoglobin (g/l)
serum Creatinine µmol/l
Hypertension %
Diabetes mellitus %
COPD %
Cerebrovascular disease %
Neurological dysfunction %
Peripheral artery disease %
Previous vascular surgery %
Carotid artery stenosis %
Previous myocardial infarction %
Recent myocardial infarction (<3 weeks)%
Atrial fibrillation %
Moderate–severe left ventricular dysfunction %
CCS class IV %
3-vessel disease %
Left main stem stenosis %
Emergency procedure %
Euroscore
Intraoperative data
Cacification of ascending aorta %
Measures undertaken because of scanning %
No. of bypasses
OPCAB %
Use of LIMA %
Other operation than isolated CABG %
AVR%
Mitral plasty%
Aortic X-clamp time min
Single-clamp%
Time on ECC min
Neurological outcome
Stroke%
Confusion%
Control
(n=34)
Treatment
(n=35)
p-value
66 ± 11
9
82.513.2
173.66.3
27.44.4
18.8
13914
10515
35.3
29.4
2.9
2.9
0
6.3
5.9
6.7
76.5
58.8
11.8
26.5
84.0
82.4
38.2
0
4.83.1
689
20
82.314.0
174.58.1
27.04.1
30.3
13815
10719
35.3
40.0
11.4
11.4
2.9
12.5
11.4
0
80.0
77.1
11.4
20.0
80.6
77.1
54.3
8.6
5.63.5
0.40
0.31
0.96
0.62
0.69
0.39
0.67
0.66
1.00
0.45
0.36
0.36
1.00
1.00
0.67
0.23
0.78
0.13
1.00
0.58
1.00
0.77
0.23
0.24
0.30
32.4
23.5
4.11.4
2.9
97.1
5.9
2.9
2.9
5922
81.8
8429
29.4
23.5
3.81.3
8.6
97.1
8.6
2.9
2.9
6423
71.9
9130
1.00
1.00
0.46
0.61
1.00
1.00
1.00
1.00
0.34
0.39
0.37
0
8.8
0
5.7
1.00
0.67
Table 2
Preoperative data
Age (years)
Females %)
Weight (kg)
Length (cm)
BMI (kg/m2 )
Active Smokers %
Blood haemoglobin (g/l)
serum Creatinine µmol/l
Hypertension %
Diabetes mellitus %
COPD %
Cerebrovascular disease %
Neurological dysfunction %
Peripheral artery disease %
Previous vascular surgery %
Carotid artery stenosis %
Previous myocardial infarction %
Recent myocardial infarction (<3 weeks)%
Atrial fibrillation %
Moderate–severe left ventricular dysfunction %
CCS class IV %
3-vessel disease %
Left main stem stenosis %
Emergency procedure %
Euroscore
Intraoperative data
Cacification of ascending aorta %
Measures undertaken because of scanning %
No. of bypasses
OPCAB %
Use of LIMA %
Other operation than isolated CABG %
AVR%
Mitral plasty%
Aortic X-clamp time min
Single-clamp%
Time on ECC min
Neurological outcome
Stroke%
Confusion%
Normal s-100B
(n=48)
Elevated s-100B
(n=21)
p-value
659
8.3
82.713.4
175.36.2
26.83.6
19.6
14114
10616
29.8
31.3
8.3
6.3
2.1
6.8
4.2
2.3
77.1
62.5
14.6
25.0
80.0
83.3
43.8
4.2
4.53.1
7010
28.6
81.614.0
171.28.7
28.05.4
36.8
13416
10620
47.6
42.9
4.8
9.5
0
30.0
19.0
5.3
80.9
81.0
4.8
19.0
87.5
71.4
52.4
4.8
7.03.1
0.10
0.057
0.76
0.029
0.28
0.21
0.07
0.87
0.18
0.42
1.00
0.64
1.00
0.022
0.07
0.52
1.00
0.17
0.42
0.76
0.71
0.33
0.60
1.00
0.003
21.3
19.1
4.01.3
2.1
97.9
4.2
0
2.1
5921
78.7
8529
52.4
33.3
3.81.3
14.3
95.2
14.3
9.5
4.8
6726
72.2
9330
0.021
0.23
0.58
0.08
0.52
0.16
0.09
0.52
0.21
0.74
0.35
0
2.1
0
19.0
1.00
0.027