- Email: [email protected]
Neutrophil Lymphocyte Ratio as a Predictor of Stroke
Neutrophil Lymphocyte Ratio as a Predictor of Stroke Serhat Tokgoz, MD,* Mehmet Kayrak, MD,† Zehra Akpinar, MD,* lu, MS,* Figen Gu € ney, MD,* and Betigu € l Yu € ru € ten, MD* Abdullah Seyithanog
Background: The aim of this study is to investigate the relationship of the neutrophil to lymphocyte ratio (NLR) with short-term mortality in acute stroke. Methods: This retrospective study included 255 patients with acute cerebral infarction who presented within 24 hours of symptom onset. A hemogram from peripheral venous blood samples was taken at the time of admission. The NLR was calculated as the ratio of neutrophils to lymphocytes. Duration of follow-up was defined as 60 days. Results: Seventy-one of 255 patients died during the follow-up period. The median NLR was significantly increased among the mortality group compared with the survival group (median 11.50, interquartile ratio [IQR] 10.40 vs median 3.79, IQR 4.72; P 5 .001). In our multivariate Cox regression model, NLR .5.0 (hazard ratio [HR] 3.30; 95% confidence interval [CI] 1.35-8.07), National Institutes of Health Stroke Scale score (HR 1.11; 95% CI 1.07-1.16), glucose values at admission (HR 1.007; 95% CI 1.002-1.011), and history of coronary artery disease (HR 2.49; 95% CI 1.26-4.92) were predictors of short-term mortality. The sensitivity for short-term mortality when the NLR was .5 was 83.10%, and the specificity was 62.00%. The positive predictive value of a NLR .5 was 45.7%, and negative predictive value was 90.50%. A strong linear association between NLR and National Institutes of Health Stroke Scale score was also observed (r 5 0.64; P 5 .001). In addition, the NLR was higher in both the atherosclerotic and cardioembolic stroke subgroups than the lacunar infarct subgroup (6.5 [IQR 7.2], 7.5 [IQR 8.9], and 3.20 [IQR 3.50], respectively; P 5 .001). Conclusions: The NLR at the time of hospital admission may be a predictor of short-term mortality in acute stroke patients. Because of the routine use and inexpensive nature of hemogram analysis, the NLR should be investigated in future prospective, randomized controlled trials investigating acute stroke. Key Words: Mortality—National Institutes of Health Stroke Scale— neutrophil lymphocyte ratio—stroke. Ó 2013 by National Stroke Association
Cerebrovascular ischemia has a high incidence of mortality and morbidity; many factors, such as acute phase reactants (C-reactive protein, etc.) and blood cell components
From the *Departments of Neurology; and †Cardiology, Necmettin Erbakan University, Meram Medical School, Konya, Turkey. Received December 5, 2012; revision received January 14, 2013; accepted January 20, 2013. Address correspondence to Serhat Tokgoz, MD, Department of Neurology, Necmettin Erbakan University, Meram Medical School, Konya 42081, Turkey. E-mail: [email protected]. 1052-3057/$ - see front matter Ó 2013 by National Stroke Association http://dx.doi.org/10.1016/j.jstrokecerebrovasdis.2013.01.011
(white blood cells [WBCs] and neutrophils, etc.) have been evaluated in acute ischemic events to predict infarct size, prognosis, and mortality.1,2 The pathophysiology of acute ischemic stroke (AIS) is an inflammatory process that involves endothelial activation, blood–brain barrier disruption, oxidant and inflammatory mediator accumulation, and the mass infiltration of leukocytes and platelets.3 This inflammatory process gradually develops within a few hours and plays an important role in ischemic damage.4,5 In particular, the release of inflammatory cytokines and neurotoxins plays an important role in exacerbating damage,6 but the prognostic impact of WBC subtype prevalence upon acute cerebral ischemia are unclear.7
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Among patients with acute myocardial infarction (MI), it has been shown that an increased neutrophil to lymphocyte ratio (NLR) is a predictor of in-hospital mortality and morbidity,8 and it has been shown that an increased NLR is related to impaired myocardial perfusion after percutaneous coronary angioplasty.9 The effect of NLR on AIS severity and mortality is not known. Because of the similarity of risk factors and prognostic markers of acute MI with AIS, we examined the relationship between the NLR at the time of hospital admission and short-term mortality in patients with AIS.
Methods Study Population This study is a hospital-based retrospective investigation. Three hundred ninety-four patients with AIS who were .18 years of age and who were admitted to Meram Medical School Hospital were screened between January 2007 and June 2012. Patients with a diagnosis of AIS on the first day of admission were included in the study. The study was approved by the local ethics committee. Exclusion criterion included patients who were admitted to the hospital .24 hours after AIS, patients with hematologic disorders, immunosuppressant drug users (steroids), those with an infection history within 2 weeks before the stroke, a stroke history within 6 months, and patients with a history of malignancy. Seventy patients with brainstem (infratentorial) infarct, which is known to have a direct effect on mortality, were also excluded. Two hundred ninety patients were included in the study protocol (Fig 1).
Study Protocol AIS cases were screened using the International Classification of Diseases, 10th revision code (G 46.8) from the hospital’s electronic record system. The demographic and clinical characteristics of 290 patients who met the criteria
were obtained from the patient’s archived records to evaluate Glasgow Coma Score (GCS), National Institutes of Health Stroke Scale (NIHSS) score, and mortality. Thirty-five patients were excluded from the study because of incomplete medical records (e.g., laboratory results and medical history). Supratentorial infarcts that were detected on cranial magnetic resonance imaging (MRI) and computed tomographic (CT) scans were divided into 2 groups: nonlacunar (.1.5 cm) infarcts and lacunar (#1.5 cm) infarcts. In addition, the nonlacunar infarct group was further separated into 2 groups: (1) cardioembolic stroke (according to the presence of the history of atrial fibrillation, valvular heart disease, and congestive heart failure) or (2) atherosclerotic stroke (based on the documented carotid artery disease and/or aortic calcific plaque without overt heart disease). Hospital mortalities of patients were determined according to the records of epicrisis. The hospital’s electronic medical record was searched for evaluation of nonhospital mortality within 60 days poststroke. Patients whose records continued after 60 days were considered alive. The families of patients with no records in the digital system after 60 days were contacted by telephone and were questioned about the mortality of the patient. Nonhospital mortality was found in 4 patients in total.
Blood Sample Analysis A hemogram was evaluated using the peripheral venous blood samples taken on admission to the emergency department. The blood sample was collected in a calcium ethylenediaminetetra-acetic acid tube. Since 2007, blood counts have been evaluated in our hospital with an autoanalyzer (Cell-Dyn Ruby Hematology Analyzer; Abbott Laboratories, Abbott Park, IL). NLR was calculated as the ratio of neutrophils to lymphocytes in peripheral blood. In addition, other routine laboratory findings were examined using the digital record systems of the hospital.
Statistical Analysis
Figure 1.
Study flow chart.
Data were analyzed using SPSS software (version 13.0; SPSS Inc, Chicago, IL) and presented as mean 6 standard deviation or median (interquartile range [IQR]). Distribution normality was analyzed with the Kolmogorov– Smirnov test. Correlation analysis was carried out using the Spearman correlation test for nonparametrically distributed variables. The difference between 2 groups was tested using independent Student t tests for normally distributed variables; the Mann–Whitney U test was used for the comparison of nonparametrically distributed variables. The difference between categorical variables was determined using the Chi-square test. Kaplan–Meier survival analysis was performed to investigate the relationship between the median NLR value and death. Cox regression analysis was performed to determine predictors of 60-day mortality. GCS score, NIHSS score, age, sex,
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hypertension, diabetes mellitus, coronary artery disease history, cerebrovascular event history, creatinin, glucose levels on admission, and NLR were included in this regression model as independent variables. The backward elimination method was applied and removal criteria for steps were accepted as $0.10. A MedCalc 9.2.0.1 (MedCalc Software, Mariakerke, Belgium) packet program was used to obtain the receiver operating curve (ROC) and to analyze the specificity, sensitivity, and negative and positive predictive values of NLR for death status. Power analysis was performed using a Minitab 16 (Minitab Inc., State College, PA) packet program. The sample volume was calculated as 250 patients to determine the difference in NLR with 90% power. The difference in NLR among stroke subtypes was analyzed using the Kruskal–Wallis test. The post hoc analysis was performed using the Mann–Whitney U test with Bonferroni correction.
Results The mean age of the 255 patients diagnosed with supratentorial AIS was 69.37 6 13.96 years. The mortality rate
was 27.8% (n 5 71) by 60 days of follow-up. Demographic and laboratory findings of the surviving and dead patients are reported in Table 1. The NLRs on admission to the hospital of patients who died were significantly higher than in the group of living patients. There were also significant differences in terms of GCS score, NIHSS score, age, serum glucose, etc. (Table 1). The study group had a median NLR of 5.0 (IQR 6.6). When analysis of 60-day survival of patients was examined according to median NLR values, the Kaplan–Meier survival curves were significantly different between the 2 groups (Fig 2). In Cox regression analysis, a NLR .5, NIHSS score, blood glucose, and coronary artery disease (CAD) history were independent predictors of the mortality (Table 2). The hazard ratio was 3.30 (1.35-8.07) when the NLR was .5 independent of other variables in the Cox regression model (P 5 .01). When the NLR is .5, the sensitivity was 83.1%, the specificity was 62.0%, the positive predictive value was 45.7% for short-term mortality, and the negative predictive value was 90.5%. The optimal cutoff value for NLR as a predictor for shortterm mortality was determined as 5.67 in the ROC curve.
Table 1. Demographic and laboratory findings of surviving and dead patients
Age, y (mean 6 SD) Sex (F/M) Hypertension (%) Diabetes mellitus (%) CAD (%) Smoking (%) Hyperlipidemia (%) GCS score, median (IQR) NIHSS score, median (IQR) Systolic BP, mm Hg (mean 6 SD) Diastolic BP, mm Hg (mean 6 SD) WBC (103/mL), median (IQR) Neutrophil (103/mL), median (IQR) Lymphocyte (103/mL), median (IQR) NLR, median (IQR) Glucose (mg/dL), median (IQR) Creatinine, mg/dL (mean 6 SD) Total cholesterol, mg/dL (mean 6 SD) Triglyceride (mg/dL), median (IQR) LDL, mg/dL (mean 6 SD) HDL, mg/dL (mean 6 SD) Drugs (%) Antihypertensive Antidiabetic ASA Coumadin Statin
Surviving (n 5 184)
Dead (n 5 71)
P value
67.5 6 14.5 89/95 51.6 28.6 19.2 27.7 36.1 15.0 (2) 4.0 (8) 141.1 6 33.2 83.8 6 16.0 8.9 (4.1) 6.03 (3.64) 1.50 (1.10) 3.80 (4.72) 113.0 (66.8) 0.94 6 .41 176.1 6 41.2 127.0 (76.9) 113.3 6 34.0 35.4 6 10.8
74.1 6 11.2 41/30 74.3 28.6 37.1 26.2 43.6 7.0 (4) 24.0 (10) 140.3 6 30.3 81.3 6 14.8 12.60 (5.10) 10.40 (5.10) 0.90 (0.70) 11.50 (10.40) 137.5 (50.5) 1.00 6 .44 162.6 6 37.5 88.50 (58.5) 97.8 6 33.8 40.7 6 11.3
.001 NS .001 NS .003 NS NS .001 .001 NS NS .001 .001 .001 .001 .001 NS .03 .001 .01 .01
42.7 32.8 28.4 8.8 26.5
55.2 30.6 32.8 12.8 39.9
NS NS NS NS .001
Z value
10.63* 8.84* 29.21 27.80 26.08 27.16 25.62 27.68 23.94 23.21
Abbreviations: ASA, acetylsalicylic acid; BP, blood pressure; CAD, coronary artery disease; F, female; GCS, Glasgow Coma Scale; HDL, high-density cholesterol; IQR, interquartile range; LDL, low-density cholesterol; M, male; NIHSS, National Institutes of Health Stroke Scale; NLR, neutrophil to lymphocyte ratio; NS, not significant; SD, standard deviation; WBC, white blood cell. *Chi-square.
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GCS score (r 5 20.433; P 5 .001; Table 3). NLR values were significantly higher in nonlacunar than in lacunar infarction (8.15 [IQR 7.8] v 3.20 [IQR 3.50]; z 5 25.43; P 5 .001, respectively). NLR values were significantly higher in both atherosclerotic and cardioembolic infarction groups than in the lacunar infarct group (Table 4), but not significantly different between the atherosclerotic and cardioembolic subtypes. In addition, NLR values were significantly higher among the deceased group than among the surviving subjects with both atherosclerotic and cardioembolic subtypes (Table 4).
Discussion
Figure 2. Kaplan–Meier survival estimates for neutrophil to lymphocyte ratio (NLR) in acute ischemic stroke (AIS) and a comparison of survival for NLR according to the median NLR value with long rank test.
On this level, the sensitivity was 81.7% and the specificity was 65.8% (positive predictive value 47.9%; negative predictive value 90.3%; Fig 3). When the linear relationships of NLR with other continuous variables were examined, the NIHSS score had a strong positive correlation with the NLR (r 5 0.64; P 5 .001) and a moderate negative correlation with the Table 2. Cox regression results for the predictors of mortality Variables First step GCS score NIHSS score Systolic BP Age Hypertension Diabetes mellitus CAD NLR* Creatinine Glucose Last step NIHSS CAD NLR (.5)* Glucose
HR (95% CI)
P value
0.92 (0.70-1.20) 1.08 (0.96-1.21) 0.99 (0.98-1.01) 0.99 (0.96-1.04) 1.48 (0.67-3.23) 0.90 (0.39-2.08) 2.76 (1.30-5.83) 3.31 (1.14-9.60) 0.89 (0.60-1.32) 1.006 (1.001-1.011)
.52 .21 .34 .89 .33 .81 .008 .03 .56 .02
1.11 (1.07-1.16) 2.49 (1.26-4.92) 3.30 (1.35-8.07) 1.007 (1.002-1.011)
.001 .01 .01 .03
Abbreviations: BP, blood pressure; CAD, coronary artery disease; CI, confidence interval; GCS, Glasgow Coma Scale; HR, hazard ratio; NIHSS, National Institutes of Health Stroke Scale; NLR, neutrophil to lymphocyte ratio. *NLR was included in the regression model as a dichotomous variable as above median (.5) and below or equal to the median (#5).
The pathogenesis of cerebral infarct is an inflammatory process that results in liquefaction necrosis. Cytokines and adhesion molecules regulate the increased migration of leukocytes 10-13 to the infarct area. It has been suggested that inflammatory cells exacerbate the damage by attacking the penumbra area in addition to the core area of infarction.14 Neutrophil migration into the damaged area is the first response to ischemic brain damage.1 Intraparenchymal perivascular neutrophil migration occurs within 6 to 24 hours.15,16 It is known that proteolytic enzymes, such as acid phosphatase and reactive oxygen species, may be released in the damaged area through the accumulation of neutrophils in ischemic and reperfused areas.17,18 It has been proposed that baseline neutrophil numbers may be related to tissue damage severity, reinfarct risk, and poor neurologic outcome.2,19-21 Haumer et al2 suggested that neutrophil numbers .5.8 103/mL result in an
Figure 3. Receiver operating curves of neutrophil lymphocyte ratio to prediction of mortality in acute ischemic stroke (AIS). Abbreviations: AUC, area under curve; CI, confidence interval; NPV, negative predictive value; PPV, positive predictive value.
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Table 3. A linear relationship between neutrophil to lymphocyte ratio and the other continuous variables Variable
r value
P value
GCS NIHSS Systolic BP Diastolic BP Age Creatinine Glucose
20.43 0.64 20.02 20.10 0.19 0.17 0.19
.001 .001 .82 .13 .002 .01 .003
Abbreviations: BP, blood pressure; GCS, Glasgow Coma Scale; NIHSS, National Institutes of Health Stroke Scale.
increased risk of major cardiovascular events. Significantly higher numbers of neutrophils in nonlacunar infarcts and deceased patients suggests that high neutrophil numbers are related to infarct size and mortality in our study. The ineffectiveness of antineutrophil treatment for AIS22 has led to new investigations. Reduced infarct size and improved clinical symptoms have been shown in studies on immunocompromised mice.12,23 Despite the contribution of the immune response to tissue damage, efforts to treat stroke with immunosuppression failed, resulting in a predisposition to infection, such as pneumonia.24,25 Similar to neutrophils, lymphocytes also play an important role in the inflammatory response. Lymphocytes begin to increase on the first day after stroke, peaking on day 7.26 Although there have been contradictory studies,13 it has been suggested that T-cell lymphocytes play an important role repairing inflamed tissues.27 This T cell–mediated repair is related to the release of cytokines and growth factors by T cells to modulate microglial activation.28 The significant negative correlation of lymphocyte values with mortality in our study suggests a relationship between severity of stroke and lymphopenia. Because of inadequate results of studies that monitored the number of neutrophils and lymphocytes separately, some studies regarding the
NLR, vascular damage, and clinical results were oriented in the final days of the follow-up period. The NLR has been investigated for its role in the cardiovascular system. In studies of coronary syndromes, it has been postulated that increased NLR is an independent predictor factor of 30-day mortality in patients with acute coronary syndrome.8,29,30 The NLR was significantly higher among the progressive group (5.0 6 5.1 v 3.2 6 3; P , .001). In multivariate analysis, the NLR was significantly related to progression (relative risk [RR] 2.27; 95% CI 1.07-4.82; P , .03).31 The NLR may be related to arteriosclerosis and coronary calcium score in the absence on infarction within the coronary arteries.32 To our knowledge, this is the first study to investigate the relationship of NLR to short-term mortality in AIS. The main finding of this study is that NLR is an independent predictor factor of short-term mortality in AIS. A statistically significant increase in mortality caused by stroke was found among patients with higher NLRs. In this study, a NLR .5.0 was determined to be a threshold value that predicts mortality in stroke. In addition, NIHSS score, glucose, and coronary artery disease (CAD) were also independent predictors of mortality. The strong correlation between the NLR, NIHSS score, and GCS score with mortality supports the importance of the clinical course. The lack of correlation between blood pressure and stroke mortality may be exposure of the patient to the emergency antihypertensive intervention at admission. An increased NLR may also predict the size of the infarct, independent of etiology.33 In light of these results, a question comes to mind: is NLR the result of physiologic stress only (i.e., steroid secretion [neutrophilia or lymphopenia]), or a factor that is concurrent and that helps increase the size of the infarction and therefore mortality. No benefit was seen with the use of immunosuppressant therapies that reduce the level of neutrophils and WBC subgroups. At this stage, we must concentrate on therapeutic approaches catalyzing the transition to the subacute lymphocytic inflammation
Table 4. A comparison of median neutrophil to lymphocyte ratio values among stroke subtypes and the relationship between neutrophil to lymphocyte ratio and survival status for each stroke subtype NLR Stroke subtype Atherosclerotic (n 5 103) Cardioembolic (n 5 65) Lacunar (n 5 87) P valuez Chi-square
Total, median (IQR) Surviving, n (median [IQR]) Dead, n (median [IQR]) P value* Z value 6.5 (7.2)y 7.5 (8.9)y 3.2 (3.5) .001 30.2
76 (5.5 [5.4]) 30 (5.2 [5.5]) — — —
27 (12.0 [11.0]) 35 (10.8 [8.9]) — — —
.001 .001 — — —
24.0 24.4 — — —
Abbreviations: IQR, interquartile range; NLR, neutrophil to lymphocyte ratio. Because death was not seen in the lacunar infarct group, P* and Z values are not shown. *Comparisons of median NLR of surviving versus dead patients for each stroke subtypes (Mann–Whitney U test). yAfter post hoc analysis, there was a statistically significant difference in the NLR between lacunar and other groups (P 5 .001). zDifference of median NLR among stroke subtypes (Kruskal–Wallis test).
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stage from the severe acute neutrophilic inflammatory phase in patients with higher NLRs.
Study Limitations This study has several limitations. A correlation between infarct volume and NLR could not be evaluated because no infarct volume measurement was made. However, it was determined that NLR evaluation is significantly lower for lacunar compared to nonlacunar infarcts. The absence of systemic infection was determined by medical history and body temperature. Infection parameters may be increased without elevated temperature, but a strong statistical relationship was found. In conclusion, NLR is a significant predictor of mortality, and an elevated NLR at the time of admission may be an important consideration for determining clinical treatment. Because of the routine use of hemogram, the easy calculation of the NLR, and the affordability of hemogram analysis, the value of NLR in AIS should be investigated in future prospective, randomized controlled trials.
References 1. Buck BH, Liebeskind DS, Saver JL, et al. Early neutrophilia is associated with volume of ischemic tissue in acute stroke. Stroke 2008;39:355-360. 2. Haumer M, Amighi J, Exner M, et al. Association of neutrophils and future cardiovascular events in patients with peripheral artery disease. J Vasc Surg 2005;41:610-617. 3. Ishikawa M, Zhang JH, Nanda A, et al. Inflammatory responses to ischemia and reperfusion in the cerebral microcirculation. Front Biosci 2004;9:1339-1347. 4. Bartus RT, Dean RL, Cavanaugh K, et al. Time-related neuronal changes following middle cerebral artery occlusion: Implications for therapeutic intervention and the role of calpain. J Cereb Blood Flow Metab 1995;15:969-979. 5. Freedman JE, Loscalzo J. Platelet-monocyte aggregates: Bridging thrombosis and inflammation. Circulation 2002; 105:2130-2132. 6. Chamorro A. Role of inflammation in stroke and atherothrombosis. Cerebrovasc Dis 2004;17:1-5. 7. Kim J, Song TJ, Park JH, et al. Different prognostic value of white blood cell subtypes in patients with acute cerebral infarction. Atherosclerosis 2012;222:464-467. 8. Tamhane UU, Aneja S, Montgomery D, et al. Association between admission neutrophil to lymphocyte ratio and outcomes in patients with acute coronary syndrome. Am J Cardiol 2008;102:653-657. 9. Akpek M, Kaya MG, Lam YY, et al. Relation of neutrophil/lymphocyte ratio to coronary flow to in-hospital major adverse cardiac events in patients with STelevated myocardial infarction undergoing primary coronary intervention. Am J Cardiol 2012;110:621-627. 10. Barone FC, Feuerstein GZ. Inflammatory mediators and stroke: New opportunities for novel therapeutics. J Cereb Blood Flow Metab 1999;19:819-834. 11. Yan J, Greer JM, Etherington K, et al. Immune activation in the peripheral blood of patients with acute ischemic stroke. J Neuroimmunol 2009;206:112-117. 12. Yilmaz G, Arumugam TV, Stokes KY, et al. Role of T lymphocytes and interferon-gamma in ischemic stroke. Circulation 2006;113:2105-2112.
S. TOKGOZ ET AL. 13. Yilmaz G, Granger DN. Leukocyte recruitment and ischemic brain injury. Neuromolecular Med 2010;12:193-204. 14. Huang J, Upadhyay UM, Tamargo RJ. Inflammation in stroke and focal cerebral ischemia. Surg Neurol 2006;66: 232-245. 15. Clark RK, Lee EV, White RF, et al. Reperfusion following focal stroke hastens inflammation and resolution of ischemic injured tissue. Brain Res Bull 1994;35:387-392. 16. Garcia JH, Liu KF, Yoshida Y, et al. Influx of leukocytes and platelets in an evolving brain infarct (Wistar rat). Am J Pathol 1994;144:188-199. 17. Frangogiannis NG, Smith CW, Entman ML. The inflammatory response in myocardial infarction. Cardiovasc Res 2002;53:31-47. 18. Chavan V, Patil N, Karnik ND. Study of leukocytic hydrolytic enzymes in patients with acute stage of coronary heart disease. Indian J Med Sci 2007;61:73-82. 19. Avanzas P, Quiles J, Lopez de Sa E, et al. Neutrophil count and infarct size in patients with acute myocardial infarction. Int J Cardiol 2004;97:155-156. 20. Akopov SE, Simonian NA, Grigorian GS. Dynamics of polymorphonuclear leukocyte accumulation in acute cerebral infarction and their correlation with brain tissue damage. Stroke 1996;27:1739-1743. 21. Fisher TC, Meiselmann HJ. Polymorphonuclear leukocytes in ischemic vascular disease. Thromb Res 1994;74:21-34. 22. Krams M, Lees KR, Hacke W, et al. Acute Stroke Therapy by Inhibition of Neutrophils (ASTIN): An adaptive doseresponse study of UK-279,276 in acute ischemic stroke. Stroke 2003;34:2543-2548. 23. Hurn PD, Subramanian S, Parker SM, et al. T- and B-celldeficient mice with experimental stroke have reduced lesion size and inflammation. J Cereb Blood Flow Metab 2007;27:1798-1805. 24. Enlimomab Acute Stroke Trial Investigators. Use of antiICAM-1 therapy in ischemic stroke: Results of the Enlimomab Acute Stroke Tril. Neurology 2001;57:1428-1434. 25. Dirnagl U, Klehmet J, Braun JS, et al. Stroke-induced immunodepression: Experimental evidence and clinical relevance. Stroke 2007;38:770-773. 26. Schroeter M, Jander S, Witte OW, et al. Local immune responses in the rat cerebral cortex after middle cerebral artery occlusion. J Neuroimmunol 1994;55:195-203. 27. Schwartz M, Moalem G. Beneficial immune activity after CNS injury: Prospects for vaccination. J Neuroimmunol 2001;113:185-192. 28. Palm NW, Medzhitov R. Not so fast: Adaptive suppression of innate immunity. Nat Med 2007;13:1142-1144. 29. Muhmmed Suliman MA, Bahnacy Juma AA, Ali Almadhani AA, et al. Predictive value of neutrophil to lymphocyte ratio in outcomes of patients with acute coronary syndrome. Arch Med Res 2010;41:618-622. 30. Duffy BK, Gurm HS, Rajagopal V, et al. Usefulness of an elevated neutrophil to lymphocyte ratio in predicting long-term mortality after percutaneous coronary intervention. Am J Cardiol 2006;97:993-996. 31. Kalay N, Dogdu O, Koc F, et al. Hematologic parameters and angiographic progression of coronary atherosclerosis. Angiology 2012;63:213-217. 32. Cho KH, Jeong MH, Ahmed K, et al. Value of early risk stratification using hemoglobin level and neutrophil-tolymphocyte ratio in patients with ST-elevation myocardial infarction undergoing primary percutaneous coronary intervention. Am J Cardiol 2011;107:849-856. 33. Ertas G, Sonmez O, Turfan M. Neutrophil/lymphocyte ratio is associated with thromboembolic stroke in patients with non-valvular atrial fibrillation. J Neurol Sci 2013;324:49-52.
Background: The aim of this study is to investigate the relationship of the neutrophil to lymphocyte ratio (NLR) with short-term mortality in acute stroke. Methods: This retrospective study included 255 patients with acute cerebral infarction who presented within 24 hours of symptom onset. A hemogram from peripheral venous blood samples was taken at the time of admission. The NLR was calculated as the ratio of neutrophils to lymphocytes. Duration of follow-up was defined as 60 days. Results: Seventy-one of 255 patients died during the follow-up period. The median NLR was significantly increased among the mortality group compared with the survival group (median 11.50, interquartile ratio [IQR] 10.40 vs median 3.79, IQR 4.72; P 5 .001). In our multivariate Cox regression model, NLR .5.0 (hazard ratio [HR] 3.30; 95% confidence interval [CI] 1.35-8.07), National Institutes of Health Stroke Scale score (HR 1.11; 95% CI 1.07-1.16), glucose values at admission (HR 1.007; 95% CI 1.002-1.011), and history of coronary artery disease (HR 2.49; 95% CI 1.26-4.92) were predictors of short-term mortality. The sensitivity for short-term mortality when the NLR was .5 was 83.10%, and the specificity was 62.00%. The positive predictive value of a NLR .5 was 45.7%, and negative predictive value was 90.50%. A strong linear association between NLR and National Institutes of Health Stroke Scale score was also observed (r 5 0.64; P 5 .001). In addition, the NLR was higher in both the atherosclerotic and cardioembolic stroke subgroups than the lacunar infarct subgroup (6.5 [IQR 7.2], 7.5 [IQR 8.9], and 3.20 [IQR 3.50], respectively; P 5 .001). Conclusions: The NLR at the time of hospital admission may be a predictor of short-term mortality in acute stroke patients. Because of the routine use and inexpensive nature of hemogram analysis, the NLR should be investigated in future prospective, randomized controlled trials investigating acute stroke. Key Words: Mortality—National Institutes of Health Stroke Scale— neutrophil lymphocyte ratio—stroke. Ó 2013 by National Stroke Association
Cerebrovascular ischemia has a high incidence of mortality and morbidity; many factors, such as acute phase reactants (C-reactive protein, etc.) and blood cell components
From the *Departments of Neurology; and †Cardiology, Necmettin Erbakan University, Meram Medical School, Konya, Turkey. Received December 5, 2012; revision received January 14, 2013; accepted January 20, 2013. Address correspondence to Serhat Tokgoz, MD, Department of Neurology, Necmettin Erbakan University, Meram Medical School, Konya 42081, Turkey. E-mail: [email protected]. 1052-3057/$ - see front matter Ó 2013 by National Stroke Association http://dx.doi.org/10.1016/j.jstrokecerebrovasdis.2013.01.011
(white blood cells [WBCs] and neutrophils, etc.) have been evaluated in acute ischemic events to predict infarct size, prognosis, and mortality.1,2 The pathophysiology of acute ischemic stroke (AIS) is an inflammatory process that involves endothelial activation, blood–brain barrier disruption, oxidant and inflammatory mediator accumulation, and the mass infiltration of leukocytes and platelets.3 This inflammatory process gradually develops within a few hours and plays an important role in ischemic damage.4,5 In particular, the release of inflammatory cytokines and neurotoxins plays an important role in exacerbating damage,6 but the prognostic impact of WBC subtype prevalence upon acute cerebral ischemia are unclear.7
Journal of Stroke and Cerebrovascular Diseases, Vol. 22, No. 7 (October), 2013: pp 1169-1174
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Among patients with acute myocardial infarction (MI), it has been shown that an increased neutrophil to lymphocyte ratio (NLR) is a predictor of in-hospital mortality and morbidity,8 and it has been shown that an increased NLR is related to impaired myocardial perfusion after percutaneous coronary angioplasty.9 The effect of NLR on AIS severity and mortality is not known. Because of the similarity of risk factors and prognostic markers of acute MI with AIS, we examined the relationship between the NLR at the time of hospital admission and short-term mortality in patients with AIS.
Methods Study Population This study is a hospital-based retrospective investigation. Three hundred ninety-four patients with AIS who were .18 years of age and who were admitted to Meram Medical School Hospital were screened between January 2007 and June 2012. Patients with a diagnosis of AIS on the first day of admission were included in the study. The study was approved by the local ethics committee. Exclusion criterion included patients who were admitted to the hospital .24 hours after AIS, patients with hematologic disorders, immunosuppressant drug users (steroids), those with an infection history within 2 weeks before the stroke, a stroke history within 6 months, and patients with a history of malignancy. Seventy patients with brainstem (infratentorial) infarct, which is known to have a direct effect on mortality, were also excluded. Two hundred ninety patients were included in the study protocol (Fig 1).
Study Protocol AIS cases were screened using the International Classification of Diseases, 10th revision code (G 46.8) from the hospital’s electronic record system. The demographic and clinical characteristics of 290 patients who met the criteria
were obtained from the patient’s archived records to evaluate Glasgow Coma Score (GCS), National Institutes of Health Stroke Scale (NIHSS) score, and mortality. Thirty-five patients were excluded from the study because of incomplete medical records (e.g., laboratory results and medical history). Supratentorial infarcts that were detected on cranial magnetic resonance imaging (MRI) and computed tomographic (CT) scans were divided into 2 groups: nonlacunar (.1.5 cm) infarcts and lacunar (#1.5 cm) infarcts. In addition, the nonlacunar infarct group was further separated into 2 groups: (1) cardioembolic stroke (according to the presence of the history of atrial fibrillation, valvular heart disease, and congestive heart failure) or (2) atherosclerotic stroke (based on the documented carotid artery disease and/or aortic calcific plaque without overt heart disease). Hospital mortalities of patients were determined according to the records of epicrisis. The hospital’s electronic medical record was searched for evaluation of nonhospital mortality within 60 days poststroke. Patients whose records continued after 60 days were considered alive. The families of patients with no records in the digital system after 60 days were contacted by telephone and were questioned about the mortality of the patient. Nonhospital mortality was found in 4 patients in total.
Blood Sample Analysis A hemogram was evaluated using the peripheral venous blood samples taken on admission to the emergency department. The blood sample was collected in a calcium ethylenediaminetetra-acetic acid tube. Since 2007, blood counts have been evaluated in our hospital with an autoanalyzer (Cell-Dyn Ruby Hematology Analyzer; Abbott Laboratories, Abbott Park, IL). NLR was calculated as the ratio of neutrophils to lymphocytes in peripheral blood. In addition, other routine laboratory findings were examined using the digital record systems of the hospital.
Statistical Analysis
Figure 1.
Study flow chart.
Data were analyzed using SPSS software (version 13.0; SPSS Inc, Chicago, IL) and presented as mean 6 standard deviation or median (interquartile range [IQR]). Distribution normality was analyzed with the Kolmogorov– Smirnov test. Correlation analysis was carried out using the Spearman correlation test for nonparametrically distributed variables. The difference between 2 groups was tested using independent Student t tests for normally distributed variables; the Mann–Whitney U test was used for the comparison of nonparametrically distributed variables. The difference between categorical variables was determined using the Chi-square test. Kaplan–Meier survival analysis was performed to investigate the relationship between the median NLR value and death. Cox regression analysis was performed to determine predictors of 60-day mortality. GCS score, NIHSS score, age, sex,
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hypertension, diabetes mellitus, coronary artery disease history, cerebrovascular event history, creatinin, glucose levels on admission, and NLR were included in this regression model as independent variables. The backward elimination method was applied and removal criteria for steps were accepted as $0.10. A MedCalc 9.2.0.1 (MedCalc Software, Mariakerke, Belgium) packet program was used to obtain the receiver operating curve (ROC) and to analyze the specificity, sensitivity, and negative and positive predictive values of NLR for death status. Power analysis was performed using a Minitab 16 (Minitab Inc., State College, PA) packet program. The sample volume was calculated as 250 patients to determine the difference in NLR with 90% power. The difference in NLR among stroke subtypes was analyzed using the Kruskal–Wallis test. The post hoc analysis was performed using the Mann–Whitney U test with Bonferroni correction.
Results The mean age of the 255 patients diagnosed with supratentorial AIS was 69.37 6 13.96 years. The mortality rate
was 27.8% (n 5 71) by 60 days of follow-up. Demographic and laboratory findings of the surviving and dead patients are reported in Table 1. The NLRs on admission to the hospital of patients who died were significantly higher than in the group of living patients. There were also significant differences in terms of GCS score, NIHSS score, age, serum glucose, etc. (Table 1). The study group had a median NLR of 5.0 (IQR 6.6). When analysis of 60-day survival of patients was examined according to median NLR values, the Kaplan–Meier survival curves were significantly different between the 2 groups (Fig 2). In Cox regression analysis, a NLR .5, NIHSS score, blood glucose, and coronary artery disease (CAD) history were independent predictors of the mortality (Table 2). The hazard ratio was 3.30 (1.35-8.07) when the NLR was .5 independent of other variables in the Cox regression model (P 5 .01). When the NLR is .5, the sensitivity was 83.1%, the specificity was 62.0%, the positive predictive value was 45.7% for short-term mortality, and the negative predictive value was 90.5%. The optimal cutoff value for NLR as a predictor for shortterm mortality was determined as 5.67 in the ROC curve.
Table 1. Demographic and laboratory findings of surviving and dead patients
Age, y (mean 6 SD) Sex (F/M) Hypertension (%) Diabetes mellitus (%) CAD (%) Smoking (%) Hyperlipidemia (%) GCS score, median (IQR) NIHSS score, median (IQR) Systolic BP, mm Hg (mean 6 SD) Diastolic BP, mm Hg (mean 6 SD) WBC (103/mL), median (IQR) Neutrophil (103/mL), median (IQR) Lymphocyte (103/mL), median (IQR) NLR, median (IQR) Glucose (mg/dL), median (IQR) Creatinine, mg/dL (mean 6 SD) Total cholesterol, mg/dL (mean 6 SD) Triglyceride (mg/dL), median (IQR) LDL, mg/dL (mean 6 SD) HDL, mg/dL (mean 6 SD) Drugs (%) Antihypertensive Antidiabetic ASA Coumadin Statin
Surviving (n 5 184)
Dead (n 5 71)
P value
67.5 6 14.5 89/95 51.6 28.6 19.2 27.7 36.1 15.0 (2) 4.0 (8) 141.1 6 33.2 83.8 6 16.0 8.9 (4.1) 6.03 (3.64) 1.50 (1.10) 3.80 (4.72) 113.0 (66.8) 0.94 6 .41 176.1 6 41.2 127.0 (76.9) 113.3 6 34.0 35.4 6 10.8
74.1 6 11.2 41/30 74.3 28.6 37.1 26.2 43.6 7.0 (4) 24.0 (10) 140.3 6 30.3 81.3 6 14.8 12.60 (5.10) 10.40 (5.10) 0.90 (0.70) 11.50 (10.40) 137.5 (50.5) 1.00 6 .44 162.6 6 37.5 88.50 (58.5) 97.8 6 33.8 40.7 6 11.3
.001 NS .001 NS .003 NS NS .001 .001 NS NS .001 .001 .001 .001 .001 NS .03 .001 .01 .01
42.7 32.8 28.4 8.8 26.5
55.2 30.6 32.8 12.8 39.9
NS NS NS NS .001
Z value
10.63* 8.84* 29.21 27.80 26.08 27.16 25.62 27.68 23.94 23.21
Abbreviations: ASA, acetylsalicylic acid; BP, blood pressure; CAD, coronary artery disease; F, female; GCS, Glasgow Coma Scale; HDL, high-density cholesterol; IQR, interquartile range; LDL, low-density cholesterol; M, male; NIHSS, National Institutes of Health Stroke Scale; NLR, neutrophil to lymphocyte ratio; NS, not significant; SD, standard deviation; WBC, white blood cell. *Chi-square.
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GCS score (r 5 20.433; P 5 .001; Table 3). NLR values were significantly higher in nonlacunar than in lacunar infarction (8.15 [IQR 7.8] v 3.20 [IQR 3.50]; z 5 25.43; P 5 .001, respectively). NLR values were significantly higher in both atherosclerotic and cardioembolic infarction groups than in the lacunar infarct group (Table 4), but not significantly different between the atherosclerotic and cardioembolic subtypes. In addition, NLR values were significantly higher among the deceased group than among the surviving subjects with both atherosclerotic and cardioembolic subtypes (Table 4).
Discussion
Figure 2. Kaplan–Meier survival estimates for neutrophil to lymphocyte ratio (NLR) in acute ischemic stroke (AIS) and a comparison of survival for NLR according to the median NLR value with long rank test.
On this level, the sensitivity was 81.7% and the specificity was 65.8% (positive predictive value 47.9%; negative predictive value 90.3%; Fig 3). When the linear relationships of NLR with other continuous variables were examined, the NIHSS score had a strong positive correlation with the NLR (r 5 0.64; P 5 .001) and a moderate negative correlation with the Table 2. Cox regression results for the predictors of mortality Variables First step GCS score NIHSS score Systolic BP Age Hypertension Diabetes mellitus CAD NLR* Creatinine Glucose Last step NIHSS CAD NLR (.5)* Glucose
HR (95% CI)
P value
0.92 (0.70-1.20) 1.08 (0.96-1.21) 0.99 (0.98-1.01) 0.99 (0.96-1.04) 1.48 (0.67-3.23) 0.90 (0.39-2.08) 2.76 (1.30-5.83) 3.31 (1.14-9.60) 0.89 (0.60-1.32) 1.006 (1.001-1.011)
.52 .21 .34 .89 .33 .81 .008 .03 .56 .02
1.11 (1.07-1.16) 2.49 (1.26-4.92) 3.30 (1.35-8.07) 1.007 (1.002-1.011)
.001 .01 .01 .03
Abbreviations: BP, blood pressure; CAD, coronary artery disease; CI, confidence interval; GCS, Glasgow Coma Scale; HR, hazard ratio; NIHSS, National Institutes of Health Stroke Scale; NLR, neutrophil to lymphocyte ratio. *NLR was included in the regression model as a dichotomous variable as above median (.5) and below or equal to the median (#5).
The pathogenesis of cerebral infarct is an inflammatory process that results in liquefaction necrosis. Cytokines and adhesion molecules regulate the increased migration of leukocytes 10-13 to the infarct area. It has been suggested that inflammatory cells exacerbate the damage by attacking the penumbra area in addition to the core area of infarction.14 Neutrophil migration into the damaged area is the first response to ischemic brain damage.1 Intraparenchymal perivascular neutrophil migration occurs within 6 to 24 hours.15,16 It is known that proteolytic enzymes, such as acid phosphatase and reactive oxygen species, may be released in the damaged area through the accumulation of neutrophils in ischemic and reperfused areas.17,18 It has been proposed that baseline neutrophil numbers may be related to tissue damage severity, reinfarct risk, and poor neurologic outcome.2,19-21 Haumer et al2 suggested that neutrophil numbers .5.8 103/mL result in an
Figure 3. Receiver operating curves of neutrophil lymphocyte ratio to prediction of mortality in acute ischemic stroke (AIS). Abbreviations: AUC, area under curve; CI, confidence interval; NPV, negative predictive value; PPV, positive predictive value.
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Table 3. A linear relationship between neutrophil to lymphocyte ratio and the other continuous variables Variable
r value
P value
GCS NIHSS Systolic BP Diastolic BP Age Creatinine Glucose
20.43 0.64 20.02 20.10 0.19 0.17 0.19
.001 .001 .82 .13 .002 .01 .003
Abbreviations: BP, blood pressure; GCS, Glasgow Coma Scale; NIHSS, National Institutes of Health Stroke Scale.
increased risk of major cardiovascular events. Significantly higher numbers of neutrophils in nonlacunar infarcts and deceased patients suggests that high neutrophil numbers are related to infarct size and mortality in our study. The ineffectiveness of antineutrophil treatment for AIS22 has led to new investigations. Reduced infarct size and improved clinical symptoms have been shown in studies on immunocompromised mice.12,23 Despite the contribution of the immune response to tissue damage, efforts to treat stroke with immunosuppression failed, resulting in a predisposition to infection, such as pneumonia.24,25 Similar to neutrophils, lymphocytes also play an important role in the inflammatory response. Lymphocytes begin to increase on the first day after stroke, peaking on day 7.26 Although there have been contradictory studies,13 it has been suggested that T-cell lymphocytes play an important role repairing inflamed tissues.27 This T cell–mediated repair is related to the release of cytokines and growth factors by T cells to modulate microglial activation.28 The significant negative correlation of lymphocyte values with mortality in our study suggests a relationship between severity of stroke and lymphopenia. Because of inadequate results of studies that monitored the number of neutrophils and lymphocytes separately, some studies regarding the
NLR, vascular damage, and clinical results were oriented in the final days of the follow-up period. The NLR has been investigated for its role in the cardiovascular system. In studies of coronary syndromes, it has been postulated that increased NLR is an independent predictor factor of 30-day mortality in patients with acute coronary syndrome.8,29,30 The NLR was significantly higher among the progressive group (5.0 6 5.1 v 3.2 6 3; P , .001). In multivariate analysis, the NLR was significantly related to progression (relative risk [RR] 2.27; 95% CI 1.07-4.82; P , .03).31 The NLR may be related to arteriosclerosis and coronary calcium score in the absence on infarction within the coronary arteries.32 To our knowledge, this is the first study to investigate the relationship of NLR to short-term mortality in AIS. The main finding of this study is that NLR is an independent predictor factor of short-term mortality in AIS. A statistically significant increase in mortality caused by stroke was found among patients with higher NLRs. In this study, a NLR .5.0 was determined to be a threshold value that predicts mortality in stroke. In addition, NIHSS score, glucose, and coronary artery disease (CAD) were also independent predictors of mortality. The strong correlation between the NLR, NIHSS score, and GCS score with mortality supports the importance of the clinical course. The lack of correlation between blood pressure and stroke mortality may be exposure of the patient to the emergency antihypertensive intervention at admission. An increased NLR may also predict the size of the infarct, independent of etiology.33 In light of these results, a question comes to mind: is NLR the result of physiologic stress only (i.e., steroid secretion [neutrophilia or lymphopenia]), or a factor that is concurrent and that helps increase the size of the infarction and therefore mortality. No benefit was seen with the use of immunosuppressant therapies that reduce the level of neutrophils and WBC subgroups. At this stage, we must concentrate on therapeutic approaches catalyzing the transition to the subacute lymphocytic inflammation
Table 4. A comparison of median neutrophil to lymphocyte ratio values among stroke subtypes and the relationship between neutrophil to lymphocyte ratio and survival status for each stroke subtype NLR Stroke subtype Atherosclerotic (n 5 103) Cardioembolic (n 5 65) Lacunar (n 5 87) P valuez Chi-square
Total, median (IQR) Surviving, n (median [IQR]) Dead, n (median [IQR]) P value* Z value 6.5 (7.2)y 7.5 (8.9)y 3.2 (3.5) .001 30.2
76 (5.5 [5.4]) 30 (5.2 [5.5]) — — —
27 (12.0 [11.0]) 35 (10.8 [8.9]) — — —
.001 .001 — — —
24.0 24.4 — — —
Abbreviations: IQR, interquartile range; NLR, neutrophil to lymphocyte ratio. Because death was not seen in the lacunar infarct group, P* and Z values are not shown. *Comparisons of median NLR of surviving versus dead patients for each stroke subtypes (Mann–Whitney U test). yAfter post hoc analysis, there was a statistically significant difference in the NLR between lacunar and other groups (P 5 .001). zDifference of median NLR among stroke subtypes (Kruskal–Wallis test).
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stage from the severe acute neutrophilic inflammatory phase in patients with higher NLRs.
Study Limitations This study has several limitations. A correlation between infarct volume and NLR could not be evaluated because no infarct volume measurement was made. However, it was determined that NLR evaluation is significantly lower for lacunar compared to nonlacunar infarcts. The absence of systemic infection was determined by medical history and body temperature. Infection parameters may be increased without elevated temperature, but a strong statistical relationship was found. In conclusion, NLR is a significant predictor of mortality, and an elevated NLR at the time of admission may be an important consideration for determining clinical treatment. Because of the routine use of hemogram, the easy calculation of the NLR, and the affordability of hemogram analysis, the value of NLR in AIS should be investigated in future prospective, randomized controlled trials.
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