In-thrombus thrombin secretion: a new diagnostic marker of atrial fibrillation in cryptogenic stroke.

Endovascularly retrieved clots may be a potential resource for diagnosing stroke etiology. This method may influence secondary prevention treatment. We measure thrombin activity eluted by serially washing clots. We concluded that an assay measuring the change in thrombin in clots retrieved during acute stroke endovascular thrombectomy procedures may serve as a diagnostic marker of the origin of the clot. The suggested mechanism for these differences may be the clot location before its retrieval, with high blood flow causing thrombin washout in atherosclerotic clots, in contrast to atrium appendage low blood flow retaining high thrombin levels.

Clinical uncertainty in large vessel occlusion ischemic stroke: does automated perfusion imaging make a difference? An intra-rater and inter-rater agreement study.

BACKGROUND: Limited research exists regarding the impact of neuroimaging on endovascular thrombectomy (EVT) decisions for late-window cases of large vessel occlusion (LVO) stroke. OBJECTIVE: T0 assess whether perfusion CT imaging: (1) alters the proportion of recommendations for EVT, and (2) enhances the reliability of EVT decision-making compared with non-contrast CT and CT angiography. METHODS: We conducted a survey using 30 patients drawn from an institutional database of 3144 acute stroke cases. These were presented to 29 Canadian physicians with and without perfusion imaging. We used non-overlapping 95% confidence intervals and difference in agreement classification as criteria to suggest a difference between the Gwet AC1 statistics (κG). RESULTS: The percentage of EVT recommendations differed by 1.1% with or without perfusion imaging. Individual decisions changed in 21.4% of cases (11.3% against EVT and 10.1% in favor). Inter-rater agreement (κG) among the 29 raters was similar between non-perfusion and perfusion CT neuroimaging (κG=0.487; 95% CI 0.327 to 0.647 and κG=0.552; 95% CI 0.430 to 0.675). The 95% CIs overlapped with moderate agreement in both. Intra-rater agreement exhibited overlapping 95% CIs for all 28 raters. κG was either substantial or excellent (0.81-1) for 71.4% (20/28) of raters in both groups. CONCLUSIONS: Despite the minimal difference in overall EVT recommendations with either neuroimaging protocol one in five decisions changed with perfusion imaging. Regarding agreement we found that the use of automated CT perfusion images does not significantly impact the reliability of EVT decisions for patients with late-window LVO.

Janus Kinase-2 V617F Mutation and Antiphospholipid Syndrome in Cerebral Sinus Venous Thrombosis: Natural History and Retrospective Bicenter Analysis.

Background: Cerebral sinus venous thrombosis (CSVT) is a rare neurovascular entity, usually associated with acquired or genetic hypercoagulable states. In up to 30% of the cases it remains idiopathic. Bone marrow proliferation disorders that are associated with Janus Kinase 2 V617F mutation (JAK-2) are known causes of the systemic and cerebral thrombosis-at times despite normal blood counts-for which hematologic treatment exists. However, JAK-2 prevalence in the CSVT cases is not clear. Methods: In this retrospective analysis, data of 236 patients with CSVT admitted to two tertiary centers between 2010 and 2020 were analyzed, with emphasis on laboratory and imaging data and clinical and interventional outcomes. Results: A total of 236 patients were included in the analysis. The patients' median age was 42 years and the average age was 44 years (±19 years), with 59% female patients. JAK-2 positivity rate was 18% (among 77 patients tested for the mutation). Patients with normal blood counts on presentation comprised 36% of the JAK-2 positive cases. Other hypercoagulability states were also investigated, with the antiphospholipid syndrome (APLA) showing the highest prevalence (11%) followed by other etiologies including oral contraceptive use, Factor V Leiden, prothrombin mutation, and malignancy. Selected JAK-2, APLA, and prothrombin mutation cases showed a more severe clinical course. Conclusion: JAK-2 mutation is underdiagnosed and its screening may be warranted in the cases of idiopathic CSVT, even despite normal blood counts, to allow disease-modifying treatment and blood cell count monitoring. JAK-2, APLA, and prothrombin mutation may be associated with a more complicated clinical course.

Can other coronavirus infections cause a cryptogenic stroke in a young patient?

We present a challenging case of a young patient who presented with focal neurological signs following a course of OC-43 coronavirus-related pneumonia almost 8 months before the COVID-19 outbreak.

Role of Thrombin in Central Nervous System Injury and Disease.

Thrombin is a Na+-activated allosteric serine protease of the chymotrypsin family involved in coagulation, inflammation, cell protection, and apoptosis. Increasingly, the role of thrombin in the brain has been explored. Low concentrations of thrombin are neuroprotective, while high concentrations exert pathological effects. However, greater attention regarding the involvement of thrombin in normal and pathological processes in the central nervous system is warranted. In this review, we explore the mechanisms of thrombin action, localization, and functions in the central nervous system and describe the involvement of thrombin in stroke and intracerebral hemorrhage, neurodegenerative diseases, epilepsy, traumatic brain injury, and primary central nervous system tumors. We aim to comprehensively characterize the role of thrombin in neurological disease and injury.

Markers for neural degeneration and regeneration: novel highly sensitive methods for the measurement of thrombin and activated protein C in human cerebrospinal fluid.

Inflammation and coagulation are tightly interconnected in the pathophysiology of neuronal diseases. Thrombin, a pro-coagulant serine protease is associated with neurodegeneration and its indirect inhibitor, activated protein C (aPC), is considered neuroprotective. While levels of thrombin and aPC activity are readily measured in the blood, similar assays in the cerebrospinal fluid (CSF) have not been described. The aim of this study was to establish a specific and sensitive enzymatic assay to measure both thrombin and aPC activity in the CSF. CSF was collected from 14 patients with suspected normal pressure hydrocephalus served as a control group, while seven patients with central nervous system infections served as an acute neuro-inflammatory study group and one sample of CSF following traumatic lumbar puncture served as a positive control. Thrombin and aPC activities were measured by fluorescence released by specific proteolytic cleavage in the presence of endopeptidase and amino-peptidase inhibitors to ensure specificity. Specificity of the method was verified by thrombin and serine-protease inhibitors N-alpha-((2-naphthylsulfinyl)glycyl)-DL-p-amidinophenylalanylpiperidine and phenylmethanesulfonyl fluoride. Inhibition of thrombin activity by CSF samples and levels of specific thrombin inhibitors were also assessed. Thrombin and aPC activities were reliably measured and were significantly higher in the CSF of patients with central nervous system infections compared to normal pressure hydrocephalus controls, suggesting the involvement of these factors in neuro-inflammation. CSF thrombin activity levels in the presence of known thrombin concentration were high in patients with central nervous system infections, and low in normal pressure hydrocephalus patients. Quantification of endogenous thrombin inhibitors protease nexin 1, amyloid precursor protein and anti-thrombin III in CSF by western blot indicated a significant elevation of amyloid precursor protein in infectious CSF. In conclusion, this study describes a novel and sensitive assay aimed at the detection of thrombin and aPC activity in CSF. This method may be useful for measuring these factors that reflect degenerative and protective influences of coagulation on neurological disorders. The study procedure was approved by the Ethics Committee of the Chaim Sheba Medical Center (approval No. 4245-17-SMC) on October 18, 2018.

In-thrombus thrombin secretion: a new diagnostic marker of atrial fibrillation in cryptogenic stroke.

BACKGROUND: Endovascularly retrieved clots are a potential resource for diagnosing stroke etiology, which may influence secondary prevention treatment. In this study we measured thrombin activity eluted by serially washing clots. METHODS: Clots were retrieved from 68 patients with acute ischemic stroke, freshly frozen and classified by standard criteria into proven atrial fibrillation (AF, 18 patients), atherosclerotic origin (AS, 15 patients), cryptogenic stroke (Cr, 17 patients) and other known causes (18 patients). Thawed clot samples were washed by transferring them into 1 mL buffer in seven hourly cycles and a fluorescent substrate assay was used to measure secreted thrombin activity. The clots were also examined histologically. Artificial fibrin and red blood cell-rich clots were similarly assayed for wash-eluted thrombin activity as an external control. RESULTS: Thrombin activity eluted from clots of AF origin decreased significantly with time in contrast to steady levels eluted from AS origin thrombi (P<0.0001 by repeated measures ANOVA). The Cr stroke group was indistinguishable from the AF group and differed statistically from the AS group (P=0.017 by repeated measures ANOVA). In artificial clots we found a biphasic activity pattern, with initially decreasing levels of eluted thrombin (AF pattern) and then, with continuing washes, steady eluted thrombin levels (AS pattern). CONCLUSIONS: An assay measuring the change in thrombin in clots retrieved during acute stroke endovascular thrombectomy procedures may serve as a diagnostic marker of the origin of the clot. The suggested mechanism for these differences may be the clot location before its retrieval, with high blood flow causing thrombin washout in atherosclerotic clots, in contrast to atrium appendage low blood flow retaining high thrombin levels.

Incorporation of relative cerebral blood flow into CT perfusion maps reduces false 'at risk' penumbra.

PURPOSE: The region defined as 'at risk' penumbra by current CT perfusion (CTP) maps is largely overestimated. We aimed to quantitate the portion of true 'at risk' tissue within CTP penumbra and to determine the parameter and threshold that would optimally distinguish it from false 'at risk' tissue, that is, benign oligaemia. METHODS: Among acute stroke patients evaluated by multimodal CT (NCCT/CTA/CTP) we identified those that had not undergone endovascular/thrombolytic treatment and had follow-up NCCT. Maps of absolute and relative CBF, CBV, MTT, TTP and Tmax as well as summary maps depicting infarcted and penumbral regions were generated using the Intellispace Portal (Philips Healthcare, Best, Netherlands). Follow-up CT was automatically co-registered to the CTP scan and the final infarct region was manually outlined. Perfusion parameters were systematically analysed - the parameter that resulted in the highest true-negative-rate (ie, proportion of benign oligaemia correctly identified) at a fixed, clinically relevant false-negative-rate (ie, proportion of 'missed' infarct) of 15%, was chosen as optimal. It was then re-applied to the CTP data to produce corrected perfusion maps. RESULTS: Forty seven acute stroke patients met selection criteria. Average portion of infarcted tissue within CTP penumbra was 15%±2.2%. Relative CBF at a threshold of 0.65 yielded the highest average true-negative-rate (48%), enabling reduction of the false 'at risk' penumbral region by ~half. CONCLUSIONS: Applying a relative CBF threshold on relative MTT-based CTP maps can significantly reduce false 'at risk' penumbra. This step may help to avoid unnecessary endovascular interventions.

Stress and Corticosteroids Modulate Muscarinic Long Term Potentiation (mLTP) in the Hippocampus.

Stress influences synaptic plasticity, learning and memory in a steroid hormone receptor dependent manner. Based on these findings it has been proposed that stress could be a major risk factor for the development of cognitive decline and dementia. Interestingly, evidence has been provided that stress also affects muscarinic, i.e., acetylcholine (ACh)-mediated neurotransmission. To learn more about the impact of stress and steroids on synaptic plasticity, in this study, we investigated the effects of stress on muscarinic long term potentiation (mLTP). We report that multiple, unpredictable exposure to stress depresses carbachol (0.5 µM)-induced mLTP, while this effect of stress is not observed in hippocampal slices prepared from mice exposed only to a single stressful procedure. Furthermore, we demonstrate that activation of distinct steroid hormone receptors is involved in stress-mediated alterations of mLTP. Activation of mineralocorticoid receptors (MR) promotes mLTP, while glucocorticoid receptor (GR) activity impairs mLTP. These effects of multiple unpredictable stress on mLTP are long-lasting since they are detected even two weeks after the last stressful experience. Thus, multiple unpredictable events rather than a single stressful experience affect mLTP in a steroid hormone receptor dependent manner, suggesting that chronic unpredictable stress can lead to lasting alterations in hippocampal cholinergic plasticity.

Thrombin Activity and Thrombin Receptor in Rat Glioblastoma Model: Possible Markers and Targets for Intervention?

High-grade gliomas constitute a group of aggressive CNS cancers that have high morbidity and mortality rates. Despite extensive research, current therapeutic approaches enable survival beyond 2 years in rare cases only. Thrombin and its main CNS target, protease-activated receptor-1, have been implicated in tumor progression and brain edema. Our aim was to study protease-activated receptor-1 (PAR-1) protein expression and thrombin-like activity levels in both in vitro and in vivo models of glioblastoma and correlate them with the volume of the surrounding edema. We measured the presence of PAR-1 protein using fluorescence immunohistochemistry and assessed thrombin activity in various glial and non-glial cell lines and in a CNS-1 glioma rat model using a thrombin-specific fluorescent assay. Thrombin activity was found to be highly elevated in various high-grade glioma cell lines as well as in non-glial malignant cell lines. In the CNS-1 glioma model, the level of PAR-1 fluorescence in the tumor was significantly elevated compared to adjacent regions of reactive gliosis or distant brain areas. The elevated level of thrombin activity observed in the high-grade glioma positively correlated with tumor-induced brain edema. In conclusion, thrombin is secreted from glioma cells and PAR-1 may be a new biological marker for high-grade gliomas.