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In this paper, the structure and the identity of fully 13C-substituted T-2 toxin were confirmed using high-resolution mass spectrometry, 1H-NMR, 13C-NMR, tandem mass spectrometry and HPLC–DAD. The purity of this compound was estimated to be at least 98.8% according to UV data. The isotopic distribution of (13C24) T-2 toxin indicated a total isotopic enrichment of 98.2 ± 1.0 atom% 13C, and the application of different MS measurement modes revealed the MS/MS fragmentation pattern of T-2 toxin. Furthermore, a stable isotope dilution mass spectrometry method for the quantification of T-2 toxin was developed using (13C24) T-2 toxin as internal standard. The method was evaluated with and without conventional clean-up and validated for maize and oats. Both cereals showed strong matrix enhancement effects, which could be compensated for through the application of the isotope-substituted internal standard. [ABSTRACT FROM AUTHOR]

Abstract Background The patho-mechanisms leading to brain damage due to cerebral malaria (CM) are yet not fully understood. Immune-mediated and ischaemic mechanisms have been implicated. The role of complement factors C1q, C3 and C5 for the pathogenesis of CM were investigated in this study. Methods C57BL/6J mice were infected with Plasmodium berghei ANKA blood stages. The clinical severity of the disease was assessed by a battery of 40 standardized tests for evaluating neurological functions in mice. Brain homogenates and sera of mice with CM, infected animals without CM and non-infected control animals were analyzed for C1q, C3 and C5 up-regulation by Western blotting. Results Densitometric analysis of Western blots of brain homogenates yielded statistically significant differences in the levels of C1q and C5 in the analyzed groups. Correlation analysis showed a statistically significant association of C1q and C5 levels with the clinical severity of the disease. More severely affected animals showed higher levels of C1q and C5. No differences in complement levels were observed between frontal and caudal parts of the brain. Densitometric analysis of Western blot of sera yielded statistically lower levels of C1q in infected animals without CM compared to animals of the control group. Conclusion The current study provides direct evidence for up-regulation of complement factors C1q and C5 in the brains of animals with CM. Local complement up-regulation is a possible mechanism for brain damage in experimental cerebral malaria.

Background: We aimed to investigate the prognostic role of β-synuclein in comparison to that of neurofilament light chain (NfL) and glial fibrillary acidic protein (GFAP) for predicting functional outcome after acute ischemic stroke (AIS)., Methods: We measured serum concentrations of β-synuclein, NfL and GFAP 24 h after hospital admission in 213 consecutive patients with moderate-to-severe AIS. We investigated the association between serum biomarkers and radiological/clinical characteristics, 3-months mortality and functional outcome on the modified Rankin Scale (mRS)., Results: In 213 patients with AIS [mean age: 76.1 (±12.5) years, 53.1% males, median NIHSS score on admission: 13 (IQR: 9-17)], higher levels of β-synuclein, NfL and GFAP were associated with higher NIHSS scores and with lower Alberta Stroke Program CT Score (ASPECTS) points on admission. Serum β-synuclein levels was significantly correlated with NfL (rho = 0.715, p < 0.001) and GFAP concentrations (rho = 0.684, p < 0.001). The inclusion of serum β-synuclein significantly improved the accuracy of prediction models without biomarkers for overall mortality (AUC: 0.836 vs. 0.752, p < 0.001) and mRS 3-6 vs. 0-2 (AUC: 0.812 vs. 0.624, p < 0.001). Combination models with NfL and/or GFAP showed a similar accuracy., Conclusions: Serum β-synuclein may be used to assess synaptic damage/dysfunction and to predict 3-months clinical outcomes in patients with AIS., (© 2024 The Author(s). European Journal of Neurology published by John Wiley & Sons Ltd on behalf of European Academy of Neurology.)

Equivalent circuits are one of the most used models for Li-ion cells in the automotive area. However, it is a challenge to these models to be able to capture the cell discharge capacity under different loads, while still being accurate on both continuous charge and dynamic tests, fast to compute, and easy to parametrize from non-specialized data. To tackle this challenge, this paper proposes an extension of the nonlinear double capacitor model by increasing its order, parameter dependency with C-rate, and an identification procedure that exploits the pseudo-linear nature of the problem to find the parameter maps. An analogy between the parts of the circuit and the single particle model is also presented to reduce the search space of the identification algorithm and to enhance model interpretability. The performance of the proposed model extension is analyzed and compared to a state-of-the-art model on a challenging LiFePO4 dataset with different characteristics and validated on a realistic drive cycle, obtaining a mean absolute average error of around 20 mV for both training and validation tests., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Background: Matrix metalloproteinases (MMPs) are implied in blood-brain barrier degradation and haemorrhagic transformation following ischaemic stroke, but their local relevance in the hyperacute disease phase is unknown. We aimed to examine ultra-early MMP-9 and MMP-2 release into collateral blood vessels, and to assess its prognostic value before therapeutic recanalisation by endovascular thrombectomy (EVT)., Methods: We report a cross-sectional proof-of-concept study including patients undergoing EVT for large-vessel ischaemic stroke at the University Hospital Würzburg, Germany. We obtained liquid biopsies from the collateral circulation before recanalisation, and systemic control samples. Laboratory workup included quantification of MMP-9 and MMP-2 plasma concentrations by cytometric bead array, immunohistochemical analyses of cellular MMP-9 and MMP-2 expression, and detection of proteolytic activity by gelatine zymography. The clinical impact of MMP concentrations was assessed by stratification according to intracranial haemorrhagic lesions on postinterventional computed tomography (Heidelberg Bleeding Classification, HBC) and early functional outcome (modified Rankin Scale, mRS). We used multivariable logistic regression, receiver-operating-characteristic (ROC) curves, and fixed-level estimates of test accuracy measures to study the prognostic value of MMP-9 concentrations., Findings: Between August 3, 2018, and September 16, 2021, 264 matched samples from 132 patients (86 [65.2%] women, 46 [34.8%] men, aged 40-94 years) were obtained. Median (interquartile range, IQR) MMP-9 (279.7 [IQR 126.4-569.6] vs 441 [IQR 223.4-731.5] ng/ml, p < 0.0001) but not MMP-2 concentrations were increased within collateral blood vessels. The median MMP-9 expression level of invading neutrophils was elevated (fluorescence intensity, arbitrary unit: 2276 [IQR 1007-5086] vs 3078 [IQR 1108-7963], p = 0.0018). Gelatine zymography experiments indicated the locally confined proteolytic activity of MMP-9 but not of MMP-2. Pretherapeutic MMP-9 release into stroke-affected brain regions predicted the degree of intracerebral haemorrhages and clinical stroke severity after recanalisation, and independently increased the odds of space-occupying parenchymal haematomas (HBC1c-3a) by 1.54 times, and the odds of severe disability or death (mRS ≥5 at hospital discharge) by 2.33 times per 1000 ng/ml increase. Excessive concentrations of MMP-9 indicated impending parenchymal haematomas and severe disability or death with high specificity., Interpretation: Measurement of MMP-9 within collateral blood vessels is feasible and identifies patients with stroke at risk of major intracerebral haemorrhages and poor outcome before therapeutic recanalisation by EVT, thereby providing evidence of the concept validity of ultra-early local stroke biomarkers., Funding: This work was funded by the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG) and the Interdisciplinary Centre for Clinical Research (IZKF) at the University of Würzburg., Competing Interests: Declaration of interests MP has received speaker honoraria from Bayer and Merck Serono; unrelated to the present study. KGH has received honoraria for acting as an advisor/speaker for Boehringer Ingelheim; unrelated to the present study. MKS declares a grant from CSL Behring, funding from the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG) project No. 424778381—CRC/TR 295, and is a co-holder of patent PCT/EP2020/068464; all unrelated to the present study. All other authors have nothing to disclose., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Objective: Conventional mock circulatory loops (MCLs) cannot replicate realistic hemodynamic conditions without inducing blood trauma. This constrains in-vitro hemocompatibility examinations of blood pumps to static test loops that do not mimic clinical scenarios. This study aimed at developing an atraumatic MCL based on a hardware-in-the-loop concept (H-MCL) for realistic hemocompatibility assessment., Methods: The H-MCL was designed for 450 ± 50 ml of blood with the polycarbonate reservoirs, the silicone/polyvinyl-chloride tubing, and the blood pump under investigation as the sole blood-contacting components. To account for inherent coupling effects a decoupling pressure control was derived by feedback linearization, whereas the level control was addressed by an optimization task to overcome periodic loss of controllability. The HeartMate 3 was showcased to evaluate the H-MCL's accuracy at typical hemodynamic conditions. To verify the atraumatic properties of the H-MCL, hemolysis (bovine blood, n = 6) was evaluated using the H-MCL in both inactive (static) and active (minor pulsatility) mode, and compared to results achieved in conventional loops., Results: Typical hemodynamic scenarios were replicated with marginal coupling effects and root mean square error (RMSE) below 1.74 ± 1.37 mmHg while the fluid level remained within ±4% of its target value. The normalized indices of hemolysis (NIH) for the inactive H-MCL showed no significant differences to conventional loops ( ∆NIH = -1.6 mg/100 L). Further, no significant difference was evident between the active and inactive mode in the H-MCL ( ∆NIH = +0.3 mg/100 L)., Conclusion and Significance: Collectively, these findings indicated the H-MCL's potential for in-vitro hemocompatibility assessment of blood pumps within realistic hemodynamic conditions, eliminating inherent setup-related risks for blood trauma.

Background: Retinal artery occlusion (RAO) may lead to irreversible blindness. For acute RAO, intravenous thrombolysis (IVT) can be considered as treatment. However, due to the rarity of RAO, data about IVT safety and effectiveness is limited., Methods: From the multicenter database ThRombolysis for Ischemic Stroke Patients (TRISP), we retrospectively analyzed visual acuity (VA) at baseline and within 3 months in IVT and non-IVT treated RAO patients. Primary outcome was difference of VA between baseline and follow up (∆VA). Secondary outcomes were rates of visual recovery (defined as improvement of VA ⩾ 0.3 logMAR), and safety (symptomatic intracranial hemorrhage (sICH) according to ECASS II criteria, asymptomatic intracranial hemorrhage (ICH) and major extracranial bleeding). Statistical analysis was performed using parametric tests and a linear regression model adjusted for age, sex and baseline VA., Results: We screened 200 patients with acute RAO and included 47 IVT and 34 non-IVT patients with complete information about recovery of vision. Visual Acuity at follow up significantly improved compared to baseline in IVT patients (∆VA 0.5 ± 0.8, p  < 0.001) and non-IVT patients (∆VA 0.40 ± 1.1, p  < 0.05). No significant differences in ∆VA and visual recovery rate were found between groups at follow up. Two asymptomatic ICH (4%) and one (2%) major extracranial bleeding (intraocular bleeding) occurred in the IVT group, while no bleeding events were reported in the non-IVT group., Conclusion: Our study provides real-life data from the largest cohort of IVT treated RAO patients published so far. While there is no evidence for superiority of IVT compared to conservative treatment, bleeding rates were low. A randomized controlled trial and standardized outcome assessments in RAO patients are justified to assess the net benefit of IVT in RAO., Competing Interests: Declaration of conflicting interestsThe author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: PB: Research funds by the Gottfried and Julia Bangerter-Rhyner Foundation and Swiss Academy of Medical Sciences and funding for travel and conference fees from BMS/Pfizer. SW: Research funds by the Swiss National Science Foundation, the UZH Clinical research priority program (CRPP) stroke, the Swiss Heart foundation, the Zurich Neuroscience Center (ZNZ), speaker honoraria from Springer, Teva Pharma and consultancy fees from Bayer and Novartis. STE has received funding for travel or speaker honoraria from Bayer, Boehringer Ingelheim and Daiichi-Sankyo. He has served on scientific advisory boards for Bayer, Boehringer Ingelheim, BMS/Pfizer, and MindMaze and on the editorial board of Stroke. His institutions have received an educational grant from Pfizer, compensation from Stago for educational efforts and research support from Daiichi-Sankyo, the Science Funds [Wissenschaftsfonds] of the University Hospital Basel, the University Basel, from the “Wissenschaftsfonds Rehabilitation” of the University Department for Geriatric Medicine Felix Platter, the “Freiwillige Akademische Gesellschaft Basel,” the Swiss Heart Foundation, and the Swiss National Science Foundation. PR has received honoraria for advisory board participation from Boehringer Ingelheim and Pfizer and lecture fees from Boehringer Ingelheim, Bayer, Pfizer and Daiichi Sankyo. His institution has received a grant from Boehringer Ingelheim for the ECASS-4 Study. RRL received funding for speaker honoraria from Pfizer, Boehringer Ingelheim, Biogen and Iscehma View. He has served on scientific advisory boards for Novo-Nordisk and Bayer.

Background and Purpose: This study aimed to investigate the effect of endovascular treatment (EVT, with or without intravenous thrombolysis [IVT]) versus IVT alone on outcomes in patients with acute ischemic stroke (AIS) and intracranial large vessel occlusion (LVO) attributable to cervical artery dissection (CeAD)., Methods: This multinational cohort study was conducted based on prospectively collected data from the EVA-TRISP (EndoVAscular treatment and ThRombolysis for Ischemic Stroke Patients) collaboration. Consecutive patients (2015-2019) with AIS-LVO attributable to CeAD treated with EVT and/or IVT were included. Primary outcome measures were (1) favorable 3-month outcome (modified Rankin Scale score 0-2) and (2) complete recanalization (thrombolysis in cerebral infarction scale 2b/3). Odds ratios with 95% confidence intervals (OR [95% CI]) from logistic regression models were calculated (unadjusted, adjusted). Secondary analyses were performed in the patients with LVO in the anterior circulation (LVOant) including propensity score matching., Results: Among 290 patients, 222 (76.6%) had EVT and 68 (23.4%) IVT alone. EVT-treated patients had more severe strokes (National Institutes of Health Stroke Scale score, median [interquartile range]: 14 [10-19] vs. 4 [2-7], P<0.001). The frequency of favorable 3-month outcome did not differ significantly between both groups (EVT: 64.0% vs. IVT: 86.8%; ORadjusted 0.56 [0.24-1.32]). EVT was associated with higher rates of recanalization (80.5% vs. 40.7%; ORadjusted 8.85 [4.28-18.29]) compared to IVT. All secondary analyses showed higher recanalization rates in the EVT-group, which however never translated into better functional outcome rates compared to the IVT-group., Conclusion: We observed no signal of superiority of EVT over IVT regarding functional outcome in CeAD-patients with AIS and LVO despite higher rates of complete recanalization with EVT. Whether pathophysiological CeAD-characteristics or their younger age might explain this observation deserves further research.

Background: The probability to receive intravenous thrombolysis (IVT) for treatment of acute ischemic stroke declines with increasing age and is consequently the lowest in very elderly patients. Safety concerns likely influence individual IVT treatment decisions. Using data from a large IVT registry, we aimed to provide more evidence on safety of IVT in the very elderly., Methods: In this prospective multicenter study from the TRISP (Thrombolysis in Ischemic Stroke Patients) registry, we compared patients ≥90 years with those <90 years using symptomatic intracranial hemorrhage (ECASS [European Cooperative Acute Stroke Study]-II criteria), death, and poor functional outcome in survivors (modified Rankin Scale score 3-5 for patients with prestroke modified Rankin Scale score ≤2 and modified Rankin Scale score 4-5 for patients prestroke modified Rankin Scale ≥3) at 3 months as outcomes. We calculated adjusted odds ratio with 95% CI using logistic regression models., Results: Of 16 974 eligible patients, 976 (5.7%) were ≥90 years. Patients ≥90 years had higher median National Institutes of Health Stroke Scale on admission (12 versus 8) and were more often dependent prior to the index stroke (prestroke modified Rankin Scale score of ≥3; 45.2% versus 7.4%). Occurrence of symptomatic intracranial hemorrhage (5.7% versus 4.4%, odds ratio adjusted 1.14 [0.83-1.57]) did not differ significantly between both groups. However, the probability of death (odds ratio adjusted 3.77 [3.14-4.53]) and poor functional outcome (odds ratio adjusted 2.63 [2.13-3.25]) was higher in patients aged ≥90 years. Results for the sample of centenarians (n=21) were similar., Conclusions: The probability of symptomatic intracranial hemorrhage after IVT in very elderly patients with stroke did not exceed that of their younger counterparts. The higher probability of death and poor functional outcome during follow-up in the very elderly seems not to be related to IVT treatment. Very high age itself should not be a reason to withhold IVT.

Purpose: The Thrombolysis in Ischemic Stroke Patients (TRISP) collaboration was a concerted effort initiated in 2010 with the purpose to address relevant research questions about the effectiveness and safety of intravenous thrombolysis (IVT). The collaboration also aims to prospectively collect data on patients undergoing endovascular treatment (EVT) and hence the name of the collaboration was changed from TRISP to EVA-TRISP. The methodology of the former TRISP registry for patients treated with IVT has already been published. This paper focuses on describing the EVT part of the registry., Participants: All centres committed to collecting predefined variables on consecutive patients prospectively. We aim for accuracy and completeness of the data and to adapt local databases to investigate novel research questions. Herein, we introduce the methodology of a recently constructed academic investigator-initiated open collaboration EVT registry built as an extension of an existing IVT registry in patients with acute ischaemic stroke (AIS)., Findings to Date: Currently, the EVA-TRISP network includes 20 stroke centres with considerable expertise in EVT and maintenance of high-quality hospital-based registries. Following several successful randomised controlled trials (RCTs), many important clinical questions remain unanswered in the (EVT) field and some of them will unlikely be investigated in future RCTs. Prospective registries with high-quality data on EVT-treated patients may help answering some of these unanswered issues, especially on safety and efficacy of EVT in specific patient subgroups., Future Plans: This collaborative effort aims at addressing clinically important questions on safety and efficacy of EVT in conditions not covered by RCTs. The TRISP registry generated substantial novel data supporting stroke physicians in their daily decision making considering IVT candidate patients. While providing observational data on EVT in daily clinical practice, our future findings may likewise be hypothesis generating for future research as well as for quality improvement (on EVT). The collaboration welcomes participation of further centres willing to fulfill the commitment and the outlined requirements., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

[Figure: see text].

Background: Transient ischemic attack (TIA) needs further diagnostic evaluation to prevent future ischemic stroke. However, prophylaxis can be harmful in elderly if the diagnosis is wrong. We aimed at characterizing differences in TIA mimics in younger and older patients to enhance diagnostic accuracy in elderly patients. Methods: In a dedicated neurological emergency room (nER) of a tertiary care University hospital, patients with transient neurological symptoms suspicious of TIA (<24 h) were retrospectively analyzed regarding their final diagnoses and their symptoms. These parameters were compared between patients aged 18-70 and >70 years using descriptive, univariable, and multivariable statistics. Results: From November 2018 until August 2019, 386 consecutive patients were included. 271 (70%) had cardiovascular risk factors and all patients received cerebral imaging, mostly CT [376 (97%)]. There was no difference in the rate of diagnosed TIA between the age groups [85 (46%) vs. 58 (39%); p = 0.213].TIA mimics in the elderly were more often internal medicine diseases [35 (19%) vs. 7 (5%); p < 0.001] and epileptic seizures [48 (26%) vs. 24 (16%); p = 0.032] but less often migraine [2 (1%) vs. 20 (13%); p < 0.001]. The most frequent symptoms in all patients were aphasia and dysarthria [107 (28%) and 92 (24%)]. Sensory impairments were less frequent in elderly patients [23 (11%) vs. 54 (30%); p < 0.001]. Impaired consciousness and orientation were independent predictors for TIA mimics ( p < 0.001) whereas facial palsy ( p < 0.001) motor weakness ( p < 0.001), dysarthria ( p = 0.022) and sensory impairment ( p < 0.001) were independent predictors of TIA. Conclusion: TIA mimics in elderly patients are more likely to be internal medicine diseases and epilepsy compared to younger patients. Excluding internal medicine diseases seems to be important in elderly patients. Facial palsy, motor weakness, dysarthria and sensory impairment are associated with TIA., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ippen, Walter, Hametner, Gumbinger, Nagel, Purrucker and Mundiyanapurath.)

Introduction: Alterations in haemoglobin levels are frequent in stroke patients. The prognostic meaning of anaemia and polyglobulia on outcomes in patients treated with intravenous thrombolysis is ambiguous., Patients and Methods: In this prospective multicentre, intravenous thrombolysis register-based study, we compared haemoglobin levels on hospital admission with three-month poor outcome (modified Rankin Scale 3-6), mortality and symptomatic intracranial haemorrhage (European Cooperative Acute Stroke Study II-criteria (ECASS-II-criteria)). Haemoglobin level was used as continuous and categorical variable distinguishing anaemia (female: <12 g/dl; male: <13 g/dl) and polyglobulia (female: >15.5 g/dl; male: >17 g/dl). Anaemia was subdivided into mild and moderate/severe (female/male: <11 g/dl). Normal haemoglobin level (female: 12.0-15.5 g/dl, male: 13.0-17.0 g/dl) served as reference group. Unadjusted and adjusted odds ratios with 95% confidence intervals were calculated with logistic regression models., Results: Among 6866 intravenous thrombolysis-treated stroke patients, 5448 (79.3%) had normal haemoglobin level, 1232 (17.9%) anaemia - of those 903 (13.2%) had mild and 329 (4.8%) moderate/severe anaemia - and 186 (2.7%) polyglobulia. Anaemia was associated with poor outcome (OR adjusted 1.25 (1.05-1.48)) and mortality (OR adjusted 1.58 (1.27-1.95)). In anaemia subgroups, both mild and moderate/severe anaemia independently predicted poor outcome (OR adjusted 1.29 (1.07-1.55) and 1.48 (1.09-2.02)) and mortality (OR adjusted 1.45 (1.15-1.84) and OR adjusted 2.00 (1.46-2.75)). Each haemoglobin level decrease by 1 g/dl independently increased the risk of poor outcome (OR adjusted 1.07 (1.02-1.11)) and mortality (OR adjusted 1.08 (1.02-1.15)). Anaemia was not associated with occurrence of symptomatic intracranial haemorrhage. Polyglobulia did not change any outcome., Discussion: The more severe the anaemia, the higher the probability of poor outcome and death. Severe anaemia might be a target for interventions in hyperacute stroke., Conclusion: Anaemia on admission, but not polyglobulia, is a strong and independent predictor of poor outcome and mortality in intravenous thrombolysis-treated stroke patients., (© European Stroke Organisation 2019.)

The mycotoxin zearalenone (ZEN) is produced by many plant pathogenic Fusarium species. It is well known for its estrogenic activity in humans and animals, but whether ZEN has a role in plant-pathogen interaction and which process it is targeting in planta was so far unclear. We found that treatment of Arabidopsis thaliana seedlings with ZEN induced transcription of the AtHSP90.1 gene. This heat shock protein (HSP) plays an important role in plant-pathogen interaction, assisting in stability and functionality of various disease resistance gene products. Inhibition of HSP90 ATPase activity impairs functionality. Because HSP90 inhibitors are known to induce HSP90 gene expression and due to the structural similarity with the known HSP90 inhibitor radicicol (RAD), we tested whether ZEN and its phase I metabolites α- and ß-zearalenol are also HSP90 ATPase inhibitors. Indeed, At HSP90.1 and wheat Ta HSP90-2 were inhibited by ZEN and ß-zearalenol, while α-zearalenol was almost inactive. Plants can efficiently glycosylate ZEN and α/ß-zearalenol. We therefore tested whether glucosylation has an effect on the inhibitory activity of these metabolites. Expression of the A. thaliana glucosyltransferase UGT73C6 conferred RAD resistance to a sensitive yeast strain. Glucosylation of RAD, ZEN, and α/ß-zearalenol abolished the in vitro inhibitory activity with recombinant HSP90 purified from Escherichia coli . In conclusion, the mycotoxin ZEN has a very prominent target in plants, HSP90, but it can be inactivated by glycosylation. This may explain why there is little evidence for a virulence function of ZEN in host plants., (Copyright © 2019 Torres Acosta, Michlmayr, Shams, Schweiger, Wiesenberger, Mitterbauer, Werner, Merz, Hauser, Hametner, Varga, Krska, Berthiller and Adam.)

Purpose: The ThRombolysis in Ischemic Stroke Patients (TRISP) collaboration aims to address clinically relevant questions about safety and outcomes of intravenous thrombolysis (IVT) and endovascular thrombectomy. The findings can provide observational information on treatment of patients derived from everyday clinical practice., Participants: TRISP is an open, investigator-driven collaborative research initiative of European stroke centres with expertise in treatment with revascularisation therapies and maintenance of hospital-based registries. All participating centres made a commitment to prospectively collect data on consecutive patients with stroke treated with IVT using standardised definitions of variables and outcomes, to assure accuracy and completeness of the data and to adapt their local databases to answer novel research questions., Findings to Date: Currently, TRISP comprises 18 centres and registers >10 000 IVT-treated patients. Prior TRISP projects provided evidence on the safety and functional outcome in relevant subgroups of patients who were excluded, under-represented or not specifically addressed in randomised controlled trials (ie, pre-existing disability, cervical artery dissections, stroke mimics, prior statin use), demonstrated deficits in organisation of acute stroke care (ie, IVT during non-working hours, effects of onset-to-door time on onset-to-needle time), evaluated the association between laboratory findings on outcome after IVT and served to develop risk estimation tools for prediction of haemorrhagic complications and functional outcome after IVT., Future Plans: Further TRISP projects to increase knowledge of the effect and safety of revascularisation therapies in acute stroke are ongoing. TRISP welcomes participation and project proposals of further centres fulfilling the outlined requirements. In the future, TRISP will be extended to include patients undergoing endovascular thrombectomy., Competing Interests: Competing interests: JFS has received speaker honoraria from W L Gore & Associates GmbH and travel support from Bayer and Boehringer-Ingelheim; HG has received research support from the Swiss National Science Foundation; AZ has received funding for speaker honoraria and consulting fees from Boehringer-Ingelheim and Medtronic-Covidien and consulting fees from Nestec; GK received grants from Swiss Parkinson Association, Swiss Heart Association, Swiss National Science Foundation and served on advisory boards for Boehringer-Ingelheim, Bayer, Daiichi Sankyo (Schweiz) AG, Zambon, Nestle, GE Healthcare within the last two years; YB received honoraria or consulting fees from AstraZeneca France, Daiichi-Sankyo, BMS-Pfizer, Covidiem, Bayer and MSD France; PM received research grants from the Swiss National Science Foundation, the Swiss Heart Foundation; speaker fees from Boehringer-Ingelheim, Bayer, Covidien and Stryker; honoraria from scientific advisory boards from Boehringer-Ingelheim, Bayer, Pfizer, Amgen; consulting fees from Pierre-Fabre and Astra-Zeneca; and travel support from Boehringer-Ingelheim and Bayer. All this support is received by the institution (CHUV) and is used for stroke education and research; DL participated during the last 5 years to 1 advisory boards, symposia or trials sponsored by Sanofi Aventis, BMS, Astrazeneca, Boeringher-Ingelheim, Servier, Ebewe, CoLucid Pharm, Brainsgate, Photothera, Lundbeck, GSK, Bayer, Pfizer et Allergan (honorarium paid to Adrinord or research account of the hospital) and was an associated editor of the Journal of neurology, neurosurgery and psychiatry 2004-2010 (personal financial compensation); CHN has received funding for travel or speaker honoraria from Bayer, Boehringer-Ingelheim, Takeda, and BMS/Pfizer; PJN has received consulting fees from Boehringer-Ingelheim; STE has received funding for travel or speaker honoraria from Bayer and Boehringer-Ingelheim, he has served on scientific advisory boards for Bayer, Boehringer-Ingelheim, BMS/Pfizer and Covidien and on the editorial board of Stroke. He has received an educational grant from Pfizer and research support from the Science Funds (Wissenschaftsfonds) of the University Hospital Basel, the University Basel, the Swiss Heart Foundation and the Swiss National Science Foundation., (© Author(s) (or their employer(s)) 2018. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Trichothecene toxins are confirmed or suspected virulence factors of various plant-pathogenic Fusarium species. Plants can detoxify these to a variable extent by glucosylation, a reaction catalyzed by UDP-glucosyltransferases (UGTs). Due to the unavailability of analytical standards for many trichothecene-glucoconjugates, information on such compounds is limited. Here, the previously identified deoxynivalenol-conjugating UGTs HvUGT13248 (barley), OsUGT79 (rice) and Bradi5g03300 ( Brachypodium ), were expressed in E. coli , affinity purified, and characterized towards their abilities to glucosylate the most relevant type A and B trichothecenes. HvUGT13248, which prefers nivalenol over deoxynivalenol, is also able to conjugate C-4 acetylated trichothecenes (e.g., T-2 toxin) to some degree while OsUGT79 and Bradi5g03300 are completely inactive with C-4 acetylated derivatives. The type A trichothecenes HT-2 toxin and T-2 triol are the kinetically preferred substrates in the case of HvUGT13248 and Bradi5g03300. We glucosylated several trichothecenes with OsUGT79 (HT-2 toxin, T-2 triol) and HvUGT13248 (T-2 toxin, neosolaniol, 4,15-diacetoxyscirpenol, fusarenon X) in the preparative scale. NMR analysis of the purified glucosides showed that exclusively β-D-glucosides were formed regio-selectively at position C-3-OH of the trichothecenes. These synthesized standards can be used to investigate the occurrence and toxicological properties of these modified mycotoxins., Competing Interests: The authors declare no conflict of interest.

Objective: To develop an NIH Stroke Scale (NIHSS)-compatible, all-in-one scale for rapid and comprehensive prehospital stroke assessment including stroke recognition, severity grading and progression monitoring as well as prediction of large vessel occlusion (LVO)., Methods: Emergency medical services (EMS) personnel and stroke physicians (n=326) rated each item of the NIHSS regarding suitability for prehospital use; best rated items were included. Stroke recognition was evaluated retrospectively in 689 consecutive patients with acute stroke or stroke mimics, prediction of LVO in 741 consecutive patients with ischaemic stroke with acute vessel imaging independent of admission NIHSS score., Results: Nine of the NIHSS items were rated as 'suitable for prehospital use.' After excluding two items in order to increase specificity, the final scale (termed shortened NIHSS for EMS, sNIHSS-EMS) consists of 'level of consciousness', 'facial palsy', 'motor arm/leg', 'sensory', 'language' and 'dysarthria'. Sensitivity for stroke recognition of the sNIHSS-EMS is 91% (95% CI 86 to 94), specificity 52% (95% CI 47 to 56). Receiver operating curve analysis revealed an optimal cut-off point for LVO prediction of ≥6 (sensitivity 70% (95% CI 65 to 76), specificity 81% (95% CI 76 to 84), positive predictive value 70 (95% CI 65 to 75), area under the curve 0.81 (95% CI 0.78 to 0.84)). Test characteristics were non-inferior to non-comprehensive scales., Conclusions: The sNIHSS-EMS may overcome the sequential use of multiple emergency stroke scales by permitting parallel stroke recognition, severity grading and LVO prediction. Full NIHSS-item compatibility allows for evaluation of stroke progression starting at the prehospital phase., Competing Interests: Competing interests: Personal fees, travel support, speaker honoraria or research grants were received from Bayer Healthcare (FH, HR, PAR, SN, SP), BeneChill (SP), BMS Pfizer (PAR, SP), Boehringer 8 Ingelheim (JCP, PAR, SN, SP), Brainomix Ltd. (SN), Covidien (SP), C.R. Bard (SP), EMCOOLS (SP), Helena Laboratories (FH, SP), HVM Medical (SP), Medtronic (SN), Pfizer (JCP, SN), Raumedic (SP) and ZOLL (SP). The other authors report no conflicts of interest., (© Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.)

Background and Purpose: Significance and management of blood pressure (BP) changes in acute stroke care are unclear. Here, we aimed to investigate the impact of 24-hour BP variability (BPV) on outcome in patients with acute ischemic stroke treated with intravenous thrombolysis., Methods: From the Safe Implementation of Treatment in Stroke International Stroke Thrombolysis registry, 28 976 patients with documented pre-treatment systolic BP at 2 and 24 hours were analyzed. The primary measure of BP variability was successive variability. Data were preprocessed using coarsened exact matching. We assessed early neurological improvement, symptomatic intracerebral hemorrhage (SICH), and long-term functional outcome (modified Rankin Scale [mRS] at 90 days) by binary and ordinal regression analyses., Results: Attempts to explain successive variation for analysis of BPV with patients characteristics at admission found systolic BP (5.5% variance) to be most influential, yet 92% of BPV variance remained unexplained. Independently from systolic BP, successive variation for analysis of BPV was associated with poor functional outcome mRS score of 0 to 2 (odds ratio [OR], 0.94; 95% confidence interval [CI], 0.90-0.98), disadvantage across the shift of mRS (OR, 1.04; 95% CI, 1.01-1.08), mortality (OR, 1.10; 95% CI, 1.01-1.08), SICH SITS (OR, 1.14; 95% CI, 1.06-1.23), and SICH ECASS (OR, 1.24; 95% CI, 1.10-1.40; ECASS [European Cooperative Acute Stroke Study 2]). Analyzing successive variation for analysis of BPV as a function of pre-treatment, systolic BP significantly improved the prediction of functional outcome (mRS score of 0-1, mRS score of 0-2, neurological improvement, mRS-shift: all P interaction <0.01). Excluding patients with atrial fibrillation in a sensitivity analysis gave consistent results overall., Conclusions: This study suggests the need for a more individual BP management accounting for pre-treatment BP and the acute BP course (ie, BPV) to achieve best possible outcome for the patient., (© 2017 American Heart Association, Inc.)

Fusarium Head Blight is a disease of cereal crops that causes severe yield losses and mycotoxin contamination of grain. The main causal pathogen, Fusarium graminearum, produces the trichothecene toxins deoxynivalenol or nivalenol as virulence factors. Nivalenol-producing isolates are most prevalent in Asia but co-exist with deoxynivalenol producers in lower frequency in North America and Europe. Previous studies identified a barley UDP-glucosyltransferase, HvUGT13248, that efficiently detoxifies deoxynivalenol, and when expressed in transgenic wheat results in high levels of type II resistance against deoxynivalenol-producing F. graminearum. Here we show that HvUGT13248 is also capable of converting nivalenol into the non-toxic nivalenol-3-O-β-d-glucoside. We describe the enzymatic preparation of a nivalenol-glucoside standard and its use in development of an analytical method to detect the nivalenol-glucoside conjugate. Recombinant Escherichia coli expressing HvUGT13248 glycosylates nivalenol more efficiently than deoxynivalenol. Overexpression in yeast, Arabidopsis thaliana, and wheat leads to increased nivalenol resistance. Increased ability to convert nivalenol to nivalenol-glucoside was observed in transgenic wheat, which also exhibits type II resistance to a nivalenol-producing F. graminearum strain. Our results demonstrate the HvUGT13248 can act to detoxify deoxynivalenol and nivalenol and provide resistance to deoxynivalenol- and nivalenol-producing Fusarium., (© The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Background and Purpose: Women have a worse outcome after stroke compared with men, although in intravenous thrombolysis (IVT)-treated patients, women seem to benefit more. Besides sex differences, age has also a possible effect on functional outcome. The interaction of sex on the functional outcome in IVT-treated patients in relation to age remains complex. The purpose of this study was to compare outcome after IVT between women and men with regard to age in a large multicenter European cohort reflecting daily clinical practice of acute stroke care., Methods: Data were obtained from IVT registries of 12 European tertiary hospitals. The primary outcome was poor functional outcome, defined as a modified Rankin scale score of 3 to 6 at 3 months. We stratified outcome by age in decades. Safety measures were symptomatic intracranial hemorrhage and mortality at 3 months., Results: In this cohort, 9495 patients were treated with IVT, and 4170 (43.9%) were women with a mean age of 71.9 years. After adjustments for baseline differences, female sex remained associated with poor functional outcome (odds ratio, 1.15; 95% confidence interval, 1.02-1.31). There was no association between sex and functional outcome when data were stratified by age. Symptomatic intracranial hemorrhage rate was similar in both sexes (adjusted odds ratio, 0.93; 95% confidence interval, 0.73-1.19), whereas mortality was lower among women (adjusted odds ratio, 0.83; 95% confidence interval, 0.70-0.99)., Conclusions: In this large cohort of IVT-treated patients, women more often had poor functional outcome compared with men. This difference was not dependent on age., (© 2017 American Heart Association, Inc.)

Zearalenone (ZEN) is an estrogenic mycotoxin occurring in Fusarium -infected cereals. Glucosylation is an important plant defense mechanism and generally reduces the acute toxicity of mycotoxins to humans and animals. Toxicological information about ZEN-glucosides is limited due to the unavailability of larger amounts required for animal studies. Hv UGT14077, a recently-validated ZEN-conjugating barley UDP-glucosyltransferase was expressed in Escherichia coli , affinity purified, and characterized. Hv UGT14077 possesses high affinity ( K m = 3 µM) and catalytic efficiency ( k cat / K m = 190 s -1 ·mM -1 ) with ZEN. It also efficiently glucosylates the phase-I ZEN-metabolites α-zearalenol and β-zearalenol, with k cat / K -glucosylation at C-14 and C-16 with preference of 14-glucoside synthesis. Furthermore, relatively slow consecutive formation of 14,16-di-glucosides was observed; their structures were tentatively identified by mass spectrometry and for ZEN-14,16-di-glucoside confirmed by nuclear magnetic resonance spectroscopy. Recombinant m of 40 and 74 s -1 ·mM -1 , respectively. Hv UGT14077 catalyzes O -glucosylation at C-14 and C-16 with preference of 14-glucoside synthesis. Furthermore, relatively slow consecutive formation of 14,16-di-glucosides was observed; their structures were tentatively identified by mass spectrometry and for ZEN-14,16-di-glucoside confirmed by nuclear magnetic resonance spectroscopy. Recombinant Hv UGT14077 allowed efficient preparative synthesis of ZEN-glucosides, yielding about 90% ZEN-14-glucoside and 10% ZEN-16-glucoside. The yield of ZEN-16-glucoside could be increased to 85% by co-incubation with a β-glucosidase highly selective for ZEN-14-glucoside. Depletion of the co-substrate UDP-glucose was counteracted by a sucrose synthase based regeneration system. This strategy could also be of interest to increase the yield of minor glucosides synthesized by other glucosyltransferases.

Background and Purpose: We hypothesized that any sex-related difference in outcome poststroke is explained by other prognostic factors and that the response to intravenous recombinant tissue-type plasminogen activator (r-tPA) is equal in males and females after adjustment for such factors., Methods: We accessed an independent collection of randomized clinical trials-the VISTA (Virtual International Stroke Trials Archive). Data were preprocessed by selecting complete cases (n=8028) and matching females to males (coarsened exact matching, n=4575, 24.3% r-tPA). Outcome was assessed by the 7-point modified Rankin Scale (mRS) measured at 90 days after ischemic stroke. Relationship among variables was estimated by adjusted regression analysis., Results: In nonthrombolyzed patients, ordinal analysis of mRS adjusting for stroke- and sex-related prognostic factors suggested comparable outcomes for females and males (odds ratio, 0.96; 95% confidence interval, 0.85-1.06). Females responded comparably to r-tPA as did males, irrespective of the outcome definition of mRS (ordinal: P Interaction =0.46, relative excess risk because of interaction=0). The number needed to treat was 6.8 and 11.2 for 1 female to achieve mRS score of 0 to 2 and 0 to 1, which was highly congruent with males. Analysis for a nonlinear variation of age-by-sex revealed a good outcome for females <45 years with significant disadvantage thereafter (mRS score of 0-2: P Interaction =0.004). No relationship between sex, r-tPA, and bleeding complications was evident., Conclusions: Functional outcome (mRS) without r-tPA was overall similar between the sexes, as was the response to r-tPA. Nonlinear sex-by-age interaction improved estimates of functional independence; this should be considered in sex-related studies in stroke., (© 2016 American Heart Association, Inc.)

Charasomes are convoluted plasma membrane domains in characean green algae. They are known to form in response to light via secretion of trans -Golgi network (TGN) vesicles and local inhibition of endocytosis. Charasomes are involved in the acidification of their aqueous environment, thereby facilitating photosynthesis-dependent carbon uptake. Charasome formation is reversible to allow cells to adapt to different light conditions. Here, we show that darkness-induced degradation of charasomes involves the formation of coated pits and coated vesicles. The darkness-induced degradation of charasomes can be inhibited by 1-2 μM ikarugamycin (IKA), which is considered to be a specific inhibitor of clathrin-dependent endocytosis. At a much higher concentration (100 μM), IKA also significantly reduces the internalization of styryl dyes, indicating uptake via clathrin-coated vesicles (CV). We are the first to present evidence, based on fine structure investigation, that IKA does not interfere with the formation of clathrin coat, but inhibits the detachment and/or further processing of coated vesicles. Both charasome degradation and constitutive endocytosis are also significantly inhibited by sterol complexing agents (methyl-ß-cyclodextrin and filipin). The absence of an additive effect, when applied together with IKA, suggests that charasome degradation and constitutive endocytosis (measured via styryl dye uptake) is not inhibited due to membrane retrieval via lipid rafts, but due to clathrin coat formation requirement of a specific set of sterols. Analysis of Chara australis clathrin proteins revealed two heavy chains and several light chains with sequence peculiarities, suggesting functional and/or species specific differences. The data obtained indicate that clathrin plays a central role not only in constitutive endocytosis but also in the degradation of charasomes, thereby representing a valuable system for studying targeted exo- and endocytosis.

Background and Purpose: We compared outcome and complications in patients with stroke treated with intravenous thrombolysis (IVT) who could not live alone without help of another person before stroke (dependent patients) versus independent ones., Methods: In a multicenter IVT-register-based cohort study, we compared previously dependent (prestroke modified Rankin Scale score, 3-5) versus independent (prestroke modified Rankin Scale score, 0-2) patients. Outcome measures were poor 3-month outcome (not reaching at least prestroke modified Rankin Scale [dependent patients]; modified Rankin Scale score of 3-6 [independent patients]), death, and symptomatic intracranial hemorrhage. Unadjusted and adjusted odds ratios (ORs) with 95% confidence intervals (OR [95% confidence interval]) were calculated., Results: Among 7430 IVT-treated patients, 489 (6.6%) were dependent and 6941 (93.4%) were independent. Previous stroke, dementia, heart, and bone diseases were the most common causes of preexisting dependency. Dependent patients were more likely to die (ORunadjusted, 4.55 [3.74-5.53]; ORadjusted, 2.19 [1.70-2.84]). Symptomatic intracranial hemorrhage occurred equally frequent (4.8% versus 4.5%). Poor outcome was more frequent in dependent (60.5%) than in independent (39.6%) patients, but the adjusted ORs were similar (ORadjusted, 0.95 [0.75-1.21]). Among survivors, the proportion of patients with poor outcome did not differ (35.7% versus 31.3%). After adjustment for age and stroke severity, the odds of poor outcome were lower in dependent patients (ORadjusted, 0.64 [0.49-0.84])., Conclusions: IVT-treated stroke patients who were dependent on the daily help of others before stroke carry a higher mortality risk than previously independent patients. The risk of symptomatic intracranial hemorrhage and the likelihood of poor outcome were not independently influenced by previous dependency. Among survivors, poor outcome was avoided at least as effectively in previously dependent patients. Thus, withholding IVT in previously dependent patients might not be justified., (© 2016 American Heart Association, Inc.)

Implementing endovascular stroke care often impedes neurologic assessment in patients who need sedation or general anesthesia. Cerebral near-infrared spectroscopy (NIRS) may help physicians monitor cerebral tissue viability, but data in hyperacute stroke patients receiving endovascular treatment are sparse. In this observational study, the NIRS index regional oxygen saturation (rSO2) was measured noninvasively before, during, and after endovascular therapy via bilateral forehead NIRS optodes. During the study period, 63 patients were monitored with NIRS; 43 qualified for analysis. Before recanalization, 10 distinct rSO2 decreases occurred in 11 patients with respect to time to intubation. During recanalization, two kinds of unilateral rSO2 changes occurred in the affected hemisphere: small peaks throughout the treatment (n=14, 32.6%) and sustained increases immediately after recanalization (n=2, 4.7%). Lower area under the curve 10% below baseline was associated with better reperfusion status (thrombolysis in cerebral infarction ≥ 2b, P=0.009). At the end of the intervention, lower interhemispheric rSO2 difference predicted death within 90 days (P=0.037). After the intervention, higher rSO2 variability predicted poor outcome (modified Rankin scale > 3, P=0.032). Our findings suggest that bi-channel rSO2-NIRS has potential for guiding neuroanesthesia and predicting outcome. To better monitor local revascularization, an improved stroke-specific set-up in future studies is necessary.

RAB5 GTPases are important regulators of endosomal membrane traffic. Among them Arabidopsis thaliana ARA7/RABF2b is highly conserved and homologues are present in fungal, animal and plant kingdoms. In land plants ARA7 and its homologues are involved in endocytosis and transport towards the vacuole. Here we report on the isolation of an ARA7 homologue (CaARA7/CaRABF2) in the highly evolved characean green alga Chara australis. It encodes a polypeptide of 202 amino acids with a calculated molecular mass of 22.2 kDa and intrinsic GTPase activity. Immunolabelling of internodal cells with a specific antibody reveals CaARA7 epitopes at multivesicular endosomes (MVEs) and at MVE-containing wortmannin (WM) compartments. When transiently expressed in epidermal cells of Nicotiana benthamiana leaves, fluorescently tagged CaARA7 localizes to small organelles (putative MVEs) and WM compartments, and partially colocalizes with AtARA7 and CaARA6, a plant specific RABF1 GTPase. Mutations in membrane anchoring and GTP binding sites alter localization of CaARA7 and affect GTPase activity, respectively. This first detailed study of a conventional RAB5 GTPase in green algae demonstrates that CaARA7 is similar to RAB5 GTPases from land plants and other organisms and shows conserved structure and localization., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: It is not established whether sex influences outcome and safety following intravenous thrombolysis (IVT) in acute stroke. As a significant imbalance exists between the baseline conditions of women and men, regression analysis alone may be subject to bias. Here we aimed to overcome this methodical shortcoming by balancing both groups using coarsened exact matching (CEM) before evaluating outcome., Methods: From our local prospective stroke database we analyzed consecutive patients who suffered anterior circulation stroke and received IVT from 1998 to 04/2013 (n = 1391, 668 female, 723 male). Data were preprocessed by CEM, balancing for age, NIHSS, lesion side, hypertension, diabetes, atrial fibrillation, smoking, coronary heart disease, and previous stroke, which yielded a matched cohort of 502 women and 436 men (n = 938). Outcome was estimated by adjusted binomial logistic regression analysis incorporating matched weights., Results: No effect of sex was seen to predict good outcome (OR 1.04, CI 0.76-1.43) or mortality (OR 1.13, CI 0.73-1.73). However, female sex was a strong independent predictor of symptomatic intracerebral hemorrhage (sICH - ECASS-II definition, OR 3.62, CI 1.77-7.41) and fatal ICH (OR 4.53, CI 1.61-12.7)., Conclusion: In balanced groups, the two sexes showed comparable outcomes following IVT. A novel finding was the higher rate of sICH and fatal ICH in women. In this analysis we also demonstrate how CEM can reduce multivariate imbalance and thereby improve estimates, already in crude, but more importantly, in adjusted regression analysis. Further investigations of multicentre data with improved analytical approaches that yield balanced sex-groups are therefore warranted.

We report the identification of deoxynivalenol-3-sulfate and deoxynivalenol-15-sulfate as two novel metabolites of the trichothecene mycotoxin deoxynivalenol in wheat. Wheat ears which were either artificially infected with Fusarium graminearum or directly treated with the major Fusarium toxin deoxynivalenol (DON) were sampled 96 h after treatment. Reference standards, which have been chemically synthesized and confirmed by NMR, were used to establish a liquid chromatography-electrospray ionization (LC-ESI)-MS/MS-based "dilute and shoot" method for the detection, unambiguous identification, and quantification of both sulfate conjugates in wheat extracts. Using this approach, detection limits of 0.003 mg/kg for deoxynivalenol-3-sulfate and 0.002 mg/kg for deoxynivalenol-15-sulfate were achieved. Matrix-matched calibration was used for the quantification of DON-sulfates in the investigated samples. In DON-treated samples, DON-3-sulfate was detected in the range of 0.29-1.4 mg/kg fresh weight while DON-15-sulfate concentrations were significantly lower (range 0.015-0.061 mg/kg fresh weight). In Fusarium-infected wheat samples, DON-3-sulfate was the only detected sulfate conjugate (range 0.022-0.059 mg/kg fresh weight). These results clearly demonstrate the potential of wheat to form sulfate conjugates of DON. In order to test whether sulfation is a detoxification reaction in planta, we determined the ability of the sulfated DON derivatives to inhibit in vitro protein synthesis of wheat ribosomes. The results demonstrate that both DON-sulfates can be regarded as detoxification products. DON-15-sulfate was about 44× less inhibitory than the native toxin, and no toxicity was observed for DON-3-sulfate in the tested range.

Deoxynivalenol (DON) is a trichothecene mycotoxin regularly occurring in cereals. Rats are often used to study toxicokinetics of DON and related compounds, yet only about 30 % of the administered dose is typically recovered. Recently, it was reported that DON is partly metabolised to previously undetected DON- and deepoxy-DON (DOM) sulfonate in rats and tentative structures were proposed. The present work describes the production and characterisation of DON-, DOM- and DON-3-glucoside (D3G) sulfonates of three different series; the development and validation of liquid chromatography tandem mass spectrometry (LC-MS/MS)-based methods for determination of DON, DOM, D3G and their sulfonates in rat faeces and urine; and application of the methods to samples from a DON and D3G feeding trial with rats. In addition to previously produced DON sulfonates (DONS) 1, 2 and 3, D3G sulfonates 1, 2 and 3; and DOM sulfonates (DOMS) 2 and 3 were synthesised, purified and characterised. The developed methods showed apparent recoveries of all investigated compounds between 68 and 151 % in faeces and between 48 and 113 % in urine. The recovery of DON, D3G and their metabolites from faeces and urine of rats (n = 6) administered in a single dose of 2.0 mg/kg b.w. DON or the equimolar amount of D3G was 75 ± 9 % for the DON group and 68 ± 8 % for the D3G group. DON-, DOM- and D3G sulfonates excreted in faeces accounted for 48 and 47 % of the total amount of administered DON and D3G. Urinary excretion of sulfonates was <1 %. In both treatment groups, DONS 2 was the major metabolite 0-24 h after treatment, whereas DOMS 2 was predominant thereafter. The developed methods can also be used for investigation of DON (conjugate) sulfonate formation in other animal species.

Introduction: Induction methods for therapeutic cooling are under investigated. We compared the effectiveness and safety of cold infusions (CI) and nasopharyngeal cooling (NPC) for cooling induction in stroke patients., Methods: A prospective, open-label, randomised (1:1), single-centre pilot trial with partially blinded safety endpoint assessment was conducted at the neurointensive care unit of Heidelberg University. Intubated stroke patients with an indication for therapeutic cooling and an intracranial pressure (ICP)/temperature brain probe were randomly assigned to CI (4°C, 2L at 4L/h) or NPC (60L/min for 1 h). Previous data suggested a maximum decrease of tympanic temperature for CI (2.1L within 35 min) after 52 min. Therefore the study period was 1 hour (15 min subperiods I-IV). The brain temperature course was the primary endpoint. Secondary measures included continuous monitoring of neurovital parameters and extracerebral temperatures. Statistical analysis based on repeated-measures analysis of variance., Results: Of 221 patients screened, 20 were randomized within 5 months. Infusion time of 2L CI was 33 ± 4 min in 10 patients and 10 patients received NPC for 60 min. During active treatment (first 30 min), brain temperature decreased faster with CI than during NPC (I: -0.31 ± 0.2 versus -0.12 ± 0.1°C, P = 0.008; II: -1.0 ± 0.3 versus -0.49 ± 0.3°C, P = 0.001). In the CI-group, after the infusion was finished, the intervention no longer decreased brain temperature, which increased after 3.5 ± 3.3 min. Oesophageal temperature correlated best with brain temperature during CI and NPC. Tympanic temperature reacted similarly to relative changes of brain temperature during CI, but absolute values slightly differed. CI provoked three severe adverse events during subperiods II-IV (two systolic arterial pressure (SAP), one shivering) compared with four in the NPC-group, all during subperiod I (three SAP, one ICP). Classified as possibly intervention-related, two cases of ventilator failure occurred during NPC., Conclusions: In intubated stroke patients, brain cooling is faster during CI than during NPC. Importantly, contrary to previous expectations, brain cooling stopped soon after CI cessation. Oesophageal but neither bladder nor rectal temperature is suited as surrogate for brain temperature during CI and NPC. Several severe adverse events in CI and in NPC demand further studying of safety., Trial Registration: ClinicalTrials.gov NCT01573117. Registered 31 March 2012.

Merkel cell carcinoma (MCC) is an aggressive, virus-associated, neuroendocrine tumor of the skin mainly affecting immunocompromised patients. Higher intratumoral infiltration with CD3 and CD8 positive T-cells is associated with a better prognosis, highlighting the relevance of the immune system for MCC development and progression. In this study 21 primary MCCs were stained with immune cell markers including CD3, CD4, CD8, CD68, CD20, and S100. Furthermore, tumor-infiltrating neutrophils, tertiary lymphoid structures and PD-L1 expression were analyzed and correlated with overall and recurrence free survival. All MCCs were Merkel Cell Polyomavirus positive. Overall and recurrence-free survival did not correlate with intra- and peritumoral CD3 and CD8 T-cell infiltration. In addition, no significant association regarding prognosis was found for tumor-associated neutrophils, tumor-associated macrophages or PD-L1 positivity in MCCs. Interestingly, the presence of tertiary lymphoid structures (TLS) in the tumor microenvironment significantly correlated with recurrence-free survival (P=0.025). In addition, TLS were significantly associated with a higher CD8/CD4 ratio in the tumor periphery (P=0.032), but not in the center of the tumor (P > 0.999). These results demonstrate for the first time that TLS, easily assessed in paraffin-embedded tissue in the tumor periphery of MCCs, may be a valuable prognostic factor indicating prolonged recurrence free survival.

The development of a reliable procedure for the synthesis of the 16-glucoside and 16-sulfate of the resorcylic acid lactone (RAL) type compound zearalenone is presented. Different protective group strategies were considered and applied to enable the preparation of glucosides and sulfates that are difficult to access up to now. Acetyl and p-methoxybenzyl protection led to undesired results and were shown to be inappropriate. Finally, triisopropylsilyl-protected zearalenone was successfully used as intermediate for the first synthesis of the corresponding mycotoxin glucoside and sulfate that are highly valuable as reference materials for further studies in the emerging field of masked mycotoxins. Furthermore, high stability was observed for aryl sulfates prepared as tetrabutylammonium salts. Overall, these findings should be applicable for the synthesis of similar RAL type and natural product conjugates.

Background and Purpose: Basilar artery occlusion remains one of the most devastating subtypes of ischemic stroke. The prognosis is poor if early recanalization is not achieved. The purpose of this study was to evaluate the safety and technical feasibility of self-expanding retrievable stents in the endovascular treatment of acute basilar artery occlusion., Materials and Methods: Twenty-four patients with acute basilar artery occlusion were treated with Solitaire FR or Revive SE devices between December 2009 and May 2012. Additional treatment included intravenous and/or intra-arterial thrombolysis (21/24) and percutaneous transluminal angioplasty/permanent stent placement (7/24). Recanalization was assessed by means of the TICI score. Clinical outcome was determined at discharge (NIHSS), and at 3 months (mRS)., Results: Median NIHSS score on admission was 24; median duration of symptoms was 254 minutes. Successful recanalization (TICI 2b +3) by thrombectomy only was achieved in 18 patients (75%). Intracranial stent deployment after thrombectomy caused by underlying atherosclerotic stenosis was performed in 7 patients. If these patients with intracranial stent placement are included, successful recanalization was achieved in 21 of 24 patients (87.5%). NIHSS improvement ≥10 points was reached in 54% of patients (n = 13/24). Mortality during the first 3 months was 29% (7/24). After 3 months, 8 patients (33%) had a favorable clinical outcome (mRS 0-2)., Conclusions: In our series, application of self-expanding retrievable stents in acute basilar artery occlusion resulted in a high recanalization rate without procedural complications and good clinical outcome in one-third of patients., (© 2014 by American Journal of Neuroradiology.)

The synthesis of 3-acyltetramic acids by C-acylation of pyrrolidine-2,4-diones was studied by density functional theory (DFT). DFT was applied to the mycotoxin tenuazonic acid (TeA), an important representative of these bioactive natural compounds. Lewis acid mediated C-acylation in combination with previous pH-neutral domino N-acylation-Wittig cyclization can be used for the efficient preparation of 3-acyltetramic acids. Nevertheless, quite harsh conditions are still required to carry out this synthetic step, leading to unwanted isomerization of stereogenic centers in some cases. In the presented study, the reaction pathway for the C-acetylation of (5S,6S-5-s-butylpyrrolidine-2,4-dione was studied in terms of mechanism, solvent effects, and Lewis acid activation, in order to obtain an appropriate theoretical model for further investigations. Crucial steps were identified that showed rather high activation barriers and rationalized previously reported experimental discoveries. After in silico optimization, aluminum chlorides were found to be promising Lewis acids that promote the C-acylation of pyrrolidine-2,4-diones, whereas calculations performed in various organic solvents showed that the solvent had only a minor effect on the energy profiles of the considered mechanisms. This clearly indicates that further synthetic studies should focus on the Lewis-acidic mediator rather than other reaction parameters. Additionally, given the results obtained for different reaction routes, the stereochemistry of this C-acylation is discussed. It is assumed that the formation of Z-configured TeA is favored, in good agreement with our previous studies.

Background and Purpose: In past years, thrombectomy has become a widely used procedure in interventional neuroradiology for the treatment of acute intracranial occlusions. However, in 10-20% of patients, there are additional occlusions or stenotic lesions of the ipsilateral cervical internal carotid artery. The purpose of this study was to evaluate the feasibility of emergency carotid artery stent placement in combination with intracranial thrombectomy and the clinical outcome of the treated patients., Materials and Methods: We analyzed clinical and angiographic data of patients who underwent emergency cervical ICA stent placement and intracranial thrombectomy with stent-retriever devices in our institution between November 2009 and July 2012. Recanalization was assessed according to the Thrombolysis in Cerebral-Infarction score. Clinical outcome was evaluated at discharge (NIHSS) and after 3 months (mRS)., Results: Overall, 24 patients were treated. The mean age was 67.2 years; mean occlusion time, 230.2 minutes. On admission, the median NIHSS score was 18. In all patients, the Thrombolysis in Cerebral Infarction score was zero before the procedure. Stent implantation was feasible in all cases. In 15 patients (62.5%), a Thrombolysis in Cerebral Infarction score ≥ 2b could be achieved. Six patients (25%) improved ≥10 NIHSS points between admission and discharge. After 90 days, the median mRS score was 3.0. Seven patients (29.2%) had a good clinical outcome (mRS 0-2), and 4 patients (16.6%) died, 1 due to fatal intracranial hemorrhage. Overall, symptomatic intracranial hemorrhage occurred in 4 patients (16.6%)., Conclusions: Emergency ICA stent implantation was technically feasible in all patients, and the intracranial recanalization Thrombolysis in Cerebral Infarction score of ≥2b was reached in a high number of patients. Clinical outcome and mortality seem to be acceptable for a cohort with severe stroke. However, a high rate of symptomatic intracranial hemorrhage occurred in our study.

The trichothecene mycotoxin deoxynivalenol (DON) is a well known and common contaminant in food and feed. Acetylated derivatives and other biosynthetic precursors can occur together with the main toxin. A key biosynthetic step towards DON involves an oxidation of the 8-OH group of 7,8-dihydroxycalonectrin. Since analytical standards for the intermediates are not available and these intermediates are therefore rarely studied, we aimed for a synthetic method to invert this reaction, making a series of calonectrin-derived precursors accessible. We did this by developing an efficient protocol for stereoselective Luche reduction at C8. This method was used to access 3,7,8,15-tetrahydroxyscirpene, 3-deacetyl-7,8-dihydroxycalonectrin, 15-deacetyl-7,8-dihydroxycalonectrin and 7,8-dihydroxycalonectrin, which were characterized using several NMR techniques. Beside the development of a method which could basically be used for all type B trichothecenes, we opened a synthetic route towards different acetylated calonectrins.

The RAB5 GTPase ARA6 (AtARA6) of Arabidopsis thaliana is known to be involved in endosomal trafficking by targeting vesicles to the plasma membrane. During this process AtARA6 is working in close relationship with the SNARE protein VAMP727 (vesicle associated membrane protein 727). Recently, ARA6 of the characean green algae Chara australis (CaARA6) was shown to have properties similar to AtARA6, pointing to similar trafficking pathways. In order to gain further insight into the vesicle trafficking machinery of characeae, C. australis was analyzed for homologous proteins of the VAMP72-family. A CaVAMP72 protein was detected and classified by protein sequence alignment and phylogenetic analyses.