Stroke Biomarkers Research Articles

Study of Morphofunctional Parameters of Blood Cells in Ischemic Stroke Using a Genome-wide Research

Biomarkers can play many useful roles in modern neurology. Early diagnosis and immediate therapy are important factors for reducing the degree of brain tissue damage in ischemic stroke, reduces the risk of death from stroke. In the current study, apolipoprotein CIII (ApoCIII), a biomarker of ischemic stroke, was found.

Proteomic analysis of extracellular vesicles enriched serum associated with future ischemic stroke

Identifying new biomarkers beyond the established risk factors that make it possible to predict and prevent ischemic stroke has great significance. Extracellular vesicles are powerful cell‒cell messengers, containing disease-specific biomolecules, which makes them powerful diagnostic candidates. Therefore, this study aimed to identify proteins derived from extracellular vesicles enriched serum related to future ischemic stroke events, using a proteomic method. Of Japanese subjects who voluntarily participated in health checkups at our institute a number of times, 10 subjects (6 males and 4 females, age: 64.2 ± 3.9 years) who developed symptomatic ischemic stroke (7.3 ± 4.4 years’ follow-up) and 10 age‒sex matched controls without brain lesions (6.7 ± 2.8 years’ follow-up) were investigated. Extracellular vesicles enriched fractions were derived from serum collected at the baseline visit. Differentially expressed proteins were evaluated using isobaric tagging for relative and absolute protein quantification (iTRAQ)-based proteomic analysis. Of the 29 proteins identified, alpha-2-macroglobulin, complement C1q subcomponent subunit B, complement C1r subcomponent, and histidine-rich glycoprotein were significantly upregulated (2.21-, 2.15-, 2.24-, and 2.16-fold, respectively) in subjects with future ischemic stroke, as compared with controls. Our study supports the concept of serum-derived extracellular vesicles enriched fractions as biomarkers for new-onset stroke. These proteins may be useful for prediction or for targeted therapy.

Muscle Synergies and Clinical Outcome Measures Describe Different Factors of Upper Limb Motor Function in Stroke Survivors Undergoing Rehabilitation in a Virtual Reality Environment.

Recent studies have investigated muscle synergies as biomarkers for stroke, but it remains controversial if muscle synergies and clinical observation convey the same information on motor impairment. We aim to identify whether muscle synergies and clinical scales convey the same information or not. Post-stroke patients were administered an upper limb treatment. Before (T0) and after (T1) treatment, we assessed motor performance with clinical scales and motor output with EMG-derived muscle synergies. We implemented an exploratory factor analysis (EFA) and a confirmatory factor analysis (CFA) to identify the underlying relationships among all variables, at T0 and T1, and a general linear regression model to infer any relationships between the similarity between the affected and unaffected synergies (Median-sp) and clinical outcomes at T0. Clinical variables improved with rehabilitation whereas muscle-synergy parameters did not show any significant change. EFA and CFA showed that clinical variables and muscle-synergy parameters (except Median-sp) were grouped into different factors. Regression model showed that Median-sp could be well predicted by clinical scales. The information underlying clinical scales and muscle synergies are therefore different. However, clinical scales well predicted the similarity between the affected and unaffected synergies. Our results may have implications on personalizing rehabilitation protocols.

Protective Properties of the Extract of Chrysanthemum on Patients with Ischemic Stroke.

Investigation of the protective effect of chrysanthemum extract in ischemic strokes patients is among the challenging issues with the traditional hospital system in general and smart technology-based hospitals in particular. In this study, we have evaluated the protective effect of chrysanthemum extract on patients with ischemic stroke by detecting the severity of stroke, neuronal indexes, and oxidative stress biomarkers. For this purpose, forty-six patients with ischemic stroke were randomly divided into the control group (n = 30) and chrysanthemum group (n = 30). The control group received standard stroke treatment, and the chrysanthemum group was treated with chrysanthemum extract 400 mg/day (200 mg/day, twice/day) on the basis of standard treatment. The groups were compared the effect of saffron capsules using the National Institute of Health Stoke Scale (NIHSS), serum neuron specific enolase (NSE), S100, brain-derived neurotrophic factor (BDNF), malondialdehyde (MDA), Su-peroxide dismutase (SOD), and total antioxidant capacity (TAC ) levels, at the time of first day and fourth day after treatment. On the first day after treatment, there was no significant difference in the NIHSS score, serum NSE, S100, BDNF, MDA, SOD, and TAC levels between the chrysanthemum group and the control group (P > 0.05). On the fourth day after treatment, the NIHSS, serum NSE, S100, and MDA levels were significantly reduced in the chrysanthemum group compared to the control group, while the BDNF, SOD, and TAC levels were higher (P < 0.05). In addition, compared to the levels on the first day, the NIHSS, serum NSE, S100, and MDA levels were significantly reduced, and the BDNF, SOD, and TAC levels were increased in the chrysanthemum group on the fourth day (P < 0.05). Chrysanthemum extract has the effects of scavenging oxygen free radicals and antioxidation and has a neuroprotective effect on ischemic stroke patients.

"Investigating Reliable Recovery Biomarkers of Ischemic Stroke: An Evidence-Based Study"

Deshwal PR1, Saikumar V2 and Tiwari P3* Author Affiliations 1Prity Rani Deshwal, Ph.D. Scholar, Department of Pharmacy Practice, National Institute of Pharmaceutical Education and Research (NIPER), Punjab, India 2Vasanthapuram Saikumar, M. Pharm. Scholar, Department of Pharmacy Practice, National Institute of Pharmaceutical Education and Research (NIPER), Punjab, India 3Pramil Tiwari, Prof. & Head, Department of Pharmacy Practice, National Institute of Pharmaceutical Education and Research (NIPER), Punjab, India Received: November 05, 2021 | Published: November 16, 2021 Corresponding author: Pramil Tiwari, Prof & Head, Department of Pharmacy Practice, National Institute of Pharmaceutical Education and Research (NIPER), S.A.S. Nagar, Punjab, India DOI: 10.26717/BJSTR.2021.40.006388

Abstract 13230: Bone Morphogenetic Protein 10 - A Novel Biomarker of Ischemic Stroke in Patients With Atrial Fibrillation

Background: Biomarkers specifically related to the atrial tissue might increase the understanding of the pathophysiology and improve risk prediction in atrial fibrillation (AF). Bone morphogenetic protein 10 (BMP10) is a novel biomarker expressed in atrial myocardium. We evaluated the association between BMP10 and the risk of stroke, bleeding, and death in patients with AF not treated with oral anticoagulation. Methods: BMP10 was measured in 2,974 plasma samples collected at randomisation in patients with AF randomized to aspirin in two cohorts, the ACTIVE-A (n=951) and AVERROES (n=2023) trials, median follow-up of 3.8 and 1.2 years, respectively. BMP10 was analysed with a prototype Elecsys electrochemiluminescence immunoassay. Associations between BMP10 and outcomes were evaluated by Cox-regression analyses adjusted for clinical characteristics, kidney function, and NT-proBNP. Results: Median age was 71 years, 58.1% were men. The median concentration of BMP10 was 2.46 (25 th -75 th percentiles, 2.09 - 2.94) ng/L. The variables most strongly associated with higher BMP10 levels were: heart rhythm (AF vs sinus), older age, female sex, lower BMI, and kidney dysfunction. BMP10 was the baseline variable most strongly associated with ischemic stroke during follow-up (Figure). After adjustments, the hazard ratio for ischemic stroke (BMP10 3rd vs 1st quartile) was 1.57 [95%CI 1.08-2.28], p=0.025), and the discriminatory value (c-index) was 0.643 (vs 0.634 of CHA 2 DS 2 -VASc score) and improved the full model from 0.737 to 0.746. BMP10 was not associated with bleeding or death. The results were consistent across both cohorts and irrespective of cardiac rhythm at blood sampling. Conclusions: In patients with AF, circulating plasma levels of BMP10 are independently associated with the risk of ischemic stroke but not with bleeding or death. BMP10 has an incremental discriminatory value for the risk of ischemic stroke, and may be a novel specific biomarker in patients with AF.

Predictive serum biomarkers of patients with cerebral infarction

ABSTRACT Objectives Stroke is the third most common cause of death and also causes seizures and disability. Biomarkers are abnormal signal indicators at the biological level that are present before the organism is seriously affected and are more sensitive to early diagnosis than are traditional imaging methods. Early diagnosis of stroke can prevent the progression of the disease. However, there are currently no widely accepted biomarkers for stroke that have been applied clinically. Methods A serum metabonomics method based on ultra-high-performance liquid chromatography-quadrupole-time of flight tandem mass spectrometry (UPLC-Q-TOF/MS) was used to identify potential biomarkers and metabolic pathways of cerebral infarction. The receiver-operating characteristic (ROC) curve was used to verify the diagnostic and classification abilities of the biomarkers, and a support vector machine (SVM) model was developed for the prediction of cerebral infarction. Results Principal component analysis revealed a clear separation between the normal and cerebral infarction groups. A total of 13 potential serum biomarkers were identified, which were mainly involved in linoleic acid metabolism; phenylalanine, tyrosine, and tryptophan biosynthesis; tyrosine metabolism; arachidonic acid metabolism; and fatty acid biosynthesis. The ROC curve analysis showed that the potential biomarkers had high specificity and sensitivity for the diagnosis of cerebral infarction. The SVM model had good diagnostic ability and could accurately distinguish the control group from the cerebral infarction group. Discussion The metabonomics approach may be a useful bioanalytical method for understanding the pathophysiology of cerebral infarction and may provide an experimental basis for the development of clinical biomarkers for stroke.

Neurofilament Light Chain Protein Is a Predictive Biomarker for Stroke After Surgical Repair for Acute Type A Aortic Dissection.

Background: Although great progress has been made in surgery and perioperative care, stroke is still a fatal complication of acute type A aortic dissection (ATAAD). Serum biomarkers may help assess brain damage and predict patient's prognosis.Methods: From March, 2019 to January, 2020, a total of 88 patients underwent surgical treatment at the Department of Cardiovascular Surgery of Beijing Anzhen Hospital, China, and were enrolled in this study. Patients were divided into two groups according to whether they had suffered a stroke after the operation. Blood samples were collected at 8 time points within 3 days after surgery to determine the level of S100β, neuron-specific enolase (NSE) and neurofilament light chain protein (NFL). Receiver operating characteristic curves (ROC) were established to explore the biomarker predictive value in stroke. The area under the curve (AUC) was used to quantify the ROC curve.Results: The patient average age was 48.1 ± 11.0 years old and 70 (79.6%) patients were male. Fifteen (17.0%) patients suffered stroke after surgery. The NFL levels of patients in the stroke group at 12 and 24 h after surgery were significantly higher than those in the non-stroke group (all P < 0.001). However, the NSE and S100β levels did not differ significantly at any time point between the two groups. The predictive value of NFL was the highest at 12 and 24 h after surgery, and the AUC was 0.834 (95% CI, 0.723–0.951, P < 0.001) and 0.748 (95% CI, 0.603–0.894, P = 0.004), respectively. Its sensitivity and specificity at 12 h were 86.7 and 71.6%, respectively. The NFL cutoff value for the diagnosis of stroke at 12 h after surgery was 16.042 ng/ml.Conclusions: This study suggests that NFL is an early and sensitive serum marker for predicting post-operative neurological prognosis of ATAAD patients. Further studies, including large-scale prospective clinical trials, are necessary to test whether the NFL can be used as a biomarker for clinical decision-making.

CircSKA3 acts as a sponge of miR-6796-5p to be associated with outcomes of ischemic stroke by regulating matrix metalloproteinase 9 expression.

This study was undertaken to screen the circular RNAs (circRNAs) influencing matrix metalloproteinase 9 (MMP9) through the competing endogenous RNA (ceRNA) network and evaluate the prognostic value of these circRNAs for acute ischemic stroke. A total of 220 ischemic stroke patients and 62healthy subjects were included in this study. RNA was isolated from blood collected in PAXgene tubes. Illumina sequencing, quantitative real-time polymerase chain reaction (qRT-PCR) validation, and luciferase reporter assay were explored to construct and verify the existence of a circRNA-microRNA (miRNA)-matrix metalloproteinase-9 (MMP9) network. The 215 ischemic stroke patients were recruited in a prognostic cohort. They were prospectively followed up for 3months after stroke onset, and a poor functional outcome was defined as a major disability or death. After Illumina sequencing, six circRNAs were predicted to bind miRNAs and then regulate MMP9messenger RNA (mRNA). qRT-PCR showed that only circSKA3 was significantly increased in ischemic stroke patients compared to healthy controls and positively associated with MMP9mRNA expression. Luciferase reporter assay further verified a direct interaction between circSKA3, MMP9, and hsa-miR-6796-5p. Patients in the top tertile of circSKA3had a 2.672-fold (p<0.05) risk of poor functional outcome, compared with those in the bottom tertile (p for trend = 0.016). The outcome was predicted by circSKA3 with area under the receiver operating characteristic curve at 0.614 (p=0.004). circSKA3 functioned as a ceRNA for hsa-miR-6796-5p to aggravate the progression of ischemic stroke via targeting MMP9. Baseline circSKA3 was positively associated with poor outcomes of ischemic stroke. circSKA3may be a potential biomarker or therapeutic target in ischemic stroke.

Sex-Specific MicroRNAs in Neurovascular Units in Ischemic Stroke.

Accumulating evidence pinpoints sex differences in stroke incidence, etiology and outcome. Therefore, more understanding of the sex-specific mechanisms that lead to ischemic stroke and aggravation of secondary damage after stroke is needed. Our current mechanistic understanding of cerebral ischemia states that endothelial quiescence in neurovascular units (NVUs) is a major physiological parameter affecting the cellular response to neuron, astrocyte and vascular smooth muscle cell (VSMC) injury. Although a hallmark of the response to injury in these cells is transcriptional activation, noncoding RNAs such as microRNAs exhibit cell-type and context dependent regulation of gene expression at the post-transcriptional level. This review assesses whether sex-specific microRNA expression (either derived from X-chromosome loci following incomplete X-chromosome inactivation or regulated by estrogen in their biogenesis) in these cells controls NVU quiescence, and as such, could differentiate stroke pathophysiology in women compared to men. Their adverse expression was found to decrease tight junction affinity in endothelial cells and activate VSMC proliferation, while their regulation of paracrine astrocyte signaling was shown to neutralize sex-specific apoptotic pathways in neurons. As such, these microRNAs have cell type-specific functions in astrocytes and vascular cells which act on one another, thereby affecting the cell viability of neurons. Furthermore, these microRNAs display actual and potential clinical implications as diagnostic and prognostic biomarkers in ischemic stroke and in predicting therapeutic response to antiplatelet therapy. In conclusion, this review improves the current mechanistic understanding of the molecular mechanisms leading to ischemic stroke in women and highlights the clinical promise of sex-specific microRNAs as novel diagnostic biomarkers for (silent) ischemic stroke.

Abstract 1122‐000183: Assessing Meningeal Lymphatic MRI Biomarkers in Acute Large Vessel Occlusion Stroke Treated with Thrombectomy

Introduction : Until recently, there was very little known about the brain’s waste removal system. Unlike the peripheral organ systems, the CNS lacked a clearly defined lymphatic vasculature and waste clearance mechanism. However, in 2015, the meningeal lymphatic system (MLS) was first visualized in the dura of humans (1). Since then, a number of studies have investigated the MLS’s potential impact on various neurological diseases, including Alzheimer’s, Parkinson’s, Multiple Sclerosis, hydrocephalus, and epilepsy (2‐6). Other studies have also demonstrated the MLS’s profound influence on stroke (7‐10). However, there is little research that characterizes MLS imaging biomarkers in relation to stroke. Our study evaluates currently proposed MLS MRI biomarkers in large vessel occlusion stroke treated with thrombectomy. Methods : Participants: We recruited twenty patients who had an acute ischemic stroke treated with mechanical thrombectomy (MT) in a large intracranial artery (middle cerebral artery, internal carotid artery, basilar and/or vertebral). The mean age of our population was 69.9 ± 15.5 years. 12 (60%) of our patients had been treated with a combined approach stentriever + aspiration catheter, 4 (20%) with stentriever only, and 4 (20%) with contact aspiration only. Study Design: Basic demographic data, clinical comorbidities, information related to the stroke, technical details about the MT and outcome were collected from electronic medical records. We implemented High Resolution MR Vessel Wall Imaging (HR‐VWI) with a black blood T1‐weighted sequence to characterize changes in the enhancing vessels around the superior sagittal sinus that have been described as potential markers for meningeal lymphatic vessels (1). Within 24 hours after the MT procedure, patients underwent a 45‐minute 3T‐ brain MRI with contrast using a 20‐channel head coil (MAGNETOM Skyra, Siemens) with a sagittal isotropic black blood sequence included with the standard protocol. Our field of view included the superior sagittal sinus. Coronal reconstructed pre‐and post‐contrast back blood T1 sequences were qualitatively evaluated by a board certified Neuroradiologist with 8 years of academic experience evaluating the cerebrovasculature for the presence of purported meningeal lymphatic prominence or asymmetry. Results : There was no significant prominence or asymmetric difference seen in the purported meningeal lymphatics based on 3D high‐resolution T1 black blood MRI sequences in acute stroke large vessel occlusion post‐MT subjects. Conclusions : While in our study no observable differences were seen in the purported meningeal lymphatics based on current MR techniques, it marks an important initial step in development of a useful biomarker for the CNS lymphatic system. Future studies are needed to evaluate alternative meningeal lymphatic quantitative biomarkers. Our study also highlights the need for improved standardization of biomarkers for the meningeal lymphatics. By continuing to evaluate imaging biomarkers, we can further understand the MLS’s potentially profound role in complex neurological diseases.

The lower expression of circulating miR-210 and elevated serum levels of HIF-1α in ischemic stroke; Possible markers for diagnosis and disease prediction.

Background Stroke, either due to ischemia or hemorrhage, causes acute neurological damages to the brain. There is shortage of reliable biomarkers for ischemic stroke (IS), and we therefore investigated the serum concentrations of microRNA‐210 (miR‐210) and hypoxia inducible factor‐1α (HIF‐1α), as possible diagnostic and/or prognostic markers for IS.MethodsSerum samples were acquired from 52 IS patients and their healthy counterparts at five time points: upon admission, 24 and 48 h after admission, upon discharge and 3 months later. Serum levels of miR‐210 and HIF‐1α were respectively analyzed using real time RT‐PCR and ELISA. Diagnostic and prognostic accuracy tests were performed to assess the value of suggested biomarkers.ResultsIS patients demonstrated higher levels of serum HIF‐1α and lower miR‐210 in comparison to the healthy subjects. MiR‐210 was suggested to be a weak diagnostic biomarker at the time of admission (AUC = 0.61; p = 0.05), while HIF‐1α was an acceptable diagnostic marker for IS (AUC = 0.73; p < 0.0001). The higher expression of miR‐210 and lower levels of HIF‐1α were associated with better survivals in IS patients.ConclusionsSerum miR‐210 is a weak diagnostic marker of IS. Serum HIF‐1α is a better biomarker in diagnosing IS patients but further work in larger groups, including those with hemorrhagic stroke is necessary to confirm its diagnostic utility. Similarly, the prognostic potentiality of miR‐210 and HIF‐1α was acceptable but needs bigger sample size and longer follow‐up to be statistically confirmed.

Hemorrhagic Stroke Induces a Time-Dependent Upregulation of miR-150-5p and miR-181b-5p in the Bloodstream.

To date, the only effective pharmacological treatment for ischemic stroke is limited to the clinical use of recombinant tissue plasminogen activator (rtPA), although endovascular therapy has also emerged as an effective treatment for acute ischemic stroke. Unfortunately, the benefit of this treatment is limited to a 4.5-h time window. Most importantly, the use of rtPA is contraindicated in the case of hemorrhagic stroke. Therefore, the identification of a reliable biomarker to distinguish hemorrhagic from ischemic stroke could provide several advantages, including an earlier diagnosis, a better treatment, and a faster decision on ruling out hemorrhage so that tPA may be administered earlier. microRNAs (miRNAs) are stable non-coding RNAs crucially involved in the downregulation of gene expression via mRNA cleavage or translational repression. In the present paper, taking advantage of three preclinical animal models of stroke, we compared the miRNA blood levels of animals subjected to permanent or transient middle cerebral artery occlusion (MCAO) or to collagenase-induced hemorrhagic stroke. Preliminarily, we examined the rat miRNome in the brain tissue of ischemic and sham-operated rats; then, we selected those miRNAs whose expression was significantly modulated after stroke to create a list of miRNAs potentially involved in stroke damage. These selected miRNAs were then evaluated at different time intervals in the blood of rats subjected to permanent or transient focal ischemia or to hemorrhagic stroke. We found that four miRNAs—miR-16-5p, miR-101a-3p, miR-218-5p, and miR-27b-3p—were significantly upregulated in the plasma of rats 3 h after permanent MCAO, whereas four other different miRNAs—miR-150-5p, let-7b-5p, let-7c-5p, and miR-181b-5p—were selectively upregulated by collagenase-induced hemorrhagic stroke. Collectively, our study identified some selective miRNAs expressed in the plasma of hemorrhagic rats and pointed out the importance of a precise time point measurement to render more reliable the use of miRNAs as stroke biomarkers.

Relationship of Alberta Stroke Program Early CT Score (ASPECTS) with the outcome of ischemic stroke and the neurocognitive stroke biomarkers

BackgroundReliable and acceptable biomarkers are needed to anticipate the outcome and cognitive impairment following ischemic stroke. The goal of this research is to examine the association of ASPECTS with cognitive decline, biomarkers of stroke, and acute ischemic stroke outcomes. This study included 120 patients with ischemic stroke in the middle cerebral artery region. The initial NIHSS, non-contrast CT brain assessed by ASPECTS, and the biomarkers of cognitive decline such as ESR, CRP, S100B, MMP9, and glutamate were investigated. The Montreal Cognitive Assessment and modified Rankin scale (mRS) were evaluated after 3 months. Correlations between ASPECTS, MoCA, biomarkers of cognitive impairment, and mRS were done by Spearman correlation.ResultsThe incidence of cognitive impairment in our patients was 25.8%. Stroke biomarkers (ESR, CRP, S100B, MMP9, and glutamate) were significantly increased in cognitively disabled individuals with significantly lower mean MoCA scores than in cognitively intact patients. There was a strong direct correlation linking the initial ASPECTS and total MoCA test score after 3 months follow-up. Cases with unfavorable outcomes were older, more incidence of hypertension, and had higher average initial NIHSS (P < 0.05). While the average ASPECTS scores for the favorable outcome group of patients were significantly higher and there was a significant negative correlation between the initial ASPECTS and modified Rankin Scale score.ConclusionsASPECTS is a reliable scale to identify the extent of acute ischemic injury and could participate in assessing the outcome. ASPECTS and particular neurocognitive stroke biomarkers will enable the early detection of post-stroke cognitive impairment.Trial registration Registration of Clinical Trial Research: ClinicalTrials.gov ID: NCT04235920

Copeptin: a potential blood biomarker for acute ischemic stroke

BackgroundCopeptin is a new blood biomarker for acute ischemic stroke which emerged to assist clinicians with decision-making. Serum copeptin can accurately reflect vasopressin concentration, which plays a role in aggravation of inflammatory responses, ions and neurotransmitters dysfunctions. The objective of this work was to investigate the relation between copeptin level as a blood biomarker and the short-term prognosis of acute ischemic stroke after 3 months. The current study included 45 patients with first ever acute ischemic stroke and 45 healthy volunteers as a control. Clinical evaluation, CT and MRI of the brain, NIHSS on admission, and mRS after 3 months were done for the patients, and all the patients and control were subjected to assessment of serum level of copeptin by ELISA technique.ResultsCopeptin level was significantly higher in patients with acute ischemic stroke compared to healthy control subjects (p-value = 0.001). Also, copeptin level was significantly higher in patients with severe stroke (NIHSS > 16) than in those with mild-to-moderate stroke (NIHSS 0–15) at presentation and in patients with unfavorable outcome (mRS 3–6) when compared to patients with favorable outcome (mRS 0–2) (p-value = 0.003 and 0.001, respectively).Copeptin level was significantly lower in patients who received thrombolytic therapy with rTPA (p-value = 0.049).ConclusionCopeptin has an interesting potential as a new prognostic biomarker for patients with acute ischemic stroke as its level was significantly higher in patients with severe stroke and in patients with unfavorable outcome.

Fingerprint of novel circulating microRNAs identify patients with stroke-embolic stroke of undetermined source

Abstract Background Stroke is the second-most common cause of death worldwide. Circulating levels of selected microRNAs (miRNAs) were found to be modulated both in animal experimental models and in patients with stroke, opening up new avenues for the identification of more effective and specific biomarkers to identify and risk-stratify stroke patients. Aim of the present study is to identify all circulating miRNAs that are modulated in patients with stroke, to select specific miRNAs to be used as disease biomarkers to improve prognosis. Methods 48 patients with stroke- ESUS were involved in the study. We have divided the patient groups based on patients who had a second stroke or TIA and did not have (safety vs safety control). Total RNA was extracted from plasma samples quality of extracted material was assessed using a fluorometric electrophoretic assay. MiRNA profiling was performed using the Affymetrix platform using. Statistical analysis was performed in TAC software. Additional analyses were performed in and R using Signal information obtained from the TAC output. We performed the following tests using log2 transformed data and all comparison groups (A-F). We performed additional FDR correction, logistic regression, Mann-whitney test t-test depending if variances were equal or differing. We calculated Area under the curve using ROCp R package. Scores were ranging from 0–1. Co-expression analysis to identify genes authentically expressed was performed using Spearman correlation (cutoff=0.9, Rpval=0.05). In order to identify the targets of DE miRNAs we used our wizbionet R package and previously developed pipelines [1,2]. We performed target screening using multimiR package, selecting top 20% predictions from all available databases. Results MiR-4786, miR-1205, miR-548ar-3p and miR-518e-3p were found the most differentially expressed miRNAs between the groups. So far, miR-4786 was studied only in patients with acute leukemia [3]. Several studies showed the importance of miR-1205 in cell carcinoma and ovarian cancer progression [4]. Moreover, so far only one study showed the regulation of miR-548ar-3p in breast cancer [5]. Finally only one study showed the alteration of miR-518e-3p in Parkinsons disease patients [6]. Besides, our enrichment analysis showed Interleukin-2 signaling pathway, Lipid and lipoprotein metabolism, BDNF signaling pathway, MAPK signaling pathway, Intellectual Disability, Alzheimer's Disease are significantly related to ESUS- patients. Conclusions Any of those miRNAs were never studied in stroke before, our results identified several novel circulating prognostic biomarkers miRNAs that are down- of up-regulated in ESUS-stroke patients (who had only one vs multiple stroke). Among those several miRNAs were identified that are known to play a role in the pathophysiology of neurovascular diseases, paving the way to a new class of smart pathophysiology-based biomarkers in stroke. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Polish National Science Center OPUS Figure 1Figure 2

Bioinformatics Identification of Ferroptosis-Related Biomarkers and Therapeutic Compounds in Ischemic Stroke.

Background: Stroke is one of the most common deadly diseases with an estimated 780,000 new cases globally, of which ischemic stroke accounts for over 80% of all cases. Ferroptosis is a new form of programmed cell death that plays a vital role in many diseases, including ischemic stroke and heart diseases. The role of the ferroptosis-related gene in the diagnosis, prognosis, or therapy of ischemic stroke was not fully clarified.Methods: Ferroptosis-related differentially expressed genes (DEGs) in ischemic stroke were identified by bioinformatic analysis of the GSE16561 and GSE22255 datasets. Subsequently, receiver operator characteristic (ROC) monofactor analysis was performed to evaluate the diagnostic value of ferroptosis-related biomarkers in ischemic stroke.Results: A total of 10 ferroptosis-related DEGs were identified in ischemic stroke vs. normal control. GO and KEGG analysis revealed that these 10 ferroptosis-related DEGs were mainly enriched in response to oxidative stress, HIF-1 signaling pathway, ferroptosis, lipid, and atherosclerosis. Moreover, the random forest model suggested three ferroptosis-related biomarkers, namely, PTGS2, MAP1LC3B, and TLR4, for ischemic stroke. Interestingly, the expression of PTGS2, MAP1LC3B, and TLR4 was upregulated in ischemic stroke. ROC monofactor analysis demonstrated a good performance of MAP1LC3B, PTGS2, and TLR4 in the diagnosis of ischemic stroke. The expression and diagnostic value of MAP1LC3B, PTGS2, and TLR4 in ischemic stroke were also verified using GSE22255. We also revealed the transcription factor regulation network and co-expressed protein network of ferroptosis-related biomarkers. Several potential therapeutic compounds corresponding to MAP1LC3B, PTGS2, and TLR4 were also identified for ischemic stroke, including Zinc12503187 (Conivaptan), Zinc3932831 (Avodart), Zinc64033452 (Lumacaftor), Zinc11679756 (Eltrombopag), Zinc100378061 (Naldemedine), and Zinc3978005 (Dihydroergotamine).Conclusion: Our results suggested MAP1LC3B, PTGS2, and TLR4 as potential diagnostic biomarkers for ischemic stroke, providing more evidence about the vital role of ferroptosis in ischemic stroke.

The importance of matrix metalloproteinases in the prognosis of acute ischemic stroke patients

Ischemic stroke is one of the leading causes of mortality and disability worldwide. Numerous studies were performed to assess the risk of clinical deterioration of acute ischemic stroke patients, including the risk of haemorrhagic transformation. The complexity of cerebral ischemia pathology raised the possibility of a multitude of candidate-molecules to be studied as stroke biomarkers. The blood brain barrier integrity biomarkers have shown promising results both in fundamental and clinical studies. Matrix metalloproteinases have been extensively analysed and gave encouraging results for predicting unfavourable neurological outcome, including the risk for haemorrhagic transformation. Matrix metalloproteinase-9 plays a crucial role in the disruption of the blood-brain barrier following focal cerebral ischemic stroke. Elevated matrix metalloproteinase-2 levels are responsible for the degradation of tight junction proteins, basal lamina and neuronal injury after ischemia, and may contribute to infarction and hemorrhagic volume. The review provides an overview of matrix metalloproteinases’ role in the prognosis of acute ischemic stroke patients, regarding the stroke outcome and the risk of haemorrhagic transformation. Conclusions: Matrix metalloproteinases, especially gelatinases, are extensively studied for their predictive value in ischemic stroke evolution. Matrix metalloproteinase-2 and matrix metalloproteinase-9 correlate with stroke severity and haemorrhagic transformation in acute ischemic stroke, but large validation studies are needed for practical translation. Future studies should focus on developing a biomarker panel for predicting outcomes in stroke patients.

Neuron-specific enolase as a biomarker of ischemic stroke

Early detection and evaluation of ischemic stroke (IS) can have a positive impact on its outcomes. One of the most promising ways is to use neuron-specific enolase (NSE). The goal of this study was to clarify the serum levels of NSE in the acute period of the IS and define their relationships with severity of neurologic deficit and short-term outcomes.

CCL2 (C-C Motif Chemokine Ligand 2) Biomarker Responses in Central Versus Peripheral Compartments After Focal Cerebral Ischemia.

Inflammatory mediators in blood have been proposed as potential biomarkers in stroke. However, a direct relationship between these circulating factors and brain-specific ischemic injury remains to be fully defined. An unbiased screen in a nonhuman primate model of stroke was used to find out the most responsive circulating biomarker flowing ischemic stroke. Then this phenomenon was checked in human beings and mice. Finally, we observed the temporospatial responsive characteristics of this biomarker after ischemic brain injury in mice to evaluate the direct relationship between this circulating factor and central nervous system–specific ischemic injury. In a nonhuman primate model, an unbiased screen revealed CCL2 (C-C motif chemokine ligand 2) as a major response factor in plasma after stroke. In mouse models of focal cerebral ischemia, plasma levels of CCL2 showed a transient response, that is, rapidly elevated by 2 to 3 hours postischemia but then renormalized back to baseline levels by 24 hours. However, a different CCL2 temporal profile was observed in whole brain homogenate, cerebrospinal fluid, and isolated brain microvessels, with a progressive increase over 24 hours, demonstrating a mismatch between brain versus plasma responses. In contrast to the lack of correlation with central nervous system responses, 2 peripheral compartments showed transient profiles that matched circulating plasma signatures. CCL2 protein in lymph nodes and adipose tissue was significantly increased at 2 hours and renormalized by 24 hours. These findings may provide a cautionary tale for biomarker pursuits in plasma. Besides a direct central nervous system response, peripheral organs may also contribute to blood signatures in complex and indirect ways.