Peptides 58 (2014) 42–46

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Prognostic value of neuropeptide proenkephalin A in patients with severe traumatic brain injury Jian-Bo Gao a,∗ , Wei-Dong Tang b , Xiao Wang a , Jia Shen c a b c

Department of Emergency Medicine, Fuyang People’s Hospital, 2-4 Guihua Road, Fuyang 311400, Zhejiang Province, China Department of Critical Care Medicine, Fuyang People’s Hospital, 2-4 Guihua Road, Fuyang 311400, Zhejiang Province, China Department of Neurosurgery, Fuyang People’s Hospital, 2-4 Guihua Road, Fuyang 311400, Zhejiang Province, China

a r t i c l e

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Article history: Received 4 May 2014 Received in revised form 6 June 2014 Accepted 6 June 2014 Available online 14 June 2014 Keywords: Proenkephalin A Traumatic brain injury Clinical outcome

a b s t r a c t High plasma proenkephalin A levels have been associated with poor clinical outcome of aneurysmal subarachnoid hemorrhage. This prospective observatory study was designed to investigate the relationship between plasma proenkephalin A levels and 1-week mortality, 6-month mortality and 6-month unfavorable outcome (defined as Glasgow Outcome Scale score of 1–3) in patients with severe traumatic brain injury. This study recruited 128 patients and 128 sex- and age-matched healthy controls. Plasma proenkephalin A levels, as measured by chemoluminescence sandwich immunoassay, were statistically significantly higher in patients than in healthy controls (239.1 ± 93.0 pmol/L vs.81.3 ± 22.1 pmol/L; P < 0.001) and were correlated with Glasgow Coma Scale scores (r = −0.540, P < 0.001). It was identified as an independent prognostic predictor of 1-week mortality [odds ratio (OR), 1.214; 95% confidence interval (CI), 1.103–1.425; P < 0.001], 6-month mortality (OR, 1.162; 95% CI, 1.101–1.372; P < 0.001) and 6-month unfavorable outcome (OR, 1.116; 95% CI, 1.097–1.281; P < 0.001). Moreover, it had high predictive value for 1-week mortality [area under curve (AUC), 0.852; 95% CI, 0.778–0.908], 6-month mortality (AUC, 0.841; 95% CI, 0.766–0.899) and 6-month unfavorable outcome (AUC, 0.830; 95% CI, 0.754–0.891). Furthermore, its predictive value was similar to Glasgow Coma Scale score’s (all P > 0.05). Yet, a combined logistic-regression model did not show that it statistically significantly improved the predictive value of Glasgow Coma Scale score (all P > 0.05). Thus, it was proposed that enhanced plasma proenkephalin A could be a useful, complementary tool to predict short- or long-term clinical outcome after severe traumatic brain injury. © 2014 Elsevier Inc. All rights reserved.

Introduction Severe traumatic brain injury (TBI) is a significant health concern and a major burden for society [23,27]. Severe TBI has a high mortality rate in the early period and survivors present many major, in-hospital complications [29]. Surviving patients after severe TBI suffer also from a lower life expectancy than the general population [1]. Outcome prediction is relevant for both clinical practice and research of severe TBI [21]. Despite the close association of Glasgow Coma Scale (GCS) score with outcome, prognostic predictions are sometimes difficult to make [28]. The determination of circulation biomarkers for outcome prediction in patients with severe TBI therefore becomes important instrument in both clinical practice and research [24].

∗ Corresponding author. Tel.: +86 0571 63101890. E-mail address: [email protected] (J.-B. Gao). http://dx.doi.org/10.1016/j.peptides.2014.06.006 0196-9781/© 2014 Elsevier Inc. All rights reserved.

The enkephalins, active as neurotransmitters, are involved in nociception and immune stimulation and also have been implicated in the pathophysiology of spinal cord trauma [31] and brain injury [10,19]. The stable precursor fragment of the neuropeptide enkephalin (proenkephalin A [PENK-A]) is secreted in an equimolar ratio to enkephalin and PENK-A concentrations mirror that of enkephalin [11]. The stability and longer ex vivo half-life of PENK-A is a practical advantage, which makes it easier to determine in the clinical laboratory [11]. Recently, PENK-A is known to have prognostic value in neurological diseases. For example, plasma PENK-A levels were highly associated with severity of cerebral injury, and may have prognostic value for fatal and nonfatal events in ischemic stroke [7]. Furthermore, plasma PENK-A levels independently predicted mortality and functional outcome at 6 months after aneurysmal subarachnoid hemorrhage [5]. Thus, we furthermore investigated the ability of PENK-A to predict short- and long-term clinical outcome in patients with severe TBI.

J.-B. Gao et al. / Peptides 58 (2014) 42–46

Materials and methods Patient and control populations A prospective observatory study over period of 3 years from March 2010 to March 2013 at the Fuyang People’s Hospital, Zhejiang Province, China was conducted. This study included the patients with isolated head trauma and postresuscitation GCS score of eight or less, and excluded the patients with less than 18 years of age, admission time >6 h, previous head trauma, neurological disease including ischemic or hemorrhagic stroke, use of antiplatelet or anticoagulant medication, diabetes mellitus, hypertension or presence of other prior systemic diseases including uremia, liver cirrhosis, malignancy, and chronic heart or lung disease. Healthy age- and sex-matched volunteers were recruited as control group. Written consent to participate in the study was obtained from study population or their relatives. This protocol was approved by the Ethics Committee of the Fuyang People’s Hospital before implementation. Clinical and radiological assessment On arrival at the Emergency Department, blood pressure, blood oxygen saturation, blood glucose and papillary reactivity were obtained. Arterial pressure was measured noninvasively using a conventional blood pressure sphygmomanometer. Mean arterial pressure was calculated from the diastolic and systolic values (mean arterial pressure = diastolic arterial pressure + 1/3[systolic arterial pressure − diastolic arterial pressure]). Head trauma severity was assessed using initial postresuscitation GCS score. All computerized tomography (CT) scans were performed according to the neuroradiology department protocol. Investigators who read them were blinded to clinical information. Abnormal cisterns, midline shift >5 mm and subarachnoid hemorrhage were recorded on initial CT scan. CT classification was performed using Traumatic Coma Data Bank criteria on initial postresuscitation CT scan according to the method of Marshall et al. [22]. End point Participants were followed up until death or completion of 6 months after head trauma. Clinical outcome included 1-week mortality, 6-month mortality and 6-month functional outcome. The functional outcome was defined by Glasgow outcome scale (GOS) score. GOS was defined as follows: 1 = death; 2 = persistent vegetative state; 3 = severe disability; 4 = moderate disability; and 5 = good recovery [16]. GOS Scores were dichotomized in favorable and unfavorable outcomes (GOS of 4–5 vs. GOS of 1–3). For follow-up, we used structure telephone interviews performed by one doctor, blinded to clinical information and PENK-A levels. Immunoassay methods Venous blood was drawn from patients on admission and from healthy controls at study entry. Plasma was frozen at −70 ◦ C until assayed. PENK-A 119–159 was determined by a chemoluminescence sandwich immunoassay as described previously (Immunochemical Intelligence GmbH, Berlin, Germany) [11]. All assessments were run in duplicates. The persons carrying out the assays were completely blinded to the clinical information. Statistical analysis Statistical analysis was performed with SPSS 19.0 (SPSS Inc., Chicago, IL, USA) and MedCalc 9.6.4.0. (MedCalc Software, Mariakerke, Belgium). The results were reported as counts (percentage)

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for the categorical variables, mean ± standard deviation if normally distributed and median (interquartile range) if not normally distributed for the continuous variables. Comparisons were made by using (1) chi-square test or Fisher exact test for categorical data, (2) student t test for continuous normally distributed variables, and (3) the Mann–Whitney U-test for continuous non-normally distributed variables. Spearman correlation coefficient was used to verify the correlation of plasma PENK-A levels with GCS scores. The relation of PENK-A to clinical outcome was assessed in a logistic-regression model with calculated odds ratio (OR) and 95% confidence interval (CI). For multivariate analysis, we included the significantly different outcome predictors as assessed in univariate analysis. The area under receiver operating characteristic curve (AUC) was calculated to assess the predictive performance of PENKA levels for clinical outcome. A combined logistic-regression model was configured to estimate the additive benefit of PENK-A to GCS scores. A P value of less than 0.05 was considered statistically significant. Results Patient characteristics During the study period, 158 patients were admitted to our Emergency Department with an isolated severe head trauma diagnosis. Of these, 30 patients were excluded according to exclusion criteria, and 128 patients were finally included in the analysis. This group of patients included 83 men and 45 women and had a mean age of 42.2 ± 16.4 years. Median initial postresuscitation GCS scores were 5 (3). 65 patients (50.8%) had unreactive pupils on admission; 60 patients (46.9%), CT classification 5 or 6; 63 patients (49.2%), abnormal cisterns on initial CT scan; 67 patients (52.3%), midline shift >5 mm on initial CT scan; 71 patients (55.5%), presence of traumatic subarachnoid hemorrhage on initial CT scan; 57 patients (44.5%), intracranial surgery in 1st 24 h; the mean admission time was 2.7 ± 1.4 h; the mean plasma-sampling time, 3.4 ± 1.4 h; the mean systolic arterial pressure, 122.4 ± 28.5 mmHg; the mean diastolic arterial pressure, 72.5 ± 19.2 mmHg; the mean value of mean arterial pressure, 89.1 ± 20.7 mmHg; the mean blood glucose level, 11.0 ± 3.9 mmol/L. Control group included 128 sexand age-matched healthy individuals. The plasma PENK-A levels were substantially higher in patients than in healthy controls (239.1 ± 93.0 pmol/L vs.81.3 ± 22.1 pmol/L; P < 0.001). Using Spearman correlation coefficient, plasma PENK-A levels were found to be correlated with GCS scores (r = −0.540, P < 0.001). 1-week mortality prediction Nineteen patients (14.8%) died from head trauma in 1 week. Higher plasma PENK-A levels were associated with 1-week mortality, as well as other variables shown in Table 1. When the above variables found to be significant in the univariate analysis were introduced into the logistic model, a multivariate analysis selected GCS scores (OR, 0.241; 95% CI, 0.139–0.617; P < 0.001) and plasma PENK-A levels (OR, 1.214; 95% CI, 1.103–1.425; P < 0.001) as the independent predictors for 1-week mortality of patients. A receiver operating characteristic curve identified that a baseline plasma PENK-A level >272.2 pmol/L predicted 1-week mortality of patients with 89.5% sensitivity and 73.4% specificity (Fig. 1). In Table 2, the predictive value of the PENK-A concentration was thus similar to that of GCS scores, but in a combined logisticregression model, PENK-A did not statistically significantly improve the AUC of GCS scores.

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Table 1 The factors associated with 1-week mortality. Characteristics

Non-survivors (n = 19)

Survivors (n = 109)

P value

Sex (male/female) Age (years) Initial postresuscitation GCS score Pupils unreactive on admission CT classification 5 or 6 Abnormal cisterns on initial CT scan Midline shift >5 mm on initial CT scan Presence of traumatic SAH on initial CT scan Intracranial surgery in 1st 24 h Admission time (h) Plasma-sampling time (h) Systolic arterial pressure (mmHg) Diastolic arterial pressure (mmHg) Mean arterial pressure (mmHg) Blood glucose level (mmol/L) Plasma proenkephalin A level (pmol/L)

12/7 41.5 ± 15.7 3(0) 18 (94.7%) 15 (79.0%) 18 (94.7%) 17 (89.5%) 16 (84.2%) 12 (63.2%) 2.2 ± 1.1 3.2 ± 0.7 126.1 ± 23.0 79.7 ± 17.4 95.1 ± 17.2 13.6 ± 5.0 337.2 ± 78.3

71/38 42.3 ± 16.6 6(3) 47(43.1%) 45(41.3%) 45(41.3%) 50(45.9%) 55(50.5%) 45(41.3%) 2.8 ± 1.5 3.4 ± 1.5 121.7 ± 29.4 71.2 ± 19.3 88.0 ± 21.2 10.6 ± 3.5 222.0 ± 84.6

0.868 0.837