Stroke Biomarkers Research Articles

Inverse correlation between stroke and urinary 3-hydroxypropyl mercapturic acid, an acrolein-glutathione metabolite

BackgroundWe found previously that increases in plasma levels of protein-conjugated acrolein and polyamine oxidases, enzymes that produce acrolein, are good biomarkers for stroke. The aim of this study was to test whether 3-hydroxypropyl mercapturic acid (3-HPMA), an acrolein-glutathione metabolite, was increased in the urine of stroke patients. MethodsThe level of 3-HPMA in urine was measured by liquid chromatography with tandem mass spectrometry (LC-MS/MS). Stroke (78 subjects) was divided into 52 cerebral infarction (CI) and 26 cerebral hemorrhage (CH) on the basis of clinical information including brain imaging. ResultsA major acrolein derivative in urine is 3-HPMA. Being different from the results of PC-Acro in plasma, 3-HPMA in urine decreased following stroke. The median value of μmol 3-HPMA/g creatinine (Cre) for 90 control subjects was 2.83, while that for 78 stroke patients was 1.56. The degree of the decrease in 3-HPMA was similar in both CI and CH patients. Furthermore, the median value of μmol 3-HPMA/g Cre in 56 patients with lesions ≥1cm in diameter (1.39) was significantly lower than that in 20 patients with lesion <1cm in diameter (2.16). ConclusionInverse correlation between stroke and urinary 3-HPMA was observed. The results suggest that stroke is aggravated when nervous system tissues have a reduced level of glutathione.

Growth-differentiation factor-15 and functional outcome after acute ischemic stroke

Blood biomarkers may improve the performance in predicting early stroke outcome beyond well-established clinical factors. We investigated the value of growth-differentiation factor-15 (GDF-15) to predict functional outcome after 90 days in a prospectively collected patient cohort with symptoms of acute ischemic stroke. Two hundred eighty-one patients with symptoms of acute ischemic stroke were prospectively investigated. Serial blood samples for GDF-15 analysis were obtained after the admission of the patient, after 6 and 24 h. Primary outcome was the dichotomized modified ranking scale (MRS) 90 days after the initial clinical event. Within the final study population (264 patients, mean age 70.3 ± 12.7 years, 55.3% male), National Institutes of Health Stroke Scale (NIH-SS) [odds ratio (OR) 1.269, 95% confidence interval (CI) 1.141–1.412, p < 0.001] and initial GDF-15 levels (OR 1.029, 95% CI 1.007–1.053, p = 0.011) were independently associated with a MRS ≥ 2 after day 90 after multiple regression analysis. Growth-differentiation factor-15 levels increase with higher NIH-SS-tertiles (p = 0.005). Receiver-operator characteristic curves demonstrated a discriminatory accuracy to predict unfavourable stroke outcome of 0.629 (95% CI 0.558–0.699), 0.753 (95% CI 0.693–812) and 0.774 (95% CI 0.717–0.832) for GDF-15, NIH-SS and the combination of these variables. The additional use of GDF-15 to NIH-SS ameliorates the model with a net reclassification index of 0.044 (p = 0.541) and integrated discrimination improvement of 0.034 (p = 0.443). Growth-differentiation factor-15 as an acute stroke biomarker independently predicts unfavourable functional 90 day stroke outcome. Discriminatory value in addition to NIH-SS is only modestly distinct.

Detection of metals and metalloproteins in the plasma of stroke patients by mass spectrometry methods

Stroke is the leading cause of adult disability, worldwide. Metalloproteins and metals play key roles in epigenetic events in living organisms, including hypertension, the most important modifiable risk factor for stroke. Thus, metalloproteins may be important target biomarkers for disease diagnosis. The primary goal of this study was to assess metal containing proteins in blood plasma, detected by ICP-MS, followed by ESIMS for peptide/protein identification. We then compared the relative concentration differences between samples from patients with ischemic stroke, hemorrhagic stroke and stroke mimics. In 29 plasma samples (10 stroke mimics, 10 ischemic stroke and 9 hemorrhagic stroke patients) previously collected from patients who presented to the University of Cincinnati Emergency Department within 12 hours of symptom onset for a plasma banking project. For the metal associated protein study, Mg, Mn, Cu, Se concentrations were statistically different when compared between stroke mimics vs. ischemic stroke patients and ischemic stroke patients vs. hemorrhagic stroke patients. Pb concentrations were statistically different when compared between stroke mimics vs. ischemic stroke patients and Mo levels were statistically the same among the three groups. In addition, we also report concentration levels and preliminary correlation studies for total elemental analysis among the three sets of patients. This pilot study demonstrates that mass spectrometry methods may be highly valuable in detecting novel stroke biomarkers in blood plasma. Expanded studies are warranted to confirm these findings.

Short Communication Circulatory microRNA-145 expression is increased in cerebral ischemia

Cerebral ischemia or ischemic stroke is mainly attributed to vascular and circulation disorders. Among protein biomarkers, RNA profiles have also been identified as markers of ischemic stroke. MicroRNA-145 expression is ostensibly recognized as marker and modulator of vascular smooth muscle cell phenotype; however, expression levels in ischemic stroke had not been investigated. Employing real-time quantitative PCR, we examined the expression profile of circulatory microRNA-145 in healthy control subjects (N = 14) and ischemic stroke patients (N = 32). Circulatory microRNA-145 expression was significantly higher in ischemic stroke patients than in control subjects. This demonstrates that hemostatic mechanisms are affected by ischemic stroke. We conclude that circulating microRNA-145 has potential as a biomarker for ischemic stroke.

Increased Left Atrial Volume Index: Potent Biomarker for First-Ever Ischemic Stroke

Increased Left Atrial Volume Index: Potent Biomarker for First-Ever Ischemic Stroke

Gene Expression Profiling of Blood in Brain Arteriovenous Malformation Patients

Brain arteriovenous malformations (BAVMs) are an important cause of intracranial hemorrhage (ICH) in young adults. Gene expression profiling of blood has led to the identification of stroke biomarkers, and may help identify BAVM biomarkers and illuminate BAVM pathogenesis. It is unknown whether blood gene expression profiles differ between 1) BAVM patients and healthy controls, or 2) unruptured and ruptured BAVM patients at presentation. We characterized blood transcriptional profiles in 60 subjects (20 unruptured BAVM, 20 ruptured BAVM, and 20 healthy controls) using Affymetrix whole genome expression arrays. Expression differences between groups were tested by ANOVA, adjusting for potential confounders. Genes with absolute fold change ≥ 1.2 (false discovery rate corrected p ≤ 0.1) were selected as differentially expressed and evaluated for over-representation in KEGG biological pathways (p ≤ 0.05). Twenty-nine genes were differentially expressed between unruptured BAVM patients and controls, including 13 which may be predictive of BAVM. Patients with ruptured BAVM compared to unruptured BAVM differed in expression of 1490 genes, with over-representation of genes in 8 pathways including MAPK, VEGF, Wnt signaling and several inflammatory pathways. These results suggest clues to the pathogenesis of BAVM and/or BAVM rupture and point to potential biomarkers or new treatment targets.

Cell-Free DNA—A Marker to Predict Ischemic Brain Damage in a Rat Stroke Experimental Model

The animal model of stroke that is most frequently used is a rat model of focal brain ischemia caused by middle cerebral artery occlusion (MCAO). Several studies have reported a link between levels of cell-free DNA (CFD) and neurologic outcome in human stroke. The purpose of this study was to assess brain injury and measure CFD levels in 2 models of MCAO in rats, and to determine whether brain injury correlates with CFD. A total of 60 rats were used for this study. Twenty rats underwent a sham procedure, 20 rats had MCAO using a monofilament, and 20 rats had MCAO with a silicon-coated filament. Groups were further divided into 2 subgroups. In 1 subgroup of 10 rats, neurologic performance [measured as a neurologic severity score, (NSS)] was measured at 1 and 24 hours after the procedure, and brain edema and infarct volume were determined at 24 hours. In the second subgroup of 10 rats, CFD was measured at 0, 1, 2, 4, 8, 12, and 24 hours and at 2, 3, 4, and 5 days. Neurologic performance (measured as a NSS) was measured at 1 and 24 hours after the procedure. The main finding was a significant increase in CFD levels observed 24 hours after the onset of MCAO. The correlation between the total infarct volume and CFD levels of the 3 groups was R=0.78, P<0.0001. Brain edema and NSS also were strongly correlated with CFD levels at 24 hours after MCAO (R=0.91, P<0.0001 and R=0.73, P<0.0001, respectively). We found that CFD levels correlate well with the extent of ischemic injury, brain edema, and neurologic outcome in rats 24 hours post-MCAO. We have also shown that CFD correlates well with the expected temporal progression of ischemic injury. These findings place CFD in a unique place as a biomarker for stroke, both experimentally and possibly clinically.

Blood biomarkers of ischemic stroke.

This review provides a summary of the protein and RNA biomarkers that have been studied for the diagnosis and assessment of ischemic stroke. Many of the biomarkers identified relate to the pathophysiology of ischemic stroke, including ischemia of CNS tissue, acute thrombosis and inflammatory response. These biomarkers are summarized by their intended clinical application in ischemic stroke including diagnosis, prediction of stroke severity and outcome, and stratification of patients for stroke therapy. Among the biomarkers discussed are recent whole genome studies using RNA expression profiles to diagnose ischemic stroke and stroke etiology. Though many candidate blood based biomarkers for ischemic stroke have been identified, none are currently used in clinical practice. With further well designed study and careful validation, the development of blood biomarkers to improve the care of patients with ischemic stroke may be achieved.

N-Terminal Probrain Natriuretic Peptide as a Biomarker of Cardioembolic Stroke

and purpose N-terminal probrain natriuretic peptide, which is mainly produced by the heart, is increased in acute stroke. We aimed to determine if N-terminal probrain natriuretic peptide could be a biomarker for ischemic stroke with a cardioembolic cause. Consecutive sample of acute stroke patients admitted to a Stroke Unit. Ischemic stroke subtype was classified using the TOAST classification. Blood samples were drawn within 72 h after stroke onset. Serum N-terminal probrain natriuretic peptide concentration was measured using an electrochemiluminescence immunoassay. Mean values of N-terminal probrain natriuretic peptide were compared between patients with hemorrhagic stroke vs. ischemic stroke, cardioembolic stroke vs. noncardioembolic stroke, cardioembolic stroke with atrial fibrillation vs. noncardioembolic stroke using t-test. Receiver operating characteristic curves were used to test the ability of N-terminal probrain natriuretic peptide values to identify cardioembolic stroke and cardioembolic stroke with atrial fibrillation. Ninety-two patients were included (66 with ischemic stroke) with a mean age of 58·6 years. Twenty-eight (42·4%) ischemic strokes had a cardioembolic cause. Mean N-terminal probrain natriuretic peptide values for cardioembolic stroke were significantly higher (P<0·001) (491·6; 95% confidence interval 283·7-852·0 pg/ml) than for noncardioembolic ischemic stroke (124·7; 86·3-180·2 pg/ml). The area under the receiver operating characteristic curve for N-terminal probrain natriuretic peptide in cardioembolic stroke was 0·77. The cut-off point with the highest sensitivity and specificity was set at 265·5 pg/ml (71·4% and 73·7% respectively). The area under the curve of N-terminal probrain natriuretic peptide for cardioembolic stroke related to atrial fibrillation was 0·92, cut-off was set at 265·5 pg/ml (sensitivity 94·4%, specificity 72·9%). N-terminal probrain natriuretic peptide is a biomarker with a good accuracy to predict ischemic stroke of cardioembolic cause, namely associated with atrial fibrillation.

Plasma glial fibrillary acidic protein levels in children with sickle cell disease

To determine if glial fibrillary acidic protein (GFAP) is associated with brain injury in children with sickle cell disease (SCD), we measured plasma GFAP among cross-sectional groups of unselected children with SCD, subsets of children with SCD and normal brain MRI or MRI evidence of cerebral infarct, healthy pediatric controls, and adults with brain injury. Children with SCD had higher plasma GFAP than healthy pediatric controls (mean concentrations 0.14 ± 0.37 vs. 0.07 ± 0.08 ng/mL; P 5 0.003); also, 16.0% (16/100) of children with SCD and cerebral infarct had GFAP elevations above the 95th percentile of healthy pediatric controls (P 5 0.04). Although not statistically significant, children with SCD and cerebral infarct had more elevated GFAP levels than with SCD and no infarct (16/100, 16.0% vs. 14/168, 8.3%; P 5 0.07). Children with SCD and acute brain ischemia had a higher proportion of elevated GFAP than SCD children with normal MRI (3/6, 50% vs.8.3%; P 5 0.01). GFAP was associated with elevated systolic blood pressure in the preceding year and correlated positively with white blood cell count and negatively with age and performance IQ. Plasma GFAP is elevated among children with SCD and may be associated with subclinical brain injury.

Advance in research of biomarker in stroke

Advance in research of biomarker in stroke

Are Stroke Biomarkers Seeing Brain Vessels in Patients With Ischemic Stroke?

The significance of asymptomatic coronary artery atherosclerosis burden (CAB) in the interpretation of stroke biomarker results is unknown. We investigated biomarker values to determine if they are related to cerebral atherosclerosis with consideration of CAB. We prospectively recruited patients with noncardioembolic ischemic cerebrovascular disease who had no history of coronary artery disease. Patient-based vascular assessment was conducted using MRA for the cerebral arteries and cardiac CT angiography for the coronary arteries. Biomarkers including the C-reactive protein and homocysteine were measured. A total of 178 patients were included in the study. The extracranial carotid stenosis was associated with the highest quartile of C-reactive protein (OR, 2.24; 95% CI, 1.04-4.55), whereas age was the only predictor of the highest quartile of homocysteine (OR, 1.06; 95% CI, 1.02-1.11). However, with consideration of CAB, CAB (≥2 segments with ≥50% stenosis) remained the only independent predictor for the highest quartile of C-reactive protein (OR, 8.69; 95% CI, 2.41-31.30) and homocysteine (OR, 11.44; 95% CI, 2.61-50.11). Our data emphasize the importance of CAB in biomarker studies of patients with ischemic stroke.

Antithrombin III associated with fibrinogen predicts the risk of cerebral ischemic stroke

Background and purpose The purpose of this study is to examine the feasibility of developing plasma predictive value biomarkers of cerebral ischemic stroke before imaging evidence is acquired. Methods Blood samples were obtained from 198 patients who attended our neurology department as emergencies – with symptoms of vertigo, numbness, limb weakness, etc. – within 4.5 h of symptom onset, and before imaging evidence was obtained and medical treatment. After the final diagnosis was made by MRI/DWI/MRA or CTA in the following 24–72 h, the above cases were divided into two groups: stroke group and non-stroke group according to the imaging results. The levels of baseline plasma antithrombin III (AT-III), thrombin–antithrombin III (TAT), fibrinogen, D-dimer and high-sensitivity C-reactive protein (hsCRP) in the two groups were assayed. Results The level of the baseline AT-III in the stroke group was 118.07 ± 26.22%, which was lower than that of the non-stroke group (283.83 ± 38.39%). The levels of TAT, fibrinogen, hsCRP were 7.24 ± 2.28 μg/L, 5.49 ± 0.98 g/L, and 2.17 ± 1.07 mg/L, respectively, which were higher than those of the non-stroke group (2.53 ± 1.23 μg/L, 3.35 ± 0.50 g/L, 1.82 ± 0.67 mg/L). All the P-values were less than 0.001. The D-dimer level was 322.57 ± 60.34 μg/L, which was slightly higher than that of the non-stroke group (305.76 ± 49.52 μg/L), but the P-value was 0.667. The sensitivities of AT-III, TAT, fibrinogen, D-dimer and hsCRP for predicting ischemic stroke tendency were 97.37%, 96.05%, 3.29%, 7.89%, but the specificity was 93.62%, 82.61%, 100% and 100%, respectively, and all the P-values were less than 0.001. High levels of D-dimer and hsCRP were mainly seen in the few cases with severe large-vessel infarction. Conclusions Clinical manifestations of acute focal neurological deficits were associated with plasma AT-III and fibrinogen. These tests might help the risk assessment of acute cerebral ischemic stroke and/or TIA with infarction tendency in the superacute stage before positive imaging evidence is obtained.

Biomarkers of Stroke

Biomarkers of Stroke

脳梗塞バイオマーカー探索研究REBIOS（Research for Biomarkers in Ischemic Stroke）における成果と今後 &amp;mdash;Biomarkerを用いたreverse translational researchを中心に&amp;mdash;

発症24時間以内の脳梗塞患者（ラクナ梗塞，アテローム血栓性脳梗塞，心原性脳塞栓症）180名と，年齢と性をマッチングした健常者172名を対照とし，100項目の血中タンパク質マーカーの経時的測定と新規マーカー探索を実施した．脳梗塞各病型に特異的あるいは共通する多くのマーカーが見出され，脳梗塞の病態解明に有用であることが期待される．さらにサル脳梗塞モデルを用いて，臨床データとの類似点や相違点を明確にすることができた．臨床バイオマーカーの探索結果をもとに，動物実験や細胞実験といった基礎研究でその機能を検証するreverse translational researchは，脳梗塞の病態解明に有用であるとともに，新たな脳梗塞治療戦略の構築に有意義であると考えられた．

The S100B Protein Could Be Used as Adjuvant Diagnostic Tool in Acute Ischemic Stroke

Background: In the emergency department, the diagnosis of ischemic stroke is difficult because the diagnostic modalities are limited to non-contrast brain CT and neurologic examination. Serum S100B protein, a bio-marker for ischemic stroke, is needed as an additional diagnostic aid in acute ischemic stroke. Methods: We retrospectively reviewed 50 patients diagnosed with ischemic stroke between August 2007 and December 2008 by brain MRI after brain CT and serum S100B measurement in the emergency department. The serum levels of S100B protein were analyzed and the diagnostic sensitivity of non-contrast brain CT combined with abnormal elevation of S100B protein was compared with that of non-contrast brain CT alone. Results: The overall sensitivity of non-contrast brain CT in the diagnosis of ischemia was 54%. S100B protein in early ischemia had a sensitivity of 58%. However, combining non-contrast brain CT and S100B increased the sensitivity to 74%. Conclusions: A biomarker-based diagnostic test would not replace the necessity for CT or other early imaging studies, and before contemplating any reperfusion strategy, neuro-imaging must be performed to rule out intracranial hemorrhage. However, S100B protein, a serum bio-marker, is able to help emergency physicians evaluate patients with suspected ischemic stroke and decide on treatment.

The usefulness of albumin-adjusted ischemia-modified albumin index as early detecting marker for ischemic stroke

This study was conducted to investigate whether albumin-adjusted ischemia-modified albumin index (IMA index) is more sensitive and accurate than ischemia-modified albumin (IMA) as early detection marker of ischemic stroke, and to compare IMA and IMA index in progression and non-progression of ischemic stroke. This case-control study was done at an emergency medical center of a university hospital. 52 patients with neurological symptoms were enrolled (28 Ischemic Stroke Group and 24 Non-Stroke Group). In the ROC analysis of IMA index to diagnose stroke, area under the curve (AUC) was 0.990 (cutoff value 91.4; 95% CI: 0.970-1.000; sensitivity: 96.4%; specificity 95.8%). The AUC for IMA value was 0.928 (cutoff value 98 U/ml; 95% CI 0.857-0.999; sensitivity 89.3%; specificity 88.5%). [corrected] The difference between progression (n = 12) and non-progression group (n = 16) in IMA and IMA index were statistically insignificant (p > 0.01). IMA index was more sensitive than conventional IMA value as diagnostic biomarker of stroke, however, arguable as a predictive biomarker for progression of ischemic stroke.

NR2 antibodies: Risk assessment of transient ischemic attack (TIA)/stroke in patients with history of isolated and multiple cerebrovascular events

Background and purpose Predicting stroke using biomarkers would enable clinicians to help prevent stroke or mitigate damage. Several stroke biomarkers have been investigated but none has shown near term predictive value. Methods We studied patients presenting with a history of stroke or transient ischemic attack (TIA) to determine whether serum levels of autoantibodies to the NMDA receptor NR2 peptide (NR2Ab) reflected the presence of recent stroke compared with controls. Antibody levels were also correlated with clinical risk factors for stroke, including diabetes, hypertension, hyperlipidemia, and history of recent TIA or stroke. Results Of the 245 patients that presented with acute stroke or TIA, 130 consented to participate and results are available for the 120. Volunteers from the community were recruited as controls. Males and females with multiple recent strokes and females with acute strokes had elevated NR2Ab levels compared to non-stroke patients or controls. Using a multiple regression model, the predictive value for NR2Ab was compared to clinical risk factors. In men, the presence of stroke correlated with hypertension ( p < 0.001) and NR2Ab levels ( p < 0.01) and in women the presence of stroke correlated with hypertension ( p < 0.001), diabetes ( p < 0.05), atrial fibrillation ( p < 0.05) and NR2Ab ( p < 0.01). Conclusion These results suggest that NR2Ab levels reflect a history of multiple strokes and may serve as a predictive factor for stroke.

Genomic biomarkers and cellular pathways of ischemic stroke by RNA gene expression profiling

The objective of this study was to provide insight into the molecular mechanisms of acute ischemic cerebrovascular syndrome (AICS) through gene expression profiling and pathway analysis. Peripheral whole blood samples were collected from 39 MRI-diagnosed patients with AICS and 25 nonstroke control subjects ≥ 18 years of age. Total RNA was extracted from whole blood stabilized in Paxgene RNA tubes, amplified, and hybridized to Illumina HumanRef-8v2 bead chips. Gene expression was compared in a univariate manner between stroke patients and control subjects using t test in GeneSpring. The significant genes were tested in a logistic regression model controlling for age, hypertension, and dyslipidemia. Inflation of type 1 error was corrected by Bonferroni and Ingenuity Systems Pathway analysis was performed. Validation was performed by QRT-PCR using Taqman gene expression assays. A 9-gene profile was identified in the whole blood of ischemic stroke patients using gene expression profiling. Five of these 9 genes were identified in a previously published expression profiling study of stroke and are therefore likely biomarkers of stroke. Pathway analysis revealed toll-like receptor signaling as a highly significant canonical pathway present in the peripheral whole blood of patients with AICS. Our study highlights the relevance of the innate immune system through toll-like receptor signaling as a mediator of response to ischemic stroke and supports the claim that gene expression profiling can be used to identify biomarkers of ischemic stroke. Further studies are needed to validate and refine these biomarkers for their diagnostic potential.

Gene Expression Profiling of Blood for the Prediction of Ischemic Stroke

A blood-based biomarker of acute ischemic stroke would be of significant value in clinical practice. This study aimed to (1) replicate in a larger cohort our previous study using gene expression profiling to predict ischemic stroke; and (2) refine prediction of ischemic stroke by including control groups relevant to ischemic stroke. Patients with ischemic stroke (n=70, 199 samples) were compared with control subjects who were healthy (n=38), had vascular risk factors (n=52), and who had myocardial infarction (n=17). Whole blood was drawn ≤3 hours, 5 hours, and 24 hours after stroke onset and from control subjects. RNA was processed on whole genome microarrays. Genes differentially expressed in ischemic stroke were identified and analyzed for predictive ability to discriminate stroke from control subjects. The 29 probe sets previously reported predicted a new set of ischemic strokes with 93.5% sensitivity and 89.5% specificity. Sixty- and 46-probe sets differentiated control groups from 3-hour and 24-hour ischemic stroke samples, respectively. A 97-probe set correctly classified 86% of ischemic strokes (3 hour+24 hour), 84% of healthy subjects, 96% of vascular risk factor subjects, and 75% with myocardial infarction. This study replicated our previously reported gene expression profile in a larger cohort and identified additional genes that discriminate ischemic stroke from relevant control groups. This multigene approach shows potential for a point-of-care test in acute ischemic stroke.