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Introduction: Hypertensive cerebral small vessel disease (HTN-cSVD) is the predominant microangiopathy in patients with a combination of lobar and deep cerebral microbleeds (CMBs) and intracerebral hemorrhage (mixed ICH/CMB). We tested the hypothesis that cerebral amyloid angiopathy (CAA) is also a contributing microangiopathy in mixed ICH/CMB patients with cortical superficial siderosis (cSS), a marker that is strongly associated with CAA. Methods: Brain MRIs from a prospective database of consecutive non-traumatic ICH patients admitted to a single referral center (2003 to 2019) were reviewed for the presence of CMBs, cSS, and non-hemorrhagic CAA markers (lobar lacunes, centrum semiovale enlarged perivascular spaces (CSO-EPVS), and multispot pattern of leukoaraiosis). The frequency of CAA markers and left ventricular hypertrophy (LVH), a marker for hypertensive end-organ damage, were compared between mixed ICH/CMB patients with cSS (mixed +cSS ) and without cSS (mixed -cSS ) in univariate and multivariate models. Results: Of the 1824 ICH patients, 40 had mixed +cSS and 256 had mixed -cSS . LVH was less common in patients with mixed +cSS compared to those with mixed -cSS (34% vs. 59%, p = 0.01). The frequency of multispot pattern (18% vs. 4%, p < 0.01) and severe CSO-EPVS (33% vs. 11%, p < 0.01) were higher in patients with mixed +cSS compared to mixed -cSS , whereas lobar lacune frequency was similar (23% vs. 20%, p = 0.67). In a logistic regression model, older age (aOR 1.04 per year, 95% CI [1.01-1.08], p = 0.01), presence of multispot pattern (aOR 5.82, 95% CI [1.82-18.60], p < 0.03), severe CSO-EPVS (aOR 4.08, 95% CI [1.73-9.61], p < 0.01), and LVH (aOR 0.43, 95% CI [0.20-0.94], p = 0.03) were independently associated with mixed +cSS after further adjustment for sex and hypertension (Table). Conclusions: CAA and HTN-cSVD appear to confer injury in patients with mixed ICH/CMB and cSS, a finding that warrants confirmation in studies incorporating advanced imaging/pathology.

Introduction: In the United States, the risk of stroke is greater among Black compared to White adults. Furthermore, hypertension has a greater impact on stroke risk among Black adults. We hypothesized that race-specific metabolites may account for some of this observed disparity. Methods: This was a case-cohort study nested in the REasons for Geographic and Racial Differences in Stroke (REGARDS) study. Incident ischemic stroke cases (n=1,075) were compared with a random cohort sample (n=968). Metabolites were measured by liquid chromatography-tandem mass spectrometry. Weighted logistic regression and Cox proportional hazard model were used to identify race-specific metabolites in association with prevalent hypertension and incident stroke respectively. Candidate metabolites were assessed for mediation between hypertension and incident ischemic stroke. Linear regression was used to explore the association of metabolites with lifestyle factors. Results: A total of 162 plasma metabolites were quantified. Of these, gluconic acid was associated with prevalent hypertension in Black adults (OR=1.86, 95% CI=1.39-2.47, P =2.58x10 -5 ) but not in White adults (OR=1.00, 95% CI=0.80-1.24, P =0.97; P for interaction=4.57x10 -4 ). Gluconic acid was also associated with incident ischemic stroke among Black adults (HR=1.53, 95% CI=1.28-1.81, P =1.76x10 -6 ) but not White adults (HR=1.16, 95% CI=1.00-1.34, P =0.057; P for interaction=0.019). Gluconic acid mediated 25.4% (95% CI=4.1%-46.8%, P =0.02) of the association between hypertension and ischemic stroke among Black adults. Specific lifestyle factors were linked to elevated gluconic acid level among Black adults in multivariable analysis, including a Southern diet (β=0.18, 95% CI=0.08-0.28, P P =0.001), and lack of exercise (β=0.26, 95% CI=0.01-0.51, P =0.045). Conclusions: Gluconic acid is a lifestyle-related biomarker that is associated with prevalent hypertension, incident ischemic stroke, and mediates the relationship between hypertension and ischemic stroke in Black adults. Key factors linked to gluconic acid include Southern diet, low educational attainment, and low physical activity.

BACKGROUND AND PURPOSE: Accurate automated infarct segmentation is needed for acute ischemic stroke studies relying on infarct volumes as an imaging phenotype or biomarker that require large numbers of subjects. This study investigates whether an ensemble of convolutional neural networks (CNN) trained on multiparametric DWI maps outperforms single networks trained on solo DWI parametric maps. MATERIALS AND METHODS: CNNs were trained on combinations of DWI, ADC, and low b-value-weighted images from 116 subjects. The performances of the networks (measured by Dice score, sensitivity and precision) were compared to one another and to ensembles of 5 networks. To assess the generalizability of the approach, the best performing model was applied to an independent evaluation cohort of 151 subjects. Agreement between manual and automated segmentations for identifying patients with large lesions volumes was calculated across multiple thresholds (21 cm(3), 31 cm(3), 51 cm(3), and 70 cm(3)). RESULTS: An ensemble of CNNs trained on DWI, ADC and low b-value-weighted images produced the most accurate acute infarct segmentation over individual networks (p

Introduction: Recently, NA1, a peptide designed to protect neurons against NMDA receptor-mediated excitotoxicity, demonstrated promise for acute stroke treatment. Enhanced efficiency of peptide delivery may result in improved efficacy. Hypothesis: The bioactive fragment of NA1 is nine amino acids of the carboxyl tail of GluN2B. Delivery of NR2B9c to cells in the brain is achieved through fusion with TAT, a peptide that binds to biological membranes without selectivity. We hypothesize that enhancing targeted delivery of NA1 to the ischemic brain improve the efficacy of NA1. Methods: We previously identified betulinic acid (BA) forming nanoparticles (NPs) as a novel formulation of stroke treatment delivery. After intravenous administration, BA NP penetrate the ischemic brain, reduce ischemia-induced infarction as an antioxidant agent, and allow for encapsulation and delivery of drugs. In this study, we engineered BA NPs for improved drug delivery preferentially to the ischemic brain through surface conjugation of AMD3100, and for acid-triggered drug release to selectively in the ischemic region, which is known to be acidic, through chemical modification. The resulting acid responsive, stroke- targeting antioxidant NPs, or ARSTA NPs, were evaluated for stroke treatment in the mouse transient middle cerebral artery occlusion (MCAO) model either alone or in combination with NA1, based on changes of infarct volumes, edema, mortality, and outcomes. Results: We found that ARSTA NPs enabled targeted delivery of NA1 preferentially to the ischemic brain and allowed for prompt drug release. We showed that intravenous delivery of 50 ug NA1 (or 1nM/g) encapsulated in 1 mg ARSTAs effectively reduced infarct volumes by 69.8%, edema by 60.3%, mortality (median survival >14 days). In contrast, delivery of ARSTA NPs alone (52.2%, 30.2%, 10) or NA1 (0.7%, 3.3%, 6 days) alone at a comparable dose exhibited significantly less efficacy. Consistently, treatment with NA1-loaded ARSTAs improved neurological outcomes in a degree significantly greater than that by either ARSTA NPs or NA1 alone. Conclusion: BA-NP encapsulating NA-1 may represent a novel platform for improved delivery to the ischemic brain. Improved delivery may result in improved efficacy.

Background and Aims: Advances in low-field MRI have enabled image acquisition at the point-of-care (POC). We aim to characterize ischemic lesions in low-field, POC MRI and assess its relationship with stroke severity in ischemic stroke patients. Methods: We performed POC MRI exams on ischemic stroke patients. T2-weighted (T2W), fluid-attenuated inversion recovery (FLAIR), and diffusion-weighted imaging (DWI) exams were acquired with a 64mT, portable bedside MRI system. Three raters computed signal intensity ratios (SIR) for each sequence. For every slice showing an infarct, an SIR was generated by dividing the mean signal intensity of the lesion by the mean signal intensity of the contralateral hemisphere. Infarct volumes were obtained by multiplying the lesion area of each slice by the slice thickness (5mm) and summing the cross-sectional areas. Volumes were correlated with National Institutes of Health Stroke Scale (NIHSS) scores at the time of scan. Results: We studied 18 ischemic stroke patients (50% women; ages 30-95 years). Two patients were studied at two and three serial timepoints, respectively. POC exams were obtained 2.7 ± 2.2 days after symptom onset. A total of 18 T2W, 17 FLAIR, and 18 DWI exams were obtained. Three exams (1 T2W; 1 FLAIR; 1 DWI) were excluded due to motion degradation. High field MRI exams (19 ± 16 hours from POC exams) demonstrated ischemic infarcts in 15 of the 18 patients. All POC T2W and FLAIR exams revealed infarcts in these patients, and 14 of the 17 DWI exams showed infarcts. Ischemic infarcts were seen as hyperintense lesions (SIR: T2W = 1.19 ± 0.10, FLAIR = 1.15 ± 0.08, DWI = 1.36 ± 0.17). Infarct volume significantly correlated with NIHSS scores (T2W: r = 0.71, p < 0.01; FLAIR: r = 0.65, p < 0.05; DWI: r = 0.65, p < 0.05). Conclusions: These preliminary data suggest that low-field, POC MRI may be useful in the clinical evaluation of ischemic stroke. Further work in larger cohorts is needed to elucidate the appearance of infarction on low-field imaging.

Background: Biomarkers may help identify patients most likely to benefit from therapies. We tested whether serum glial fibrillary acidic protein (GFAP), a biomarker elevated early after intracerebral hemorrhage (ICH) in response to blood-brain barrier disruption, is associated with hematoma expansion (HE) and outcome after ICH and whether GFAP levels modify the effect of factor VII treatment. Methods: We performed an exploratory analysis of the recombinant activated factor VII for acute ICH (FAST) trial. FAST collected serum GFAP levels were collected at admission within 4 hours of ICH onset prior to factor VII treatment. We used regression analyses to evaluate the associations between serum GFAP, HE (dichotomized as &gte; 33% or > 6 mL increase in ICH volume from baseline to 24h and as the absolute volume of expansion), and 3-month poor outcome (modified Rankin Scale score 4-6). We tested for interaction between GFAP and factor VII treatment by adding product terms to multivariable regression models. Results: Of 841 enrolled patients, we included 567 (67%) with available GFAP levels (mean age 64 [SD 13], female sex 203 [37%]). GFAP was associated with HE (adjusted odds ratio [OR] 1.54, 95% CI 1.10-2.17) and poor outcome (adjusted OR 1.86, 95% CI 1.18-3.09). Compared to patients in the lowest GFAP quartile, those in the highest quartile had 2 times the odds of HE (95% CI 1.08-3.89) and 2.7 times the odds of a poor outcome (95% CI 1.33-5.70). GFAP modified the association between factor VII treatment and HE volume (multivariable interaction p=0.04): treatment was not associated with reduced HE volume in the lowest GFAP quartile (β -0.44, 95% CI -3.54 to 2.67), but was associated with reduced HE volume in higher quartiles (β -3.82, 95% CI -7.58 to -0.06). Conclusions: In the FAST trial population, early GFAP levels were associated with HE and poor functional outcome. Factor VII treatment was associated with a greater reduction in HE volume in patients with higher GFAP levels. Serum GFAP may be useful for risk-stratifying patients early after ICH onset.

Introduction: The incidence of substance-related cardiac arrests (SRCA) has increased due to the opioid epidemic. Previous studies have shown that SRCA present differently than non-SRCA. Weinvestigated differences in the clinical courses between the two groups for patients who were initially comatose post-resuscitation. Methods: Data from out-of-hospital arrest patients seen between 2007-2019 were retrospectively analyzed. Arrests were classified as SRCA if the patient had a positive toxicology screen, witnesses testified to substance use, or drug paraphernalia was present at the scene. Arrest types were categorized as shockable or non-shockable. Cerebral edema classifications were based on MRI or CT radiology reports. Poor outcomes were defined as Cerebral Performance Category (CPC) > 3 at discharge. Differences in demographics, presentation and outcomes were compared (Fisher’s Exact Test or Exact Wilcoxon-Mann-Whitney). Multivariable backward stepwise logistic regression was performed. Results: SRCA patients (N=90) were younger (p Discussion: SRCA patients are more likely to have non-shockable arrests which in turn leads to cerebral edema development and potentially worse outcomes.

Background: Magnetic resonance imaging (MRI) is a powerful modality for diagnosing stroke. Conventionally, patients must travel to the location of a high-field MRI device. Advances in low-field MRI have enabled acquisition of clinically useful images using a portable device at the bedside. The feasibility of using point of care (POC) MRI in a clinical stroke setting is unknown. Objective: To determine the safety and feasibility of portable, bedside, low-field MRI in a clinical setting. Design/Methods: POC MRI exams were performed in Yale’s Neuroscience Intensive Care Unit (NICU) from July 2018 to August 2019. Images were acquired at the bedside using a standard 110V, 15A power outlet. The environment included the bedside vitals monitor, ventilators and intravenous infusion pumps. Exams were performed by research staff trained to operate the POC scanner in the absence of a trained MRI technician. No special precautions were necessary to remove ferrous metals from the room. Scan parameters were controlled using a tablet computer interface, and images were available immediately after acquisition. Results: POC MRI was obtained in 85 stroke cases (46% female, ages 18-96 years, 46% ischemic stroke, 34% intracerebral hemorrhage, 20% subarachnoid hemorrhage). Scans were obtained within 7 days of symptom onset. NIHSS scores ranged from 1 to 29 (median of 7). Of the 85 patients analyzed, 68 underwent T2-weighted imaging, 72 underwent FLAIR imaging, and 39 underwent diffusion weighted imaging (DWI). DWI was only tested in ischemic stroke cases. Patients’ BMI ranged from 20.0 to 46.5 with a median of 26.7. The majority 74 (87%) of patients completed the entire exam. Five patients (6%) were unable to fit in the scanner’s 30 cm opening, while 6 patients (7%) experienced claustrophobia resulting in early termination of the exam. Mean exam time was 28.9 ± 8.4 minutes. The 64 mT static magnetic field, gradient and RF pulses of the POC MRI scanner did not interfere with NICU equipment, and no significant adverse events occurred. Conclusions: We report the first use of a portable, low-field MRI system to image stroke patients at the bedside. This early work suggests our approach is safe and viable in a complex clinical care environment.

Introduction: Identification of patients likely to develop midline shift (MLS) after large-vessel occlusion (LVO) stroke is essential for appropriate triage and patient disposition. Studies have identified clinical and radiographic predictors of MLS, but with limited accuracy. Using an innovative assessment of cerebral autoregulation, we sought to develop an accurate predictive model for MLS. Methods: We prospectively enrolled 73 patients with LVO stroke. Beat-by-beat cerebral blood flow (transcranial Doppler) and arterial pressure (arterial catheter or finger photoplethysmography) were recorded within 24 hours of the stroke, and a 24-hour brain MRI was obtained to determine infarct volume and MLS. Autoregulatory function was quantified from pressure-flow relation via projection pursuit regression (PPR), allowing for characterization of 5 hemodynamic markers (Figure 1A). We assessed the predictive relation of autoregulatory capacity and radiological and clinical variables to MLS using recursive classification tree models. Results: PPR successfully quantified autoregulatory function in 50/73 (68.5%) patients within 24 hours of LVO ischemic stroke (age 63.9±13.6, 66% F, NIHSS 15.8±6.7). Of these 50 patients, most (78%) underwent endovascular therapy. Thirteen (26%) experienced 24-h MLS; in these patients, infarct volumes were larger (140.2 vs. 48.6 mL, P ), and ipsilateral (but not contralateral) falling slopes were steeper (1.1 vs. 0.7 cm·s -1 ·mmHg -1 , P=0.001 ). Among all clinical, demographic, and hemodynamic variables, only two (infarct volume, ipsilateral falling slope) significantly contributed to prediction of MLS (accuracy 94%; Figure 1B). Conclusions: This predictive model of MLS wields translatable potential for triaging level of care in patients suffering from LVO ischemic stroke, but further research, including optimization of the PPR algorithm as well as prospective use of the predictive model, is needed.

Background: Radiographic diagnosis of intracranial hemorrhage (ICH) is a critical determinant of stroke care pathways requiring patient transport to a neuroimaging suite. Advances in low-field MRI have made it possible to obtain clinically useful imaging at the point of care (POC). Aim: The aim of this study was to obtain preliminary data regarding the ability of a bedside POC MRI scanner to detect ICH. Methods: We studied 36 patients with a diagnosis of ICH (n=18) or ischemic stroke (n=18). Five blinded readers independently evaluated T2W and FLAIR exams acquired prospectively on a 64 mT, portable bedside MRI system (Hyperfine Research, Inc). Kappa coefficients (κ) were calculated to determine inter-rater agreement. Ground truth was obtained from the clinical report of the closest conventional imaging study (17.9 ± 10.4 hours) and verified by a core reader. For each exam, majority consensus among raters was used to determine sensitivity. Results: ICH volume ranged from 4 to 101 cc (median of 13 cc). Exams were acquired within 7 days of symptom onset (51.1 ± 28.8 hours). A pathologic lesion was identified on every exam with 100% sensitivity. Sensitivity for distinguishing any hemorrhage was 89% and specificity was 83%. The mean sensitivity and specificity for individual raters was 79% and 69%, respectively. When limited to supratentorial hemorrhage, consensus sensitivity was 94%. For ICH cases detected by all raters (n=9), there was 100% accuracy for localizing the bleed (lobar vs. non-lobar) with perfect agreement among raters (κ = 1, p Conclusions: These data suggest that low-field, POC MRI may be used to detect hemorrhagic stroke at the bedside. Further work is needed to evaluate this approach in the hyperacute setting and across a wide range of ICH characteristics.

Introduction: Prior studies have shown a linear relationship between CT-derived density and brain water uptake. We sought to determine whether intravenous (IV) glibenclamide (glyburide; BIIB093) would reduce water uptake and mass effect on serial CT scans from patients enrolled in the phase 2 GAMES-RP trial. Methods: Non-contrast CT scans performed between admission and day 7 (n=263) were analyzed in patients in the GAMES-RP modified intention-to-treat sample. Midline shift (MLS) was measured at the level of maximal lateral displacement of the septum pellucidum. CT-derived radiodensity was measured in pre-defined regions of interest (ROI) within the infarct. The same ROIs were mirrored to the contralateral hemisphere. The change in CT radiodensity (change in water uptake) was averaged among gray and white matter regions separately, and across both regions together. Repeated measures mixed effects models were used to assess the effect of IV glibenclamide on MLS or CT-measured water uptake. Results: There was a median of 3 CT scans [IQR 2-5] per patient during the first 7 days after stroke. Greater change in water uptake was associated with more MLS (β=0.17, 95% CI 0.15 to 0.20, p p =0.037) and reduced MLS (β= -1.78, 95% CI -2.87 to -0.67, p =0.002), and adjusted for time. Treatment with IV glibenclamide reduced both gray and white matter water uptake independently. In mediation analysis, gray matter edema (β=0.10, 95% CI 0.05 to 0.16, p p =0.427). Conclusions: IV glibenclamide reduced water uptake and MLS after large hemispheric infarction. A phase 3 study is underway to determine whether IV glibenclamide treatment improves clinical outcome.

Introduction: Galectin-3 (Gal-3) is a member of the beta-galactoside-binding protein family that has been implicated in cardiovascular disease and stroke. We sought to determine if baseline Gal-3 predicts functional outcome after acute ischemic stroke, and to compare its performance relative to several other prognostic biomarkers. Methods: We measured plasma Gal-3, soluble ST2 (sST2), and fatty acid binding protein 4 (FABP4) in 383 patients who presented with acute ischemic stroke and were enrolled in a two-center biomarker study. Additional analyses that include copeptin are currently underway. Functional outcome was assessed at 3 months with the modified Rankin Scale (mRS), with good outcome defined as mRS 0-2, and poor outcome defined as mRS 3-6. The relationships between each marker and outcome were evaluated using univariate and multivariate logistic regression. Results: Median admission Gal-3 level was 7.4 ng/mL [IQR 5.2-11.7], median sST2 was 41.3 ng/mL [IQR 30.1-57.3] and median FABP4 was 37.8 ng/mL [21.2-57.1] in 383 patients (mean age 69+/- 15 years; 41.5% women). In univariate analysis, each biomarker predicted poor outcome (Gal-3: OR 1.8, 95% CI 1.4 - 2.4, p Conclusion: Among three previously validated biomarkers, plasma Gal-3 level predicted 3-month functional outcome, and Gal-3 and sST2 predicted mortality. Replication in a second cohort is planned.

Background: For cardiac arrest survivors initially comatose after restoration of spontaneous circulation (ROSC), the extent of brain injury and expected neurologic outcome are crucial for patient management decisions. Advanced diffusion imaging approaches such as neurite orientation dispersion and density imaging (NODDI) or diffusion kurtosis imaging may provide additional insight into tissue integrity and potential for recovery of consciousness complementary to standard diffusion tensor imaging (DTI). Methods: Multi-shell diffusion imaging was acquired in a prospective study of comatose cardiac arrest patients and in 5 controls. Neurite orientation dispersion (OD), intracellular volume fraction (ICVF), mean kurtosis (MK), axial kurtosis (AK), radial kurtosis (RK), mean diffusivity (MD), axial diffusivity (AD), radial diffusivity (RD) and fractional anisotropy (FA) were calculated. Median whole-brain values in patients with poor outcomes (no arousal recovery [AR] by discharge) were compared with those with AR and to controls (1-way ANOVA, post-hoc 1-sided Wilcoxon exact test). Results: 18 patients (mean ±SD 48±23 y, 39% men) and 5 controls (37±19 y, 40% men) were analyzed. Median (range) Glasgow Coma Scale was 3 [3-5]. 10 patients exhibited AR, 8 did not. Median [IQR] time-to-MRI was 5 [4-8] days. FA (P=0.009), MK (P=0.017), AK (P=0.026), RK (P=0.014), OD (P=0.018) and ICVF (P=0.0038) were significantly different (see Figure). FA control values were greater than AR and no AR (P Discussion: This is the first report investigating early NODDI and diffusional heterogeneity changes in post-cardiac arrest comatose patients. Patients who failed to recover arousal demonstrated greater values for all kurtosis and NODDI metrics compared to controls. Potential bias from early withdrawal of life sustaining treatment and small cohorts are limitations.

Introduction: Soluble ST2 (sST2) is a member of the interleukin-1 receptor family known to predict outcome and mortality in cardiovascular disease and ischemic stroke. We sought to determine whether plasma sST2 concentration is associated with functional outcome after intraparenchymal hemorrhage. Methods: Concentration of sST2 was measured on plasma samples from a prospectively collected cohort of 75 patients who presented with spontaneous intraparenchymal hemorrhage. Blood samples were drawn between 24 and 72 hours after symptom onset. Soluble ST2 was measured using a commercially-available ELISA (Critical Diagnostics, San Diego, CA). Injury severity was determined using the admission Glasgow Coma Scale (GCS), with GCS ≤ 8 defined as severe injury. Outcome was assessed by modified Rankin Scale (mRS) at 90 days, with poor outcome defined as mRS 4-6. The association between sST2 and outcome, adjusted for clinical variables, was determined using ordinal logistic regression and receiver operating curve (ROC) analysis. Results: Median sST2 concentration was 54.6ng/mL [IQR 25.7-156.5] in the entire cohort. Patients with a more severe presentation (GCS ≤ 8) had a higher sST2 concentration (165.1ng/ml [IQR 91.2-238.3] vs. 36.8ng/ml [IQR 23.0-62.6], p114.0ng/mL predicted poor outcome with a sensitivity of 63.3%, specificity of 82.2%, and an area under the curve of 0.766. Plasma sST2 concentration was also associated with ordinal mRS at 90 days (OR 2.99, 95% CI 1.84-4.88, p < 0.0001). Other univariate predictors of ordinal mRS included admission GCS, admission NIHSS, hematoma volume, and ICH score. Plasma sST2 remained an independent predictor of mRS after adjustment for ICH score (adjusted OR 1.97, 95% CI 1.09-3.53, p = 0.024). Similarly, adjustment for admission NIHSS, admission GCS or hematoma volume demonstrated that sST2 remained an independent predictor of outcome. Conclusion: Plasma sST2 is independently associated with functional outcome after intraparenchymal hemorrhage. These results support the role of sST2 as a candidate biomarker for both ischemic and hemorrhagic stroke, and argue for further investigation of the ST2 pathway in the pathogenesis of neurologic injury.

Introduction/Hypothesis: The sterile inflammatory response involves activation of the immune system in the absence of infection following trauma, ischemia, or toxins. Matrix metalloproteinase-9 (MMP-9) is a neuro-inflammatory zinc metalloproteinase implicated in secondary injury after acute ischemic stroke. In order to determine whether MMP-9 was associated with a sterile inflammatory response, we examined the association between circulating MMP-9 and extracellular ATP [a marker of a damage-associated molecular pattern (DAMP)] and acute neutrophil count. Methods: We measured plasma MMP-9 and ATP levels in 338 patients who presented to two institutions with acute ischemic stroke and were enrolled in the Specialized Program of Translational Research in Acute Stroke (SPOTRIAS) study. Plasma samples and admission leukocyte subsets were collected at 7.0 ± 3.0 hours and 47.7 ± 7.3 hours from stroke onset. ATP was measured using a liquid chromatography-tandem mass spectrometry metabolomics assay. MMP-9 levels were measured by ELISA using a commercially available assay. Results: 338 acute stroke patients were available for analysis. MMP-9 was associated with plasma ATP on admission (Pearson r = 0.24, P Conclusions: MMP-9 is associated with markers of the sterile inflammatory response, including circulating plasma ATP levels and absolute neutrophil count. In vitro studies have demonstrated that an initial rise in extracellular ATP is associated with increased neutrophil-mediated MMP-9 release. These findings suggest that ATP may be functioning as a DAMP in response to acute ischemia, and is associated with MMP-9 acutely in patients. This study marks the first clinical cohort to begin to characterize determinants of the sterile inflammatory response in acute ischemic stroke.

Introduction: Soluble ST2 (sST2) is a member of the interleukin-1 receptor family that predicts mortality and outcome in cardiovascular disease. We aimed to determine whether baseline sST2 predicts functional outcome after stroke and examined its potential role in hemorrhagic transformation (HT). Methods: We measured plasma sST2 and matrix metalloproteinase-9 (MMP-9) in 646 patients who presented with acute ischemic stroke and were enrolled in the Specialized Program of Translational Research in Acute Stroke (SPOTRIAS) network biomarker study. Soluble ST2 and total MMP-9 were measured by ELISA. Functional outcome was assessed at 3 months with the modified Rankin Scale (mRS), with good outcome defined as mRS 0-2, and poor outcome defined as mRS 3-6. HT was classified in 246 patients with available CT scans, using the ECASS III criteria. The relationships between sST2, outcome, and HT were evaluated using multivariate logistic regression. Results: Median admission sST2 level was 35.0 ng/mL [IQR 25.7—49.8] in 646 patients (mean age 69 years; 44% women). The plasma concentration of sST2 was independently associated with poor outcome (OR 3.06, 95% CI 1.15-8.31, P =0.02) and mortality (OR 8.94, 95% CI 2.13-39.55, P =0.003) after adjustment for age, sex, NIHSS, admission glucose, cardioembolic stroke subtype, and tPA treatment. Of the 246 patients with imaging data available, HT occurred in 37 (15%) patients: 18 (48%) with hemorrhagic infarction type 1 (HI1), 11 (30%) with hemorrhagic infarction type 2 (HI2), 4 (11%) with parenchymal hemorrhage type 1 (PH1) and 4 (11%) with parenchymal hemorrhage type 2 (PH2). Admission sST2 level was associated with the subsequent development of any HT ( P =0.03) and with hemorrhagic infarction type 2 or greater (HI2, PH1, or PH2; P =0.008). Elevated sST2 independently predicted HI2 or greater after adjustment for age, MMP-9, blood glucose level, and tPA treatment (OR 9.27, 95% CI, 1.05-86.76, P =0.045). Conclusion: Plasma sST2 levels predicted mortality, functional outcome, and risk of HT after ischemic stroke. Further study of the potential link between sST2 and the brain injury response after stroke is warranted.

Introduction: Hemorrhagic transformation (HT) is an uncommon but adverse complication after a stroke that can cause secondary injury. We sought to determine whether glyburide can reduce HT in an ischemia/reperfusion model of stroke. Materials and Methods: We used a transient filament occlusion of the middle cerebral artery (tMCAo), with three hours of cerebral ischemia. Glyburide (n=10) was administered as an intravenous bolus followed by a subcutaneous continuous infusion using an osmotic pump to deliver a dose of 39.5 ug/day. Control animals (n=10) were administered dimethyl sulfoxide vehicle. At 48 hours, infarct size, brain edema and HT were analyzed. HT was quantified using three methods, 1) a categorical designation based on European Cooperative Acute Stroke Study (ECASS) criteria, 2) densitometry of color images and 3) direct measurement of the hemoglobin concentration using spectrophotometry. Results: Compared to vehicle control, continuous glyburide treatment improved neurological outcome (7.6±1.0 vs. 6.0±0.9, P3 , P Conclusion: In a prolonged ischemia/reperfusion model, glyburide reduced the amount of hemorrhagic transformation. Clinical evaluation of glyburide in combination with mechanical thrombectomy may be warranted in patients.

Background: Prior evidence suggests that younger patients are at increased risk for the deleterious consequences associated with edema after large hemispheric infarction (LHI). GAMES-RP was a double-blind, randomized, placebo controlled phase 2 study. Patients presenting to 18 centers with baseline infarct volumes between 82 and 300 cm 3 on MRI were randomized 1:1 within 10 hours of symptom onset. Although the primary analysis, which included decompressive craniectomy as part of a composite endpoint, suggested no difference between groups, there was a trend toward decreased mortality for patients treated with IV glyburide. Objective: Exploratory analyses of the GAMES-RP trial suggested that overall trend of decreased mortality for patients treated with IV glyburide may be more pronounced with younger patients. We hypothesized that patients age 70 and under who were treated with IV glyburide would have improved long term outcomes compared to placebo. Methods: This exploratory analysis included the subset of patients aged < 70 who met the Per-Protocol definition: IV glyburide (n=35) and placebo (n=30). We analyzed 12-month mortality, functional outcome by analysis of the modified Rankin Scale (mRS), functional improvement on the Barthel Index (BI), and quality of life using the EQ-5D. Differences were analyzed using Fisher’s exact test except for shift analysis, which used a two sided Mann-Whitney test. Results: Subjects treated with IV glyburide had lower mortality at 12 months (14% vs 40%, p=0.025, OR 4.00). Shift analysis revealed a trend towards improved functional outcome in IV glyburide treated patients (OR 2.24; p=0.080), higher score on the BI (mean 60.1; SD 37.9 vs mean 39.5; SD 40.1, p=0.03), and higher score on the EQ-5D (mean 0.49; SD 0.31 vs mean 0.33; SD 0.32, p=0.04). Conclusion: This post-hoc analysis suggests that patients under 70, who may be at highest risk for poor outcomes secondary to edema after large hemispheric infarction, may have improved survival, better functional outcome and improved quality of life following treatment with IV glyburide. A prospective study is required in this population in order to definitively establish that IV glyburide can improve outcome. Registry: NCT01794182

Background: In acute ischemic stroke (AIS), therapeutic decisions are increasingly being based upon the volume of likely-unsalvageable brain tissue, which is often estimated using DWI. Deep learning algorithms, e.g. convolutional neural networks (CNN), have been employed for chronic stroke lesion segmentation. Here we investigate the applicability of CNN for DWI lesion measurement in acute stroke. Methods: 50 AIS patients underwent DWI < 12h from last known well. Apparent diffusion coefficient maps, T2WI, and DWI were used as covariates in a 2D CNN (5-fold cross validation). Including convolutional, inception and fully connected dense layers, a CNN of 15 layers was trained using manually outlined DWI lesions. To avoid overfitting, statistical dropout, L1- and L2-regularization and batch-normalization were used. Automatically segmented lesion volumes (ALV) using a 50% risk threshold were compared to the manual lesion volumes (MLV) using Dice similarity index (DSI, a measure of overlap) and Spearman’s correlation coefficient. Subset analysis was performed evaluating results between small ( Results: The figure shows examples of CNN segmentation. The median [IQR] measured lesion volume and DSI were 25 [13-46] mL and 66% [35-75%], respectively. The correlation of MLV with ALV was 86% (P Discussion: Automatic DWI lesion segmentation for large lesions is feasible using CNN. CNN tended to overestimate the volumes of small lesions. Prior methods have used a priori heuristics or morphometric operations to remove artifacts. CNN methods show promise for “learning” to discriminate artifacts from real lesions.

Objective Commonly, patients undergoing craniotomy are admitted to an intensive care setting postoperatively to allow for close monitoring. We aim to determine the frequency with which patients who have undergone elective craniotomies require intensive care unit (ICU)–level interventions or experience significant complications during the postoperative period to identify a subset of patients for whom an alternative to ICU-level care may be appropriate. Methods Following Institutional Review Board approval, a prospective, consecutive cohort of adult patients undergoing elective craniotomy was established at the Massachusetts General Hospital between the dates of April 2010 and March 2011. Inclusion criteria were intradural operations requiring craniotomy performed on adults (18 years of age or older). Exclusion criteria were cases of an urgent or emergent nature, patients who remained intubated postoperatively, and patients who had a ventriculostomy drain in place at the conclusion of the case. Results Four hundred patients were analyzed. Univariate analysis revealed that patients with diabetes ( P = 0.00047), those who required intraoperative blood product administration ( P = 0.032), older patients ( P P = 0.041), and those who underwent longer surgical procedures ( P = 0.021) were more likely to require ICU-level interventions or experience significant postoperative complications. Multivariate analysis only found diabetes ( P = 0.0005) and age ( P = 0.0091) to be predictive of a patient's need for postoperative ICU admission. Conclusions Diabetes and older age predict the need for ICU-level intervention after elective craniotomy. Properly selected patients may not require postcraniotomy ICU monitoring. Further study of resource utilization is necessary to validate these preliminary findings, particularly in different hospital types.

Background: Increases in FLAIR MRI signal intensity (SI) after acute ischemic stroke (AIS) have been proposed as “tissue clocks”, reflecting the degree of ischemic injury and potential for recovery in brain tissue. We hypothesize that lower SI increases will be a biomarker for less severe tissue injury and hence is potentially salvageable. To test this, we investigated patients who were treated with tPA to determine whether better outcomes were associated with lower FLAIR SI. Methods: Using our Get with the Guidelines database, we retrospectively analyzed AIS patients admitted between 2011 to 2013 who received full-dose tPA based on institutional protocols. Patients were included if they received a CT scan at our hospital before tPA therapy, and MRI performed < 1 h post CT included a usable FLAIR scan and evidence of stroke on acute DWI. SI ratio (SIR) was calculated on the FLAIR by selecting region of interests in hyperintense FLAIR areas that coincided with the acute DWI lesion and matching contralateral regions. Logistic regression analysis was performed using forward stepwise analysis to combine age, sex, admission NIHSS, SIR and onset-to-treatment (OTT) time to predict discharge outcome. Good outcome was defined as discharge to home or in-patient rehabilitation hospital. Results: There were 129 AIS patients who received tPA, and 57 met our imaging criteria. Patient characteristics were: mean±SD age 70±15 years, median [IQR] NIHSS 13 [7-18], OTT 2.1±1.1 h, time-to-MRI 2.1±1.0 h, 58% female and median SIR 1.13 [1.05-1.26]. On a univariate basis, only age (P Discussion: Controlling for age and admission NIHSS, we found that lower FLAIR SIR was a significant predictor of discharge outcome while OTT was not. This suggests that “tissue clocks” may be more accurate than “time clocks” for predicting tissue outcome, and ultimately functional outcome.

Introduction: Perihematomal edema (PHE) expansion rate may be an independent predictor of poor functional outcome following spontaneous intracerebral hemorrhage (ICH). We examined whether this association varies by ICH location. Hypothesis: The effect size of PHE expansion rate on mortality and poor functional outcome will be greater for deep ICH compared to lobar ICH. Methods: Subjects (n=139) were retrospectively identified from a prospective ICH cohort enrolled from 2000-2013. Inclusion criteria: ≥18 years of age, spontaneous supratentorial ICH, and known time of onset. Exclusion criteria: infratentorial or primary intraventricular hemorrhage, subsequent surgery, trauma, or warfarin-related ICH. ICH, PHE, and intraventricular hemorrhage (IVH) volumes were measured from CT scans. PHE expansion rates were calculated from serial PHE volume measurements. Logistic regression assessed the association between PHE expansion rate and mortality or poor functional outcome (modified Rankin Scale >2) at 90 days. Odds ratios are per 0.04 mL/h. Results: PHE expansion rate from baseline to 24 hours (PHE24) predicts mortality for deep (p=0.03, OR 1.13[1.02-1.26]) and lobar ICH (p=0.02, OR 1.03[1.00-1.06]) in unadjusted regression, and in models adjusted for age (Deep: p=0.02; Lobar: p=0.03), blood pressure (Deep: p=0.02; Lobar: p=0.04), IVH volume (Deep: p=0.02; Lobar: p=0.05), Glasgow Coma Scale (Deep: p=0.03; Lobar: p=0.02), or time to baseline CT (Deep: p=0.05; Lobar: p=0.05). PHE24 also predicts mortality for lobar ICH adjusting for ICH volume (p=0.05, OR 1.03[1-1.06]). A significant interaction exists between ICH location and PHE expansion rate from baseline to 72 hours (PHE72) in models predicting mRS>2 (p=0.04). PHE72 predicts mRS>2 for deep but not lobar ICH (p-values not shown) in models that are unadjusted (p=0.02, OR 4.04[1.25-13.04]) or adjusted for ICH volume (p=0.02, OR 4.3[1.25-14.98]), age (p=0.03, OR 5.4[1.21-24.11]), blood pressure (p=0.05, OR 3.28[1.02-10.57]), IVH volume (p=0.02, OR 4.59[1.28-16.41]), GCS (p=0.02, OR 4.19[1.2-14.55]), or time to first CT (p=0.03, OR 4.02[1.19-13.56]). Conclusion: PHE72 predicts poor functional outcomes exclusively after deep ICH, whereas PHE24 predicts mortality for deep and lobar ICH.

Neuropathologic studies suggest an association between cerebral amyloid angiopathy (CAA) and small ischemic infarctions as well as hemorrhages. We examined the prevalence and associated risk factors for infarcts detected by diffusion-weighted imaging (DWI).We performed retrospective analysis of MR images from 78 subjects with a diagnosis of probable CAA and a similar aged group of 55 subjects with Alzheimer disease or mild cognitive impairment (AD/MCI) for comparison. DWI and apparent diffusion coefficient (ADC) maps were inspected for acute or subacute infarcts. We also examined the association between DWI lesions and demographic variables, conventional vascular risk factors, and radiographic markers of CAA severity such as number of hemorrhages on gradient-echo MRI and volume of T2-hyperintense white matter lesions.Twelve of 78 subjects with CAA (15%) had a total of 17 DWI-hyperintense lesions consistent with subacute cerebral infarctions vs 0 of 55 subjects with AD/MCI (p = 0.001). The DWI lesions were located primarily in cortex and subcortical white matter. CAA subjects with DWI lesions had a higher median number of total hemorrhages (22 vs 4, p = 0.025) and no difference in white matter hyperintensity volume or conventional vascular risk factors compared to subjects with CAA without lesions.MRI evidence of small subacute infarcts is present in a substantial proportion of living patients with advanced cerebral amyloid angiopathy (CAA). The presence of these lesions is associated with a higher burden of hemorrhages, but not with conventional vascular risk factors. This suggests that advanced CAA predisposes to ischemic infarction as well as intracerebral hemorrhage.

Introduction: In malignant infarction, brain edema leads to secondary neurological deterioration and poor outcome. We investigated whether swelling is associated with outcome in a broader range of stroke severity. Methods: Two research cohorts (NBO and EPITHET) comprising 98 acute stroke subjects with prospective serial brain magnetic resonance imaging (MRI) were analyzed. The categorical presence of swelling and/or infarct growth (IG) was assessed on diffusion weighted MRI (DWI) by comparing baseline and 3-5 day follow-up scans. IG was defined using the ASPECTS system (small IG = decrease in ASPECTS ≥ 1; large IG = decrease in ASPECTS ≥ 2). The increase in lesion volume (ΔDWI) was then subdivided into swelling and IG volumes using region-of-interest analysis. The relationship of these imaging markers with outcome was evaluated using univariate and multivariate regression, with poor outcome defined as a 90 day modified Rankin Scale score of 3-6. Receiver operating characteristic (ROC) curve analysis was undertaken to define a threshold for prediction of poor outcome. Results: The presence of swelling independently predicted poor outcome after adjustment for age, admission NIH stroke scale score, admission glucose, and baseline DWI volume (OR4.55, 95%CI 1.21-18.9, p Conclusions: Both swelling and IG contribute to total lesion growth in the days following stroke. Swelling is an independent predictor of poor outcome in moderate to severe stroke.

Background: In acute ischemic stroke (AIS) patients, multi-parametric MRI-based predictive algorithms have shown promise in identifying tissue at risk of infarction, but do not consider the intrinsic variations of normal or pathological tissue. We hypothesized that extending MRI-based algorithms to take into consideration tissue type will improve predictions of tissue outcome. Methods: We retrospectively analyzed AIS patients who received neither revascularization nor experimental interventional treatment, who underwent MRI within 12 h from the time since they were last known well and who had follow-up imaging >4 days. Perfusion- and diffusion-parametric maps were combined to predict tissue outcome using 2 models: 1) a generalized linear model (GLM) trained with data from the whole ipsilateral hemisphere (sGLM), irrespective of tissue type, or 2) an anatomically-weighted GLM (aGLM) that was calculated using a weighted average to combine results from models generated using entire white or gray matter regions only. Both methods were evaluated using jack-knifing and predicted and follow-up regions were compared in terms of accuracy (measured as area under the receiver operator characteristic curve, AUC), Dice similarity index (DSI) and root mean square error (RMSE). Results: Results from 109 patients (65% male, median 68 y IQR [55-77], NIHSS 14 [9-25]) showed that, compared to sGLM, aGLM’s predictions had higher DSI (0.48 [0.19-0.59], P Discussion: We showed that anatomically-weighted algorithms may better capture differences in tissue vulnerability in acute ischemic stroke, contributing to improved MRI-based tissue outcome predictions.

Introduction: Swelling is common after acute ischemic stroke but little is known about predictors for its development. We explored several putative markers of swelling using brain magnetic resonance imaging (MRI). Methods: We retrospectively analyzed a cohort of acute stroke subjects with serial research brain MRI at baseline, 6 hours later, and then at 24 and 48 hours after the baseline MRI. We defined swelling as the expansion in stroke volume from baseline to the follow-up scan up about 2 days after stroke onset. Change in volume was also examined between each MRI interval. Apparent diffusion coefficient (ADC) and fluid-attenuated inversion recovery (FLAIR) signal intensity ratios were developed by normalizing the average signal intensity within the stroke lesion to that of the normal contralateral hemisphere. The association between ADC ratio and FLAIR ratio with expansion was assessed. Results: Twenty-one subjects had 71 scans available at four time points with a median baseline stroke volume of 98 mL [interquartile range (IQR) 66-135 mL]. The median increase in volume was 55 mL (IQR 35-84 mL) from baseline MRI. The changes from baseline to 24 hours (r=0.89, p Conclusions: Acute stroke subjects exhibit differing trajectories of expansion, and those with greater total swelling show larger increases early on. The nadir ADC ratio, a marker of stroke severity and cytotoxic injury, correlated with total swelling. FLAIR ratio, a potential marker of vasogenic edema, was not correlated to lesion expansion in this cohort. Future studies are warranted to better understand the relationship between these imaging markers and clinical outcome.

Introduction: Diffusion tensor imaging (DTI) measurements of fractional anisotropy (FA) and apparent diffusion coefficient (ADC) within the cerebral peduncle (CP) and corticospinal tract (CST) correlate with functional outcomes in ischemic stroke. We evaluated the utility of FA and ADC as quantitative biomarkers of cytotoxic edema in acute stroke subjects treated with intravenous glyburide (RP-1127), which may prevent brain swelling after ischemia. Methods: Nine acute hemispheric stroke subjects enrolled in the Glyburide Advantage in Malignant Edema and Stroke (GAMES) Pilot study were compared to a cohort of seven stroke patients not treated with RP-1127. GAMES subjects received an intravenous bolus of RP-1127 within 10 hours of symptom onset, followed by an intravenous drip for 72 hours. The cohorts were matched for admission NIHSS score, infarct volume, and time from infarct to DTI data acquisition. CPs were manually traced on color FA maps and used as seed regions to identify CST fiber tracts using deterministic, streamline tractography. Ipsilesional FA and ADC values were normalized to contralesional values, yielding four quantitative metrics: CP FA ratio, CP ADC ratio, CST FA ratio, and CST ADC ratio. Results: Median NIHSS score was 19 in both cohorts. Mean time to DTI data acquisition was 2.5 ± 0.4 and 2.4 ± 0.6 days, and mean infarct volume was 170 ± 49 and 188 ± 13 cc in the GAMES and control cohorts, respectively. CP ADC ratios were significantly higher in patients treated with RP-1127 compared to the control cohort (1.05 vs. 0.98, p=0.045). No difference between the GAMES and control cohorts was observed for the CP FA ratio (1.02 vs. 0.98, p=0.30), CST FA ratio (1.0 vs. 0.99, p=0.78), or CST ADC ratio (0.92 vs. 0.93, p=0.75). Conclusions: CP ADC ratio may provide a quantitative imaging biomarker of response to RP-1127 in acute stroke. Higher CP ADC ratios reflect a reduction of cytotoxic edema within CST axons at the level of the CP, consistent with RP-1127’s proposed mechanism of preventing cellular edema.

Introduction: Malignant infarction is characterized by the formation of cerebral edema, and decompressive craniectomy (DC) is the only proven therapy. The NC (Ca-ATP) channel, regulated by the SUR1 receptor, is blocked by the sulfonylurea drug glyburide, which attenuates brain water content in preclinical models of large stroke. A phase I study has been safely completed. We assessed the hypothesis that a pilot study of RP-1127 (Glyburide for Injection) in patients at high risk for malignant infarction was safe and feasible. Methods: The objective of this four-center prospective, open label, phase IIa trial was to assess the safety and feasibility of treating severe anterior circulation ischemic stroke patients with RP-1127, whether or not treated with IV rtPA. The study enrolled patients with a baseline MRI DWI lesion between 82 cm3 and 210 cm3, age 18-80 years, and time from symptom onset to drug infusion of ≤ 10 hours. Patients received a RP-1127 bolus followed by continuous infusion for 72 hours. Results: Recruitment of 10 patients was completed within 9.6 months. The mean initial DWI lesion volume was 102 ± 23 cm3. There were no serious adverse events related to the drug. The incidence of malignant edema was 20%, compared to 88% in a prospective observational study with DWI ≥82 cm3. Further, 8/10 patients did not require any bolus osmotherapy or DC. The mean increase in ipsilateral hemisphere volume was 50±33 cm3 compared to historical controls of 71.5±27 cm3. The proportion of 30 day mRS ≤ 4 was 90% compared to 23.8% (at 12 months) in control patients from a pooled analysis of DC trials. Conclusions: These findings suggest that a Phase II trial for patients at risk of malignant infarction is feasible and will provide important insights into the safety and potential efficacy of RP-1127.

Background: We investigated whether MRI-based algorithms combining acute DWI & PWI can be used to identify patients likely to experience lesion expansion. Methods: We analyzed MRI from 181 unilateral stroke patients who underwent DWI & PWI ≤ 12h from last seen well and had follow-up imaging (F/u) ≥4 days with lesions ≥ 1 cm 3 . Apparent diffusion coefficient, T2, DWI, CBF, CBV, MTT and Tmax (time of peak of deconvolved residue function) maps were co-registered, normalized and used as covariates in a model that produced infarction risk maps. Coefficients were calculated from 111 non-lysed patients and applied to 70 patients who received mechanical or drug lysis. Area under generated receiver operating characteristic curves (AUC) were calculated. Regional analyses were performed comparing mean risk values in Core (acute DWI), Growth (F/u - Core), Reverse (Core - F/u) and Normal (ipsilesional hemisphere - F/u) regions. Predicted lesion volumes (PLV) using a 50% risk threshold and post-hoc artifact removal for classifying infarcted tissue were correlated with the measured lesion volumes (MLV) on F/u. Values are median [IQR] or mean±SD. Results: The thrombolysis-treated group differed significantly from the non-lysis group in admission NIH Stroke Scale scores (12 [9-16] vs 6 [3-13]; P3 ; P=0.02) and F/u lesion volumes (39 [12-72] vs 17 [6-56] cm 3 ; P=0.02). No significant difference was found between age (67±15 vs 65±17 years old), male sex (59% vs 67%) and time to F/u (15 [6-47] vs 10 [6-46] days). Significant differences in predicted risk between the 4 regions were found for both lysed (Core: 0.72±0.15; Growth: 0.41±0.13; Reverse: 0.64±0.16; Normal: 0.25±0.06; P Conclusion: Mismatch in PLV and core can be used to identify patients most likely to experience lesion expansion if not given reperfusion therapy. The performance of the models was reduced in thrombolysed patients, which we propose is due to salvage of tissue that would have otherwise infarcted. MRI-based algorithms may identify patients who are not candidates for thrombolysis, yet have tissue to salvage. This may be useful as imaging selection criteria for future investigational trials of reperfusion therapy in extended time windows or for patients with unclear onsets.

The presence of anti-neutrophil cytoplasmic Abs (ANCA) in many patients with systemic vasculitis suggests that ANCA may play a role in disease pathogenesis. Neutrophils from patients with Wegener's granulomatosis often express ANCA target Ags (myeloperoxidase (MPO) and proteinase 3 (PR3)) on their surface, making these intracellular primary granule enzymes accessible to these autoantibodies. Similarly, normal neutrophils can be induced to translocate MPO and PR3 to the cell surface in vitro, and we demonstrate that murine mAb ANCA IgG, but not IgM, binds to the ANCA target and engages the Fc gamma RIIa ligand-binding site on the surface of human neutrophils. In contrast to ANCA IgM, ANCA IgG also induces an oxidative burst in neutrophils (oxidation of dihydrorhodamine = 91 +/- 15 fluorescence units with anti-PR3 IgG vs 17 +/- 2 with anti-PR3 IgM, p < 0.001). Blockade of the ligand-binding site of Fc gamma RIIa with an antibinding site mAb Fab significantly reduces this ANCA IgG-triggered production of reactive oxygen species (p < 0.01). Similarly, human ANCA bind the ANCA target, engage Fc gamma RIIa, and induce an oxidative burst in neutrophils. The allelic phenotype of Fc gamma RIIa strongly influences the Fc gamma receptor engagement by ligand, and Fc gamma RIIa homozygous donors differ by more than threefold in the quantitative production of reactive oxygen intermediates (ROI) (p < 0.01). Thus, engagement of Fc gamma RIIa by the Fc region of ANCA is one mechanism by which these autoantibodies activate receptor-mediated signal transduction systems in human neutrophils to initiate programs of inflammation and tissue injury. Fc gamma receptor alleles may represent heritable disease risk factors influencing the magnitude of such a process.

β- and γ-secretases, the proteases responsible for liberating the amyloid-β peptide (Aβ) from its precursor protein (APP), are considered important targets for the development of therapeutics for Alzheimer’s disease (AD). γ-Secretase has not been definitively identified, but its activity is closely linked to the multitransmembrane presenilins. Evidence using transition-state analogue inhibitors, site-directed mutagenesis, and molecular modeling suggests that γ-secretase is an aspartyl protease that catalyzes an intramembranous proteolysis. Two conserved transmembrane aspartates in presenilins are each critical for γ-secretase activity, and transition-state analogue γ-secretase inhibitors bind directly to presenilins. These results strongly suggest that presenilins are novel polytopic aspartyl proteases. The presenilins are also involved in the intramembranous proteolysis of the Notch receptor, a critical signaling event during cell fate decision in embryonic development. Transition-state analogue γ-secretase inhibitors also block the intramembranous cleavage of Notch, and the transmembrane aspartates of presenilins are required for this proteolysis as well. Thus, presenilins appear to play a similar role in the intramembranous processing of APP and Notch, raising concerns that toxic effects might result from chronic inhibition Notch signaling. We recently established in vitro γ-secretase assays using APP- and Notch-based substrates, and these assays should allow a rigorous biochemical comparison of these two related proteolytic processes. We also found that γ-secretase inhibitors can induce phenotypes mimicking Notch deficiencies in Drosophila, demonstrating that targeting this protease can interfere with Notch signaling in whole organisms.

Objective: We hypothesize that MRI based hemisphere volume increases in the first days after stroke is associated with neurological deterioration and functional outcome in patients with malignant edema. Background 70% of patients with malignant infarction die secondary to cerebral edema. A fundamental challenge in the development of treatment strategies is the lack of a validated imaging marker for brain edema. Design/Methods: We conducted a retrospective analysis of the EPITHET study MRI9s at stroke onset and at 3-5 days. We selected patients with an acute diffusion weighted imaging volume > 82cc, which is highly predictive for malignant edema. The involved hemisphere was manually outlined by two independent readers, and volumes were measured using Analyze software. We assessed inter-rater agreement, volume change between scans and the association of volume change with early neurological deterioration (END: NIHSS score worsening ≥4 points), 90-day modified Rankin Scale (mRS) score 0-4 and mortality. Results: Of 12 patients who met study criteria, average baseline and follow-up DWI lesion size was 138 cm 3 and 234 cm 3 , respectively. Mean time to follow-up MRI was 62 hours. Concordance correlation coefficients between readers were >0.90 for both hemisphere and ventricle volume assessment. Mean percent hemisphere volume increase was 16.2±8.3% (p Conclusions: In this exploratory analysis of severe ischemic stroke patients, statistically significant changes in hemisphere and ventricular volumes within the first week are consistent with expected changes of cerebral edema. MRI-based analysis of hemisphere growth appears to be a suitable biomarker for edema formation. Disclosure: Dr. Sheth has nothing to disclose. Dr. Yoo has nothing to disclose. Dr. Gonzalez has nothing to disclose. Dr. Kimberly has nothing to disclose. Dr. Chaudhry has nothing to disclose. Dr. Elm has nothing to disclose. Dr. Jacobson has received personal compensation for activities with Remedy Pharmaceuticals as the CEO.Dr. Jacobson has received compensation for serving as the CEO of Remedy Pharmaceuticals.Dr. Jacobson holds stock and/or stock options in Remedy Pharmaceuticals. Dr. Davis has received personal compensation for activities with Boehringer Ingelheim, Sanofi- Aventis, and Ever Pharma. Dr. Davis has received personal compensation in an editorial capacity for Associate Editor of Cerebrovascular Diseases. Dr. Donnan has nothing to disclose. Dr. Albers has received personal compensation for activities with Genentech and Lundbeck as a member of the Advisory Board. Dr. Albers has received research support from Genetech, Aventis, Sanofi, BI, Ono, and AstraZeneca. Dr. Stern has received personal compensation in an editorial capacity for the Neurologist. Dr. Stern has received research support from Remedy Pharmaceuticals, Inc.

The presence of anti-neutrophil cytoplasmic Abs (ANCA) in many patients with systemic vasculitis suggests that ANCA may play a role in disease pathogenesis. Neutrophils from patients with Wegener's granulomatosis often express ANCA target Ags (myeloperoxidase (MPO) and proteinase 3 (PR3)) on their surface, making these intracellular primary granule enzymes accessible to these autoantibodies. Similarly, normal neutrophils can be induced to translocate MPO and PR3 to the cell surface in vitro, and we demonstrate that murine mAb ANCA IgG, but not IgM, binds to the ANCA target and engages the Fc gamma RIIa ligand-binding site on the surface of human neutrophils. In contrast to ANCA IgM, ANCA IgG also induces an oxidative burst in neutrophils (oxidation of dihydrorhodamine = 91 +/- 15 fluorescence units with anti-PR3 IgG vs 17 +/- 2 with anti-PR3 IgM, p0.001). Blockade of the ligand-binding site of Fc gamma RIIa with an antibinding site mAb Fab significantly reduces this ANCA IgG-triggered production of reactive oxygen species (p0.01). Similarly, human ANCA bind the ANCA target, engage Fc gamma RIIa, and induce an oxidative burst in neutrophils. The allelic phenotype of Fc gamma RIIa strongly influences the Fc gamma receptor engagement by ligand, and Fc gamma RIIa homozygous donors differ by more than threefold in the quantitative production of reactive oxygen intermediates (ROI) (p0.01). Thus, engagement of Fc gamma RIIa by the Fc region of ANCA is one mechanism by which these autoantibodies activate receptor-mediated signal transduction systems in human neutrophils to initiate programs of inflammation and tissue injury. Fc gamma receptor alleles may represent heritable disease risk factors influencing the magnitude of such a process.

This study evaluated microRNA (miRNA) changes in cerebrospinal fluid (CSF) and their association with the occurrence of delayed cerebral ischemia (DCI) and poor functional outcome after SAH. Forty-three selected miRNAs were measured in daily CSF samples from a discovery cohort of SAH patients admitted to Rigshospitalet, Copenhagen, Denmark, and compared with neurologically healthy patients. Findings were validated in CSF from a replication cohort of SAH patients admitted to Massachusetts General Hospital, Boston, Massachusetts. The CSF levels of miRNA over time were compared with the occurrence of DCI, and functional outcome after 3 months. miRNAs were quantified in 427 CSF samples from 63 SAH patients in the discovery cohort, in 104 CSF samples from 63 SAH patients in the replication cohort, and in 11 CSF samples from 11 neurologically healthy patients. The miRNA profile changed remarkably immediately after SAH. Elevated miR-9-3p was associated with a poor functional outcome in the discovery cohort (p