Circadian Biology and the Neurovascular Unit.

Mammalian physiology and cellular function are subject to significant oscillations over the course of every 24-hour day. It is likely that these daily rhythms will affect function as well as mechanisms of disease in the central nervous system. In this review, we attempt to survey and synthesize emerging studies that investigate how circadian biology may influence the neurovascular unit. We examine how circadian clocks may operate in neural, glial, and vascular compartments, review how circadian mechanisms regulate cell-cell signaling, assess interactions with aging and vascular comorbidities, and finally ask whether and how circadian effects and disruptions in rhythms may influence the risk and progression of pathophysiology in cerebrovascular disease. Overcoming identified challenges and leveraging opportunities for future research might support the development of novel circadian-based treatments for stroke.

Peripheral priming induces plastic transcriptomic and proteomic responses in circulating neutrophils required for pathogen containment.

Neutrophils rapidly respond to inflammation and infection, but to which degree their functional trajectories after mobilization from the bone marrow are shaped within the circulation remains vague. Experimental limitations have so far hampered neutrophil research in human disease. Here, using innovative fixation and single-cell-based toolsets, we profile human and murine neutrophil transcriptomes and proteomes during steady state and bacterial infection. We find that peripheral priming of circulating neutrophils leads to dynamic shifts dominated by conserved up-regulation of antimicrobial genes across neutrophil substates, facilitating pathogen containment. We show the TLR4/NF-κB signaling-dependent up-regulation of canonical neutrophil activation markers like CD177/NB-1 during acute inflammation, resulting in functional shifts in vivo. Blocking de novo RNA synthesis in circulating neutrophils abrogates these plastic shifts and prevents the adaptation of antibacterial neutrophil programs by up-regulation of distinct effector molecules upon infection. These data underline transcriptional plasticity as a relevant mechanism of functional neutrophil reprogramming during acute infection to foster bacterial containment within the circulation.

Not open and shut: Complex and prolonged blood-brain barrier responses after stroke.

Blood-brain barrier dysfunction (BBB) occurs rapidly after stroke and contributes to edema, inflammation, and secondary brain injury including haemorrhage. Two recent studies shed light on the temporal extent of post-stroke BBB dysfunction as well as its consequences for drug delivery. Zhang et al. found increases in BBB permeability that persist up to one-year post-ischemia. Despite increased paracellular leakage, Stanton et al. showed that transcellular transporter systems are required to deliver therapeutics into brain parenchyma. Both studies remind us of the complexity of BBB responses after stroke and provide novel entry points for future research into the underlying mechanisms.

Brain-derived Tau for Monitoring Brain Injury in Acute Ischemic Stroke.

The evolution of infarcts varies widely among patients with acute ischemic stroke (IS) and influences treatment decisions. Neuroimaging is not applicable for frequent monitoring and there is no blood-based biomarker to track ongoing brain injury in acute IS. Here, we examined the utility of plasma brain-derived tau (BD-tau) as a biomarker for brain injury in acute IS. We conducted the prospective, observational Precision Medicine in Stroke [PROMISE] study with serial blood sampling upon hospital admission and at days 2, 3, and 7 in patients with acute ischemic stroke (IS) and for comparison, in patients with stroke mimics (SM). We determined the temporal course of plasma BD-tau, its relation to infarct size and admission imaging-based metrics of brain injury, and its value to predict functional outcome. Upon admission (median time-from-onset, 4.4h), BD-tau levels in IS patients correlated with ASPECTS (ρ=-0.21, P<.0001) and were predictive of final infarct volume (ρ=0.26, P<.0001). In contrast to SM patients, BD-tau levels in IS patients increased from admission (median, 2.9 pg/ml [IQR, 1.8-4.8]) to day 2 (median time-from-onset, 22.7h; median BD-tau, 5.0 pg/ml [IQR, 2.6-10.3]; P<.0001). The rate of change of BD-tau from admission to day 2 was significantly associated with collateral supply (R2=0.10, P<.0001) and infarct progression (ρ=0.58, P<.0001). At day 2, BD-tau was predictive of final infarct volume (ρ=0.59, P<.0001) and showed superior value for predicting the 90-day mRS score compared with final infarct volume. In conclusion, in 502 patients with acute IS, plasma BD-tau was associated with imaging-based metrics of brain injury upon admission, increased within the first 24 hours in correlation with infarct progression, and at 24 hours was superior to final infarct volume in predicting 90-day functional outcome. Further research is needed to determine whether BD-tau assessments can inform decision-making in stroke care.

Larger ischemic cores and poor collaterals among large vessel occlusions presenting in the late evening.

BACKGROUND: Components critical to cerebral perfusion have been noted to oscillate over a 24-h cycle. We previously reported that ischemic core volume has a diurnal relationship with stroke onset time when examined as dichotomized epochs (i.e. Day, Evening, Night) in a cohort of over 1,500 large vessel occlusion (LVO) patients. In this follow-up analysis, our goal was to explore if there is a sinusoidal relationship between ischemic core, collateral status (as measured by HIR), and stroke onset time. METHODS: We retrospectively examined collection of LVO patients with baseline perfusion imaging performed within 24 h of stroke onset from four international comprehensive stroke centers. Both ischemic core volume and HIR, were utilized as the primary radiographic parameters. To evaluate for differences in these parameters over a continuous 24-h cycle, we conducted a sinusoidal regression analysis after linearly regressing out the confounders age and time to imaging. RESULTS: A total of 1506 LVO cases were included, with a median ischemic core volume of 13.0 cc (IQR: 0.0-42.0) and median HIR of 0.4 (IQR: 0.2-0.6). Ischemic core volume varied by stroke onset time in the unadjusted (p = 0.001) and adjusted (p = 0.003) sinusoidal regression analysis with a peak in core volume around 7:45PM. HIR similarly varied by stroke onset time in the unadjusted (p = 0.004) and adjusted (p = 0.002) models with a peak in HIR values at around 8:18PM. CONCLUSION: The results suggest that critical factors to the development of the ischemic core vary by stroke onset time and peak around 8PM. When placed in the context of prior studies, strongly suggest a diurnal component to the development of the ischemic core.

Periprocedural unfractionated heparin bolus during endovascular treatment in acute ischemic stroke does more harm than good.

BACKGROUND: Unfractionated heparin (UFH) bolus is occasionally administered during endovascular treatment (EVT) to reduce thrombotic complications in acute ischemic stroke patients. However, the MR CLEAN-MED trial showed an increase in symptomatic intracranial hemorrhages (sICH) and a non-significant shift towards worse functional outcome with UFH administration. We aimed to analyze the impact of periprocedural UFH bolus in a real-world setting in anterior (ACS) and posterior circulation stroke (PCS) patients. METHODS: We analyzed data from the German Stroke Registry-Endovascular Treatment using propensity score matching. Primary outcome was the modified Rankin Scale at 3 months, and secondary outcome measures included mortality, angiographic outcomes, post-EVT National Institute of Health Stroke Scale scores and ICH at 24 hours. RESULTS: Among 13,082 patients, 7948 with ACS (UFH bolus use in 15%) and 841 with PCS (UFH bolus use in 16.3%) were included in the propensity score matching analysis. Applying MR CLEAN-MED study criteria, UFH bolus was associated with worse functional outcomes (odds ratio [OR] 1.44; 95% CI 1.06-1.96). Analyzing all ACS and PCS patients, UFH bolus did not provide any net benefit. In ACS patients treated with intravenous thrombolysis (IVT), UFH bolus use was associated with worse functional outcomes (OR 2.40; 95% CI 1.34 to 5.06). CONCLUSION: Our findings show transferability of the MR CLEAN-MED results into a real-world setting, confirming a negative effect of periprocedural UFH on functional outcome in this subgroup of patients. Considering all ACS and PCS patients, periprocedural UFH did not provide a net benefit and appears to be harmful, particularly in IVT-treated patients.

Impact of ongoing intravenous thrombolysis until completion of endovascular treatment in large vessel occlusion stroke patients.

Background: Recent studies have implied that ongoing intravenous thrombolysis (IVT) during endovascular treatment (ET) improves functional outcomes in patients who have undergone stroke caused by a large vessel occlusion (LVO). In this study, we investigated the effect of ongoing IVT until completion of ET on procedure duration, first-pass thrombectomy rate, and periprocedural complications. Methods: We analyzed patients from the German Stroke Registry-Endovascular Treatment dataset, collected between June 2015 and December 2021. Primary outcomes were modified Rankin Scale (mRS) score after 3 months and achievement of a Thrombolysis In Cerebral Infarction (TICI) score of 2b-3. Secondary parameters included ET duration, first-pass thrombectomy, and periprocedural complications. Results: Of the 13,082 patients in the dataset, 1,639 met the study inclusion criteria. A total of n = 317 patients (19.3%) underwent ongoing IVT until completion of ET, while IVT was completed prior to ET in 1,322 patients (80.7%). Ongoing IVT was associated with higher rates of achievement of an mRS score of 0-2 (or a back-to-baseline) after 3 months [odds ratio (OR) 1.53; 95% confidence interval (CI) 1.08-2.17]. Furthermore, ongoing IVT was predictive of achievement of a TICI score of 2b-3 (OR 1.37; 95% CI 1.03-1.83) and of first-pass thrombectomy (OR 2.07; 95% CI 1.51-2.84), while reducing the rate of peri-interventional complications (OR 0.64; 95% CI 0.44-0.94) and reducing ET duration by 24 min [ß = -24.35; 95% CI -32.92-(-15.79)]. Conclusion: Our findings suggest that ongoing IVT until ET completion has a favorable impact on both clinical and angiographic outcomes, as well as on periprocedural conditions, regardless of the overall time intervals involved. Therefore, rapid ET after IVT should be sought in order to take advantage of the additive effect of ongoing IVT during ET. Future studies should consider IVT timing in the context of ET as a potential confounder and treatment target.

Association of the time of day of EVT with clinical outcomes and benefit from successful recanalization after stroke.

Experimental and neuroimaging studies suggest an influence of the time of day on acute infarct growth, but whether this could inform patient selection for acute treatments is uncertain. In a multicenter cohort of 9357 stroke patients undergoing endovascular treatment, morning treatment (05:00-10:59) was associated with lowest 90-day mRS scores (adjusted odds ratio, 1.27 [95% CI, 1.08-1.47]; p = 0.004). The association between successful recanalization and outcome was stronger in morning compared to evening-treated patients (pia = 0.046) with treatment benefit persisting until 24 h for morning-treated compared to 11.5 h for evening-treated patients suggesting that the time of day might inform patient selection for EVT.

Endovascular therapy in patients with internal carotid artery occlusion and patent circle of Willis.

BACKGROUND: Occlusion of the internal carotid artery (ICA) may extend into the middle or anterior cerebral artery (ICA-T) or be confined to the intracranial (ICA-I) or extracranial segment (ICA-E). While there is excellent evidence for endovascular therapy (EVT) in ICA-T occlusions, studies on EVT in non-tandem ICA-I or ICA-E occlusions are scarce. OBJECTIVE: To characterize EVT-treated patients with ICA-I- and ICA-E occlusion by comparing them with ICA-T occlusions. METHODS: The German Stroke Registry (GSR), a national, multicenter, prospective registry was searched for EVT-treated patients with isolated ICA occlusion between June 2015 and December 2021. We stratified patients by ICA occlusion site: (a) ICA-T, (b) ICA-I, (c) ICA-E. Baseline factors, procedural variables, technical (modified Thrombolysis in Cerebral Infarction (mTICI)), and functional outcomes (modified Rankin scale score at 3 months) were analyzed. RESULTS: Of 13 082 GSR patients, 2588 (19.8%) presented with an isolated ICA occlusion, thereof 1946 (75.2%) ICA-T, 366 (14.1%) ICA-I, and 276 (10.7%) ICA-E patients. The groups differed in age (77 vs 76 vs 74 years, Ptrend=0.02), sex (53.4 vs 48.9 vs 43.1% female, Ptrend<0.01), and stroke severity (median National Institutes of Health Stroke Scale score at admission 17 vs 14 vs 13 points, Ptrend<0.001). In comparison with ICA-T occlusions, both ICA-I and ICA-E occlusions had lower rates of successful recanalization (mTICI 2b/3: 85.4% vs 80.4% vs 76.3%; aOR (95% CI for ICA-I vs ICA-T 0.71 (0.53 to 0.95); aOR (95% CI) for ICA-E vs ICA-T 0.57 (0.42 to 0.78)). In adjusted analyses, ICA-E occlusion was associated with worse outcome when compared with ICA-T occlusion (mRS ordinal shift, cOR (95% CI) 0.70 (0.52 to 0.93)). CONCLUSION: Patient characteristics and outcomes differ substantially between ICA-T, ICA-I, and ICA-E occlusions. These results warrant further studies on EVT in ICA-I and ICA-E patients.

Consensus Recommendations for Standardized Data Elements, Scales, and Time Segmentations in Studies of Human Circadian/Diurnal Biology and Stroke.

Increasing evidence indicates that circadian and diurnal rhythms robustly influence stroke onset, mechanism, progression, recovery, and response to therapy in human patients. Pioneering initial investigations yielded important insights but were often single-center series, used basic imaging approaches, and used conflicting definitions of key data elements, including what constitutes daytime versus nighttime. Contemporary methodologic advances in human neurovascular investigation have the potential to substantially increase understanding, including the use of large multicenter and national data registries, detailed clinical trial data sets, analysis guided by individual patient chronotype, and multimodal computed tomographic and magnetic resonance imaging. To fully harness the power of these approaches to enhance pathophysiologic knowledge, an important foundational step is to develop standardized definitions and coding guides for data collection, permitting rapid aggregation of data acquired in different studies, and ensuring a common framework for analysis. To meet this need, the Leducq Consortium International pour la Recherche Circadienne sur l'AVC (CIRCA) convened a Consensus Statement Working Group of leading international researchers in cerebrovascular and circadian/diurnal biology. Using an iterative, mixed-methods process, the working group developed 79 data standards, including 48 common data elements (23 new and 25 modified/unmodified from existing common data elements), 14 intervals for time-anchored analyses of different granularity, and 7 formal, validated scales. This portfolio of standardized data structures is now available to assist researchers in the design, implementation, aggregation, and interpretation of clinical, imaging, and population research related to the influence of human circadian/diurnal biology upon ischemic and hemorrhagic stroke.

Decline of thrombolysis rates before endovascular therapy in patients with acute anterior circulation large vessel occlusion ischemic stroke: A multicenter analysis from the German Stroke Registry.

INTRODUCTION: In recent years, the role of intravenous thrombolysis (IVT) before endovascular stroke treatment (EVT) has been discussed intensively. Whether the discussion was accompanied by changing rates of bridging IVT is unknown. METHODS: Data were extracted from the prospectively maintained German Stroke Registry, including patients treated with EVT at one of 28 stroke centers in Germany between 2016 and 2021. Primary outcome parameters were the rate of bridging IVT (a) in the entire registry cohort and (b) in patients without formal contraindications to IVT (i.e. recent oral anticoagulants, time window ⩾4.5 h, extensive early ischemic changes) adjusted for demographic and clinical confounders. RESULTS: 10,162 patients (52.8% women, median age 77 years, median National Institutes of Health Stroke Scale score 14) were analyzed. In the entire cohort, the rate of bridging IVT decreased from 63.8% in 2016 to 43.6% in 2021 (average absolute annual decrease 3.1%, 95% CI 2.4%-3.8%), while the proportion of patients with at least one formal contraindication increased by only 1.2% annually (95% CI 0.6%-1.9%). Among 5460 patients without record of formal contraindications, the rate of bridging IVT decreased from 75.5% in 2016 to 63.2% in 2021 and was significantly associated with admission date in a multivariable model (average absolute annual decrease 1.4%, 95% CI 0.6%-2.2%). Clinical factors associated with lower odds of bridging IVT included diabetes mellitus, carotid-T-occlusion, dual antiplatelet therapy, and direct admission to a thrombectomy center. CONCLUSION: We observed a substantial decline in bridging IVT rates independent of demographic confounders and not explained by an increase in contraindications. This observation deserves further exploration in independent populations.

RCT versus real-world cohorts: Differences in patient characteristics drive associations with outcome after EVT.

Background: The selection of patients with large-vessel occlusion (LVO) stroke for endovascular treatment (EVT) depends on patient characteristics and procedural metrics. The relation of these variables to functional outcome after EVT has been assessed in numerous datasets from both randomized controlled trials (RCT) and real-world registries, but whether differences in their case mix modulate outcome prediction is unknown. Methods: We leveraged data from individual patients with anterior LVO stroke treated with EVT from completed RCTs from the Virtual International Stroke Trials Archive (N = 479) and from the German Stroke Registry (N = 4079). Cohorts were compared regarding (i) patient characteristics and procedural pre-EVT metrics, (ii) these variables' relation to functional outcome, and (iii) the performance of derived outcome prediction models. Relation to outcome (functional dependence defined by a modified Rankin Scale score of 3-6 at 90 days) was analyzed by logistic regression models and a machine learning algorithm. Results: Ten out of 11 analyzed baseline variables differed between the RCT and real-world cohort: RCT patients were younger, had higher admission NIHSS scores, and received thrombolysis more often (all p < 0.0001). Largest differences at the level of individual outcome predictors were observed for age (RCT: adjusted odds ratio (aOR), 1.29 (95% CI, 1.10-1.53) vs real-world aOR, 1.65 (95% CI, 1.54-1.78) per 10-year increments, p < 0.001). Treatment with intravenous thrombolysis was not significantly associated with functional outcome in the RCT cohort (aOR, 1.64 (95 % CI, 0.91-3.00)), but in the real-world cohort (aOR, 0.81 (95% CI, 0.69-0.96); p for cohort heterogeneity = 0.056). Outcome prediction was more accurate when constructing and testing the model using real-world data compared to construction with RCT data and testing on real-world data (area under the curve, 0.82 (95% CI, 0.79-0.85) vs 0.79 (95% CI, 0.77-0.80), p = 0.004). Conclusions: RCT and real-world cohorts considerably differ in patient characteristics, individual outcome predictor strength, and overall outcome prediction model performance.

Immobility-associated thromboprotection is conserved across mammalian species from bear to human.

Venous thromboembolism (VTE) comprising deep venous thrombosis and pulmonary embolism is a major cause of morbidity and mortality. Short-term immobility-related conditions are a major risk factor for the development of VTE. Paradoxically, long-term immobilized free-ranging hibernating brown bears and paralyzed spinal cord injury (SCI) patients are protected from VTE. We aimed to identify mechanisms of immobility-associated VTE protection in a cross-species approach. Mass spectrometry-based proteomics revealed an antithrombotic signature in platelets of hibernating brown bears with heat shock protein 47 (HSP47) as the most substantially reduced protein. HSP47 down-regulation or ablation attenuated immune cell activation and neutrophil extracellular trap formation, contributing to thromboprotection in bears, SCI patients, and mice. This cross-species conserved platelet signature may give rise to antithrombotic therapeutics and prognostic markers beyond immobility-associated VTE.

A unified classification approach rating clinical utility of protein biomarkers across neurologic diseases.

A major evolution from purely clinical diagnoses to biomarker supported clinical diagnosing has been occurring over the past years in neurology. High-throughput methods, such as next-generation sequencing and mass spectrometry-based proteomics along with improved neuroimaging methods, are accelerating this development. This calls for a consensus framework that is broadly applicable and provides a spot-on overview of the clinical validity of novel biomarkers. We propose a harmonized terminology and a uniform concept that stratifies biomarkers according to clinical context of use and evidence levels, adapted from existing frameworks in oncology with a strong focus on (epi)genetic markers and treatment context. We demonstrate that this framework allows for a consistent assessment of clinical validity across disease entities and that sufficient evidence for many clinical applications of protein biomarkers is lacking. Our framework may help to identify promising biomarker candidates and classify their applications by clinical context, aiming for routine clinical use of (protein) biomarkers in neurology.

Endovascular thrombectomy for basilar artery occlusion stroke: Analysis of the German Stroke Registry-Endovascular Treatment.

BACKGROUND AND PURPOSE: Acute ischemic stroke due to basilar artery occlusion (BAO) causes the most severe strokes and has a poor prognosis. Data regarding efficacy of endovascular thrombectomy in BAO are sparse. Therefore, in this study, we performed an analysis of the therapy of patients with BAO in routine clinical practice. METHODS: Patients enrolled between June 2015 and December 2019 in the German Stroke Registry-Endovascular Treatment (GSR-ET) were analyzed. Primary outcomes were successful reperfusion (modified Thrombolysis in Cerebral Infarction [mTICI] score of 2b-3), substantial neurological improvement (≥8-point National Institute of Health Stroke Scale [NIHSS] score reduction from admission to discharge or NIHSS score at discharge ≤1), and good functional outcome at 3 months (modified Rankin Scale [mRS] score of 0-2). RESULTS: Out of 6635 GSR-ET patients, 640 (9.6%) patients (age 72.2 ± 13.3, 43.3% female) experienced BAO (median [interquartile range] NIHSS score 17 [8, 27]). Successful reperfusion was achieved in 88.4%. Substantial neurological improvement at discharge was reached by 45.5%. At 3-month follow-up, good clinical outcome was observed in 31.1% of patients and the mortality rate was 39.2%. Analysis of mTICI3 versus mTICI2b groups showed considerable better outcome in those with mTICI3 (38.9% vs. 24.4%; p = 0.005). The strongest predictors of good functional outcome were intravenous thrombolysis (IVT) treatment (odds ratio [OR] 3.04, 95% confidence interval [CI] 1.76-5.23) and successful reperfusion (OR 4.92, 95% CI 1.15-21.11), while the effect of time between symptom onset and reperfusion seemed to be small. CONCLUSIONS: Acute reperfusion strategies in BAO are common in daily practice and can achieve good rates of successful reperfusion, neurological improvement and good functional outcome. Our data suggest that, in addition to IVT treatment, successful and, in particular, complete reperfusion (mTICI3) strongly predicts good outcome, while time from symptom onset seemed to have a lower impact.

Circadian rhythm of ischaemic core progression in human stroke.

INTRODUCTION: Experimental stroke studies suggest an influence of the time of day of stroke onset on infarct progression. Whether this holds true after human stroke is unknown, but would have implications for the design of randomised controlled trials, especially those on neuroprotection. METHODS: We pooled data from 583 patients with anterior large-vessel occlusion stroke from three prospectively recruited cohorts. Ischaemic core and penumbra volumes were determined with CT perfusion using automated thresholds. Core growth was calculated as the ratio of core volume and onset-to-imaging time. To determine circadian rhythmicity, we applied multivariable linear and sinusoidal regression analysis adjusting for potential baseline confounders. RESULTS: Patients with symptom onset at night showed larger ischaemic core volumes on admission compared with patients with onset during the day (median, 40.2 mL vs 33.8 mL), also in adjusted analyses (p=0.008). Sinusoidal analysis indicated a peak of core volumes with onset at 11pm. Core growth was faster at night compared with day onset (adjusted p=0.01), especially for shorter onset-to-imaging times. In contrast, penumbra volumes did not change across the 24-hour cycle. DISCUSSION: These results suggest that human infarct progression varies across the 24-hour cycle with potential implications for the design and interpretation of neuroprotection trials.

Machine Learning-Based Identification of Target Groups for Thrombectomy in Acute Stroke.

Whether endovascular thrombectomy (EVT) improves functional outcome in patients with large-vessel occlusion (LVO) stroke that do not comply with inclusion criteria of randomized controlled trials (RCTs) but that are considered for EVT in clinical practice is uncertain. We aimed to systematically identify patients with LVO stroke underrepresented in RCTs who might benefit from EVT. Following the premises that (i) patients without reperfusion after EVT represent a non-treated control group and (ii) the level of reperfusion affects outcome in patients with benefit from EVT but not in patients without treatment benefit, we systematically assessed the importance of reperfusion level on functional outcome prediction using machine learning in patients with LVO stroke treated with EVT in clinical practice (N = 5235, German-Stroke-Registry) and in patients treated with EVT or best medical management from RCTs (N = 1488, Virtual-International-Stroke-Trials-Archive). The importance of reperfusion level on outcome prediction in an RCT-like real-world cohort equaled the importance of EVT treatment allocation for outcome prediction in RCT data and was higher compared to an unselected real-world population. The importance of reperfusion level was magnified in patient groups underrepresented in RCTs, including patients with lower NIHSS scores (0-10), M2 occlusions, and lower ASPECTS (0-5 and 6-8). Reperfusion level was equally important in patients with vertebrobasilar as with anterior LVO stroke. The importance of reperfusion level for outcome prediction identifies patient target groups who likely benefit from EVT, including vertebrobasilar stroke patients and among patients underrepresented in RCT patients with low NIHSS scores, low ASPECTS, and M2 occlusions.