Cognitive deficit occurs commonly after coronary artery bypass surgery (CABG). Such dysfunction is multifactorial in origin and may be due to cerebral embolization , cerebral ischaemia-reperfusion , cerebral hyperthermia after discontinuation after cardiopulmonary bypass (CPB)  and the systemic inflammatory response induced by CPB . Nitric oxide (NO) is an important mediator of cerebral ischaemia-reperfusion and the cerebral inflammatory response in this setting. Adverse neurological outcome is classified as: Type I (focal neurological injury, stupor or coma at discharge) and Type II (deterioration in intellectual function, memory deficit or seizures). The incidence of a Type I deficit is of the order of 3% . Cognitive dysfunction is reported in 53% of patients at discharge, 36% at 6 weeks and 42% at 5 yr . Adverse neurological outcome has important financial, social and care implications for patients, their families and society .
The identification of an effective biochemical marker in the assessment of intervention intended to decrease postoperative cognitive deficit would be clinically valuable. S100β protein (a stable product derived largely from glial tissue) has been most extensively studied but its specificity as a marker of neurological injury associated with cardiac surgery and CPB has been challenged by its identification in cardiotomy suction .
There are three known isoenzymes of nitric oxide synthase (NOS) that oxidize arginine to NO and citrulline. In animal models, acute global ischaemia upregulates neuronal NOS within minutes , upregulates inducible NOS within hours  and results in a very marked production of NOS mRNA and associated NO activity . Such an insult also results in upregulation of endothelial NOS . The resulting NO activity is an important mediator of the subsequent cerebral injury but may also play a role in the repair process . NO promotes neural injury by causing oxidative injury, energy depletion, DNA damage, inhibiting DNA synthesis and triggering programmed cell death .
NO is difficult to measure in vivo because of its short half-life. However, the stable and inactive end products of NO oxidation, nitrate (Symbol) and nitrite (Symbol), can be quantified in biological fluids and provide a useful method of indirectly estimating endogenous NO production . Increased cerebrospinal fluid (CSF) and plasma concentrations of NO oxidation products have been demonstrated to be associated with head injury severity score . In this study by Clarke and colleagues, however, there was no difference over time in post-traumatic plasma Symbol and Symbol concentrations. Increased CSF NO oxidation products and a correlation to infarct volume (coefficient 0.39, P < 0.001) has been demonstrated in stroke patients .
We hypothesized that perioperative plasma concentrations of NOx (Symbol plus Symbol) are associated with cognitive dysfunction after CABG. It may thus serve as an effective biomarker in the assessment of intervention intended to decrease postoperative cognitive deficit.
With Institutional Ethics Committee approval and written informed consent, 36 adult ASA III-IV patients undergoing elective CABG, were studied in a prospective observational trial. Exclusion criteria were: psychiatric or active neurological disease; severe visual or auditory disorders; alcoholism; active renal or liver disease. Patients receiving drugs capable of NO release (e.g. Symbol) 24 h before surgery were also excluded. Patients were recruited sequentially (if no contraindication) with advanced age not being an exclusion criterion. All patients were without food for at least 12 h before surgery as dietary conditions markedly influence plasma NOx .
Oral lorazepam 0.02-0.03 mg kg−1 was administered to patients as a premedicant 2 h before surgery. General anaesthesia was induced with fentanyl 15-20 μg kg−1 and propofol 0.5-1.0 mg kg−1 and maintained using a propofol infusion (2-3 mg kg−1 h−1). Muscle relaxation was achieved with pancuronium 0.1 mg kg−1. Patients received morphine 1-3 mg h−1 and midazolam 1-3 mg h−1 while intubated postoperatively in the intensive care unit (ICU).
CPB was instituted using a hollow fibre oxygenator (Cobe Optima, Sorin Biomedica UK Ltd, Gloucester, UK) with crystalloid priming and non-pulsatile flow at mild hypothermia (32-34°C). Pump priming consisted of Ringers' lactate 2000 mL, sodium bicarbonate 8.4% 50 mL and mannitol 20% 3 mL kg−1. The pump flow rate was 2.4 L min−1 m−2 during the period of aortic cross clamping. During CPB, mean arterial pressure was maintained at 55-70 mmHg and haematocrit at 20-25%. Cardiotomy suction, 0.5-1 L min−1 (Adult Sump sucker, Lifestream International, TX, USA) was used from pericardiotomy to closure of pericardium. A cell saver device was not used.
One of two surgeons operated on each patient studied. One surgeon (A. O'D.) administered intermittent antegrade and retrograde cold (15°C) blood cardioplegia and completed vein grafts with a DLP octopus retractor (Medtronic Europe, Tolochenaz, Switzerland). A single cross clamp was used to complete distal and proximal anastomosis. The other surgeon used antegrade crystalloid cold (4°C) cardioplegia with proximal anastamosis after declamping using an aortic side-biting clamp. St. Thomas solution was used as cardioplegia (Na+ 120 mmol, K+ 16 mmol, Mg2+ 16 mmol, Ca2+ 1.2 mmol, Symbol 10 mmol and Cl− 160 mmol). Aortic venting was used in all patients.
Mood was assessed by the Hospital Anxiety and Depression (HAD) Scale . The diagnosis of delirium was based on the Diagnostic and Statistical Manual of Mental Disorders (Third Edition - Revised) criteria (DSM-III-R), and the Mini Mental State Examination. Delirium was assessed on a daily basis until hospital discharge. A detailed neurological examination was also performed daily. A battery of cognitive tests (including those recommended by the Statement of Consensus 1995 ) was administered on the day before, 4 days and 3 months after surgery. Subjects were unaware of their original scores. A single clinical psychologist, blinded to NOx concentrations, performed all cognitive assessments under standard conditions. Cognitive assessment was designed to take approximately 40 min to complete.
Domains of cognitive function assessed and the tests used were as follows: Verbal memory: Rey Auditory Verbal Learning Test (RAVLT). Attention: Trail-Making Test parts A and B (TMT A & B). Motor speed: The Purdue Pegboard test (Pegs Tot). Executive function/Verbal fluency: Controlled Oral Word Association Test (COWAT). Psychomotor speed: Digit Symbol Test (Dig Symb).
Spouses of patients included in study were also recruited if they consented and no exclusion criteria were met. The same exclusion criteria as applied to patients pertained. The battery of cognitive tests was administered to spouses at the times described above for patients.
Definition of cognitive deficit
Using the methodology outlined by Jacobson and Traux , the reliable change (RC) index was calculated for each neuropsychological measure using the baseline and follow-up data of the control subjects. First, the test-retest reliability coefficient (rxx) was computed for each measure (Pearson correlation coefficient between preoperative and postoperative scores), from which the standard error of measurement (SEm) was calculated using the formula SEm = SD1 (Symbol), where SD1 is the SD of the preoperative control score. The standard error of the difference (SEdiff) then was calculated using the formula SEdiff = Symbol. The SEdiff describes the distribution of changes in scores that would be expected if no true change had occurred. To establish a 90% RC confidence interval (CI) (two-tailed prediction) the SEdiff was multiplied by ± 1.64SD1. A correction representing the practice effect then was added to the two-tailed cut-off points. The practice effect was calculated for each measure as the mean of the difference between each pair of pre- and postoperative control scores. Thus, an RC 90% CI was calculated from this formula for each variable: RC interval = (SEdiff) × (±1.64SD1) + practice effect. The change index limits were rounded to the nearest whole number outside the 90% RC interval. For each neuropsychological measure, a postoperative minus preoperative difference score was calculated for each patient. When this score fell outside the RC intervals, a significant change in performance on that measure was considered to have occurred.
Plasma samples were obtained for NOx estimation immediately prior to induction of anaesthesia (t0), 10 min after commencement of CPB (t1), 5 min after removal of the aortic cross clamp (t2), immediately prior to protamine administration (t3), 10 min after protamine administration (t4) and 2 (t5), 4 (t6), 12 (t7), 24 (t8) and 48 h (t9) after arrival in the ICU. Blood samples were drawn from a radial arterial catheter at all times except at 48 h when a central venous sample was obtained because arterial access was uncommon at 48 h. The relationship between arterial and venous NOx concentrations associated with CABG and CPB has not been described. Blood samples (4 mL) were collected in lithium heparin bottles (Greiner bio-one, Kremsmunster, Austria) and placed immediately on ice. Following centrifugation at 4000g for 5 min at 4°C, plasma was stored at −80°C. NOx concentrations were determined within a month of storage. Plasma NOx is affected by renal impairment . Plasma creatinine concentrations were measured at each sampling time.
NOx were measured using a Nitric Oxide Chemiluminescent Analyzer, Sievers 280 NOA™ (Sievers Instruments, Boulder, CO, USA). Acid and reducing agents (vanadium III chloride in 1 M hydrochloric acid) at 95°C were added to samples, converting Symbol and Symbol to NO. An inert gas was then used to purge NO from solution, which was detected by chemiluminescence, with a sensitivity limit of detection of 20 nmol L−1. Standard curves were constructed and the amount of Symbol and Symbol in plasma was determined. NOx concentrations were estimated sequentially in batches.
Differences between pre- and postoperative concentrations of NOx were compared using two-tailed, paired t-test. Differences in concentrations between deficit and non-deficit groups were compared using two-tailed unpaired t-test. Association between glyceryl trinitrate (GTN) therapy and plasma creatinine or plasma NOx concentrations, respectively, was measured using Pearson correlation. Categorical variables were compared using the χ2-test. P < 0.05 was considered significant. Patient characteristics data and biochemical concentrations are reported as mean (SD or range).
Thirty-six adult ASA II-III patients undergoing CABG were studied. Ten patient spouses were also studied as a control group. No patient had a Type I adverse neurological outcome. Five patients had postoperative delirium (14%). On the fourth postoperative day three patients could not be assessed due to respiratory complications (increased FiO2 requirement) and were assessed on the following (fifth) postoperative day. These data have been included in the tables with figures referring to the fourth postoperative day. One patient was excluded from the 3-month assessment due to a non-operative-related illness.
Patients who did not develop a postoperative cognitive dysfunction and controls were similar in terms of patient characteristics and preoperative cognitive test scores (Table 1). Compared to the non-deficit group (4 days postoperatively), patients in whom a postoperative cognitive dysfunction was identified were older (P = 0.02), and scored more poorly on preoperative cognitive tests (Table 1). Preoperatively patients and controls were similar in terms of depression and anxiety scores (Table 1).
The data obtained from control subjects used to calculate the RC indices for the interval from preoperatively to 4 days postoperatively are summarized in Table 2. RC indices were similarly calculated for the interval from preoperatively to 3 months postoperatively. Each cognitive test showed acceptable test-retest reliability, ranging from 0.68 (Pegs Tot) to 0.86 (COWAT). Postoperative cognitive dysfunction (defined by a change in one or more cognitive domains using the RC method) was present in 22/36 (61%) and in 16/35 (49%) patients, 4 days and 3 months postoperatively. The tests most sensitive to postoperative decline (4 days postoperatively) were TMT A & B and RAVLT Tot. The least sensitive test was the COWAT (Table 3). Three months postoperatively the deficit and non-deficit groups were similar in terms of depression (1.6 ± 1.8 vs. 1.8 ± 1.7, P = 0.7) and anxiety scores (2.7 ± 3.0 vs. 2.8 ± 2.3, P = 0.9).
Plasma NOx (μmol L−1) was less after end of CPB (t3 and t4) than preoperatively (P = 0.006) and greater 24 h after arrival in ICU than at end of CPB (P = 0.03) (Table 4). Perioperative plasma concentrations of NOx were greater in the group with deficit at 4 days postoperatively (n = 22) as compared to the non-deficit group (n = 14) at seven time points (Table 5). Patients with deficit at 3 months postoperatively (n = 16) had greater plasma NOx compared to the non-deficit group (n = 19) at five time points (Table 5). There was no difference, however, in magnitude of alteration of plasma NOx over time (t0-t8) between the deficit and non-deficit groups.
The proportion of patients taking nitroso-donor drugs up to 24 h preoperatively, was similar in deficit and non-deficit groups (P = 0.9) (Table 1). The decrease in plasma NOx was similar in patients who received GTN intraoperatively (4.62 (0.4-15.1) μmol L−1, n = 19) and those who did not (5.1 (2.2-1.5) μmol L−1, n = 17), (P = 0.7). Plasma NOx increased both in patients who received GTN postoperatively (5.9 (−18.8-28.8) μmol L−1, n = 32) and in those who did not (1.1 (−0.4-9.7) μmol L−1, n = 4) (P = 0.56). The proportion of patients with cognitive deficit 4 days after surgery in the groups who did and did not receive GTN intraoperatively (11/19 vs. 11/17, P = 0.67) and postoperatively (20/32 vs. 2/4, P = 0.57) was similar.
The volumes of fluid administered prior to CPB were similar in the deficit and non-defict groups (1.5 ± 0.3 L vs. 1.4 ± 0.2 L, P = 0.4). When plasma NOx concentrations 10 min after start of CPB (t1) and 10 min after protamine administration (t4) were standardized to haemoglobin (Hb) change (from t0 to t1 and t4, respectively) statistically significant differences remained between the groups P = 0.003 and P = 0.002. Plasma creatinine concentrations demonstrated no significant correlation with plasma NOx 5 min after removal of aortic cross clamp (r = 0.32, P = 0.18), or at 24 h postoperatively (r = 0.37, P = 0.14).
Prospective longitudinal studies now indicate that cognitive dysfunction following CABG is common, consistent and persistent . Our finding of 61% and 49% incidences of cognitive deficit at 4 days and 3 months postoperatively are greater than that reported by Newman and colleagues (53% of patients at discharge and 36% at 6 weeks) . Potential explanations for this are due to our use of the RC index, a validated and sensitive method to define deficit and different times of postoperative assessment chosen. The greatest incidences of cognitive deficit were observed for immediate memory (RAVLT) and attention (TMT A & B). This finding is consistent with those of Smith and colleagues .
The main finding of this study was that an association between perioperative plasma NOx concentrations and cognitive dysfunction after CABG could not be established. The plasma NOx concentrations reported in this study are similar to those reported by Mathie and colleagues . There was a time course similarity in NOx elevations for both deficit and non-deficit groups. If certain perioperative cerebral insults causing elevated NO predispose to postoperative cognitive defects, then it might be expected that the pattern of elevation would vary accordingly. This did not occur in the deficit and non-deficit groups in this study. It thus appears that the only difference between the groups was preoperatively and that this difference was carried through the study period.
It is unclear how NO metabolism in tissues is affected by cardiac surgery and in the clinical setting it is not possible to confirm a cerebral origin of the measured products. However, NO release from coronary vessels does not increase during or after cardioplegic arrest . NOx is a reliable measure of NO metabolism in vivo but interpretation of elevated plasma NOx concentrations requires certain factors to be taken into account. The interpretation of plasma component concentrations during cardiac surgery is difficult because of the fluid compartmental shifts that occur and changes in renal function. Normalization of parameters to the haematocrit or Hb and indices of renal function overcome these problems. In this study volumes of fluid administered prior to CPB were similar. When plasma NOx concentrations were standardized to Hb, statistically significant differences remained between the groups. Therefore, fluid shifts during surgery are unlikely to account for the observed difference in NOx concentrations. In this study also plasma creatinine concentrations demonstrated no significant correlation with plasma NOx at 5 min after removal of aortic cross clamp.
Increased concentrations of plasma NOx were identified preoperatively in the deficit compared to non-deficit groups. The baseline value of NOx (22.6 (9.2) μmol L−1) in the non-deficit group is similar to control values reported in previous studies . Identification of a pre-existing cognitive deficit is associated with increased risk of a subsequent postoperative deficit . In this study there was a statistically significant difference in several preoperative cognitive test scores in the deficit compared to non-deficit groups. This is an interesting finding in that the possibility exists that preoperative plasma NOx could be used to predict cognitive outcome. There is conflicting evidence regarding increased plasma NOx concentrations and neurological disease . In patients with dementia, increased concentrations have been reported . In the current study, patients in the deficit group had a trend towards increased incidence of hypertension and were older, both factors that could predispose to cognitive deficit and potential compensatory upregulation of NO production. There is conflicting evidence, however, regarding increased plasma NOx concentrations in healthy elderly patients [25,26]. Older age is associated with cognitive deficit after both cardiac and non-cardiac surgery . Confounding variables that could result in increased plasma NOx concentrations include diet, renal function and drugs capable of NO release. All patients fasted for 12 h preoperatively thus diet was unlikely to account for observed differences. Patients with renal impairment were excluded from the study and the proportion of patients taking drugs capable of NO release preoperatively, were similar in the two groups (Table 1). No association appears to exist between NOx and preoperative  or intraoperative  GTN administration.
The most appropriate control group for comparison with patients undergoing CABG has not been defined. The ‘Statement of Consensus on Assessment of Neurobehavioural Outcomes after Cardiac surgery’  recommends that measurement error and practice effects are taken into account. Estimation of practice effects and measurement error should match for factors of socio-economic background, educational attainment and mood factors and probably most importantly preoperative test scores . Patient spouses have previously been used to make calculations of practice effects and measurement error . In our study anxiety and depression scores, estimated IQ and years of education were similar in the control and patient groups (Table 1). Although the control group had a different male/female ratio to the study groups, this was offset by using tests free rom sex bias. The ‘Statement of Consensus on Assessment of Neurobehavioural Outcomes After Cardiac surgery’ recommends that tests used should be free from sex bias . These tests were used in this study. Thus differences in male/female ratio were unlikely to have had influence on our results. A limitation to the use of patient spouses in the RC method is the need to obtain a suitably large group that will allow matching for preoperative test scores. By choosing not to use a surgical control group, we have not provided controls for the effects of surgery and anaesthesia. In the early postoperative assessment of cognitive dysfunction this has relevance but not at long-term assessment (plateau cognitive function) when direct effects of surgery and anaesthesia are unlikely to influence test scores. After 3 months, patients' cognitive function appears to plateau and assessment at this time may be more predictive of long-term outcome . In a recent study , however, cognitive deficit at hospital discharge was most predictive of long-term outcome.
We have avoided the potentially misleading analysis of cognitive dysfunction preoperatively, though this represents a study limitation. Cognitive dysfunction was defined in this study by the RC index. This requires a control group, who are administered the same cognitive tests for a specified time interval. This information was not available to make a preoperative diagnosis of cognitive dysfunction. It will be necessary to examine this role of NOx as a potential marker for cognitive dysfunction in such patients. A further study limitation was that patients were operated on by two surgeons who used different preservation methods and techniques of aortic manipulation that may affect cognitive outcome.
In conclusion, we have demonstrated that in patients who develop a new cognitive deficit after CABG, compared to those who did not, (a) preoperative plasma concentrations of NOx are greater; (b) preoperative performance of some cognitive function tests (TMT A & B, Dig Symb) are poorer; (c) intra- and postoperative plasma concentrations of NOx are greater; (d) there was a time course similarity in NOx elevations for both groups. The implications of these findings are that (a) our data support the contention that a proportion of patients presenting for CABG surgery have a preoperative cognitive deficit, (b) preoperative estimation of NOx may serve as a predictor of subsequent new postoperative cognitive deficit and (c) perioperative plasma NOx concentrations do not serve as an effective biomarker in the assessment of intervention intended to decrease postoperative cognitive deficit.
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Keywords:© 2005 European Society of Anaesthesiology
CORONARY ARTERY BYPASS SURGERY; MENTAL PROCESSES; cognition; HYPOXIA-ISCHAEMIA; brain; BIOLOGICAL MARKERS; REACTIVE NITROGEN SPECIES; nitric oxide; NEUROPSYCHOLOGICAL TESTS