Our aim was to update the evidence-based and consensus-based recommendations for prehospital coagulation management and fluid replacement therapy in patients with multiple and/or severe injuries on the basis of current evidence. This guideline topic is part of the 2022 update of the German Guideline on the Treatment of Patients with Multiple and/or Severe Injuries. MEDLINE and Embase were systematically searched to May 2021. Further literature reports were obtained from clinical experts. Randomised controlled trials, prospective cohort studies, and comparative registry studies were included if they compared interventions for fluid replacement therapy, the transfusion of blood products, the management of coagulation, or intravenous/intraosseous access in patients with multiple and/or severe injuries in the prehospital setting. We considered patient-relevant clinical outcomes such as mortality and bleeding control, or coagulation parameters as surrogate outcomes. Risk of bias was assessed using NICE 2012 checklists. The evidence was synthesised narratively, and expert consensus was used to develop recommendations and determine their strength. Thirty-five new studies were identified. Interventions covered were prehospital fluid replacement therapy (n = 5 studies), infusions (n = 3), transfusions (n = 11), coagulation management (n = 13), and intraosseous access (n = 2). Four recommendations were modified, and six additional recommendations were developed. All achieved strong consensus. The following key recommendations are made. Fluid replacement therapy should be initiated in severely injured patients. In patients with uncontrolled bleeding, fluid replacement therapy should be limited (MAP of 65 mmHg, SBP of 80 mmHg) in order to maintain minimum haemodynamic stability while not increasing blood loss. In hypotensive patients with suspected isolated or concomitant significant traumatic brain injury, the objective of fluid replacement should be to maintain normal blood pressure (MAP of 85 mmHg, SBP of 110 mmHg). Intravenous access is used in trauma patients. If intravenous access cannot be achieved in trauma patients, intraosseous access is used for the delivery of fluids and medications. If there are no signs and symptoms of volume depletion, fluid replacement therapy should not be provided. If the administration of a sufficient volume of fluids fails to achieve adequate blood pressure in a polytrauma patient, the titrated use of vasopressors for circulatory support may be considered. The lethal triad of hypothermia, acidosis, and coagulopathy should be addressed at the prehospital stage. One gram of tranexamic acid should be administered in cases of existing or imminent haemorrhagic shock. The administration of fibrinogen may also be considered in cases of uncontrollable bleeding, as may the administration of erythrocyte and plasma concentrates.

Introduction:Parvovirus B19 transmitted by umbilical cord blood (UCB) products may cause severe disease in allogenic hematopoietic stem cell transplant recipients. Thus, commercially available nucleic acid test (NAT) assays for highly sensitive detection of parvovirus B19 DNA validated for the specimen cord blood plasma (CBP) are required to avoid parvovirus B19 transmission by umbilical hematopoietic stem cell preparations. Methods:The multiplex cobas DPX NAT assay was validated for detection of parvovirus B19 DNA in CBP derived from citrate anticoagulated UCB units which have been processed by the Rubinstein method. In total, 363 retained CBP samples pretested negative for parvovirus B19 DNA were prepared for analyzing sensitivity, specificity, and interference of that NAT assay. The 3rd WHO International Standard for parvovirus B19 DNA was used for determining the 95% limit of detection (LOD95) by probit analysis. Results:The validation of the parvovirus B19 NAT assay for CBP demonstrated high sensitivity, specificity, intra- and inter-assay precision. Dilution series and replicate analyses showed a high linearity of the assay with a coefficient of determination above 0.99 and revealed a LOD95 of 17 International Units (IU)/mL (95% confidence interval, 14-44 IU/mL) for parvovirus B19 DNA in CBP samples. Conclusion:The validation of a commercially available parvovirus B19 NAT assay for the specimen CBP demonstrated a high assay performance fulfilling German guidelines and international regulations.

Patients with the chronic need of transfusions, such as patients with sickle cell disease, have a higher risk of alloimmunization, for example patients with Duffy Null Fy(a- b-) Genotype. In the case of alloantibody formation, the supply of compatible red cell units for patients is more difficult. In this regard, anti-Fy-3 antibody is an antibody formed in individuals with Duffy null genotype (Fya-, Fyb-). This alloantibody can bind both Duffy A and Duffy B antigens. The Duffy null phenotype, Fy(a-b-), is very rare in Caucasians but occurs in 68% of blacks [1].Our patient is of African descent and has the allele FY*02 N.01 (Duffy Null). During a sickle cell crisis, a positive crossmatch as well as alloantibodies, anti-M and anti-Jk (b)) were present. The anti-M titer was 64 and the anti-Jk(b) titer was 32. Genotyping revealed the Duffy null genotype. Furthermore, genotyping showed that the patient was M-negative and Jk(b)-negative, which supported the diagnosis of anti-M and anti-Jk(b) antibodies. The patient received a transfusion of antigen-compatible red cells, hemoglobin levels increased, and her condition improved. No further signs of hemolysis occurred. Additional diagnostics confirmed the suspected alloantibodies anti-M- and anti-Jk(b). In addition, an anti-Fy-3 antibody was suspected.This case highlights that genotyping of blood group antigens can provide additional information to serological findings. This information supports the prevention of alloimmunization in frequently transfused patients.

ZusammenfasssungPatienten, die im Kontext einer Erythrozytenbildungsstörung wie z. B. einer Sichelzellerkrankung chronisch substitutionsbedürftig sind, haben ein erhöhtes Risiko einer Alloimmunisierung, z. B. bei Patienten mit dem Genotyp Duffy null Fy(a- b-). Im Falle der Bildung von Alloantikörpern ist die Versorgung der Patienten mit kompatiblen Erythrozytenkonzentraten erschwert. Anti-Fy-3-Antikörper können bei Personen mit dem Genoytp Duffy null Fy(a-, b-) gebildet werden. Diese Antikörper können sowohl Duffy A als auch Duffy B Antigene binden. Der Duffy-Null-Phänotyp, Fy (a-b-), ist bei Kaukasiern sehr selten, kommt aber bei 68% der Patienten mit afrikanischer Abstammung vor [1].Die von uns betreute Patientin ist afrikanischer Abstammung und weist genotypisch das Allel FY*02 N.01 (Duffy null) auf. Im Rahmen einer Sichelzellkrise und geplanten Bluttransfusionen fielen eine positive Kreuzprobe sowie nachweisbare Alloantikörper (Verdacht auf Anti-M, Anti-Jk(b)) auf. Der anti-M-Titer betrug 64 und der anti-Jk(b)-Titer betrug 32. Die Genotypisierung ergab den Duffy-Null-Genotyp Fy*02 N.01 (SSP inno-Train kits KKD SKP0121, MNS SMN1120, Rare ID SSP0620). Ferner zeigte sich in der Genotypisierung, dass die Patientin M-negativ und Jk(b)-negativ war, was die Diagnose von anti-M- und anti-Jk(b)-Antikörpern bestärkte. Die Patientin erhielt eine Transfusion mit Antigen-kompatiblen Erythrozytenkonzentraten. Der Hämoglobinwert stieg an und ihr klinischer Zustand verbesserte sich deutlich. Anzeichen einer Hämolyse traten nicht auf. Die ergänzende serologische Diagnostik bestätigte die vermuteten Alloantikörper anti-M- und anti-Jk(b). Aufgrund dessen wurde ein zusätzlicher anti-Fy-3-Antikörper vermutet.Dieser Fall zeigt die Bedeutung einer Genotypisierung ergänzend zu den serologischen Befunden. Diese Information unterstützt bei häufig transfundierten Patienten die Vermeidung einer Alloimmunisierung.

Background: Therapeutic plasma exchange (TPE) is a well-known apheresis technology since many years and is available worldwide. Myasthenia gravis is one of the first neurological diseases successfully treated with TPE. TPE is also frequently applied in acute inflammatory demyelinating polyradiculoneuropathy (Guillain-Barré syndrome). Both neurological disorders are immunologically mediated and might cause life-threatening symptoms in patients. Summary: There is a large body of evidence from many randomized controlled trials (RCTs) that the application of TPE in myasthenia gravis crisis or in acute Guillain-Barré syndrome is effective and safe. Thus, TPE is recommended as first-line therapy with a grade 1A recommendation during the critical course of these neurological diseases. Even chronic inflammatory demyelinating polyneuropathies characterized by complement-fixing autoantibodies to myelin are successfully treated with TPE. The plasma exchange reduces inflammatory cytokines, complements activating antibodies, and leads to an improvement of neurological symptoms. TPE is no standalone treatment but often combined with immunosuppressive therapy. Recent studies (clinical trials, retrospective analysis, meta-analysis, and systematic reviews) evaluate special apheresis technology (i.e., immunoadsorption [IA], small volume plasma exchange), compare different treatments of these neuropathies, or report on the therapy of rare immune-mediated neuropathies in case reports. Key Messages: TA is a well-established treatment and is safe in acute progressive neuropathies (myasthenia gravis, Guillain-Barré syndrome) with an immune etiology. TPE has been applied for decades and thus has the best evidence so far. The indication for IA depends on the availability of that technology and the evidence by RCTs in special neurological diseases. The treatment with TA should improve the clinical outcome of patients, reducing acute or chronic (chronic inflammatory demyelinating polyneuropathies) neurological symptoms. The informed consent of the patient should carefully weight risks and benefits of the apheresis treatment and consider alternative therapies.

The incidence of head and neck squamous cell carcinomas (HNSCC) is increasing worldwide, especially when triggered by the human papilloma virus (HPV). Radiotherapy has immune-modulatory properties, but the role of macrophages present in HNSCC and having contact with irradiated tumor cells remains unclear. The influence of irradiated (2 × 5Gy) HNSCC cells on the (re-)polarization and phagocytosis of human macrophages, either non-polarized or with a more M1 or M2 phenotype, was therefore investigated. Human monocytes were differentiated with the hematopoietic growth factors M‑CSF (m) or GM-CSF (g) and additionally pre-polarized with either interleukin (IL)-4 and IL-10 or interferon (IFN)-γ and lipopolysaccharides (LPS), respectively. Subsequently, they were added to previously irradiated (2 × 5Gy) and mock-treated HPV-positive (UD-SCC-2) and HPV-negative (Cal33) HNSCC cells including their supernatants. The HNSCC cells treated with hypofractionated irradiation died via apoptosis and were strongly phagocytosed by M0m and M2 macrophages. M0g and M1 macrophages phagocytosed the tumor cells to a lesser extent. Irradiated HNSCC cells were better phagocytosed by M1 macrophages compared to mock-treated controls. The polarization status of the macrophages was not significantly changed, except for the expression of CD206 on M2 macrophages, which was reduced after phagocytosis of irradiated HPV-negative cells. Further, a significant increase in the uptake of irradiated HPV-positive cells by M0g macrophages when compared to HPV-negative cells was observed. HNSCC cells treated with hypofractionated irradiation foster phagocytosis by anti-tumorigenic M1 macrophages. The data provide the first evidence on the impact of the HPV status of HNSCC cells on the modulation of the macrophage response to irradiated tumor cells.

Cystic fibrosis patients suffer from a progressive, often fatal lung disease, which is based on a complex interplay between chronic infections, locally accumulating immune cells and pulmonary tissue remodeling. Although group-2 innate lymphoid cells (ILC2s) act as crucial initiators of lung inflammation, our understanding of their involvement in the pathogenesis of cystic fibrosis remains incomplete. Here we report a marked decrease of circulating CCR6+ ILC2s in the blood of cystic fibrosis patients, which significantly correlated with high disease severity and advanced pulmonary failure, strongly implicating increased ILC2 homing from the peripheral blood to the chronically inflamed lung tissue in cystic fibrosis patients. On a functional level, the CCR6 ligand CCL20 was identified as potent promoter of lung-directed ILC2 migration upon inflammatory conditions in vitro and in vivo using a new humanized mouse model with light-sheet fluorescence microscopic visualization of lung-accumulated human ILC2s. In the lung, blood-derived human ILC2s were able to augment local eosinophil and neutrophil accumulation and induced a marked upregulation of pulmonary type-VI collagen expression. Studies in primary human lung fibroblasts additionally revealed ILC2-derived IL-4 and IL-13 as important mediators of this type-VI collagen-inducing effect. Taken together, the here acquired results suggest that pathologically increased CCL20 levels in cystic fibrosis airways induce CCR6-mediated lung homing of circulating human ILC2s. Subsequent ILC2 activation then triggers local production of type-VI collagen and might thereby drive extracellular matrix remodeling potentially influencing pulmonary tissue destruction in cystic fibrosis patients. Thus, modulating the lung homing capacity of circulating ILC2s and their local effector functions opens new therapeutic avenues for cystic fibrosis treatment.

BACKGROUND:Applicability of thrombin generation tests to clinical routine has been sought for many years. The aim of this study was to compare thrombin generation measured in fresh platelet poor plasma (f-PPP) and frozen-thawed platelet poor plasma (ft-PPP) to prove the consistency of results.METHODS:In this prospective study, thrombin generation was measured in twenty-fold repetitions in 3.2% citrate PPP obtained from male healthy blood donors aged 19 - 39 years (n = 54 donations). The tests were performed with fresh PPP and repeated after storing the PPP at -60°C. In two subgroup analyses, the effect of higher and lower normal baseline platelet counts on the Calibrated Automated Thrombogram (CAT) assay and the influence of ABO blood groups on thrombin generation were analyzed.RESULTS:Referring to the parameters of thrombin generation most frequently used in studies, peak thrombin of f-PPP and ft-PPP agreed in about 50% of the samples. Endogenous thrombin potential (ETP) of f-PPP and ft-PPP agreed in nearly two-thirds of the samples. A slightly but significantly slower kinetic was found in the thrombin generation of ft-PPP compared with f-PPP. At least in f-PPP, ETP correlates with baseline platelet counts of the whole blood sample. Peak thrombin was significantly higher in non-O blood groups compared to O blood group.CONCLUSIONS:A low level of agreement between the results of f-PPP and ft-PPP is shown. In terms of practicability of sample collection using semi-automated thrombin-generation assays ft-PPP should be preferred over f-PPP. We therefore recommend using ft-PPP in clinical studies.

Background Oral Factor Xa inhibitors for the prevention of stroke in atrial fibrillation require dose adjustment based on certain clinical criteria, but the off-label use of the reduced doses is common. Methods Data from an observational registry including patients admitted with acute cerebral ischemia while taking oral Factor Xa inhibitors for atrial fibrillation between April 2016 and December 2018 were investigated. The dose regimen of the Xa inhibitor was classified as “appropriate”, “underdosed” and “overdosed” in conformity with the European Medicines Agency labelling. The effect of underdosing on the functional factor Xa plasma level on admission, the clinical stroke severity and the functional outcome after 3 months were investigated. Results 254 patients with cerebral ischemia while on Factor Xa inhibitors were included. The dose regimen of the Factor Xa inhibitor was appropriate in 166 patients (65%), underdosed in 67 patients (26%) and overdosed in 21 patients (8%). Underdosing was associated with female sex, diabetes mellitus and higher CHA 2 DS 2 –Vasc scores. Underdosing independently predicted lower anti-Xa plasma levels on admission [median 69.4 ng/ml (IQR 0.0–121.6) vs. 129.2 ng/ml (65.5–207.2); p < 0.001], was associated with higher NIHSS scores on admission [median 5 (IQR 1–10) vs. 3 (1–7); p = 0.041] and worse functional outcome after 3 months (favorable outcome 26.9% vs. 46.9%; p = 0.025). Conclusion One in three patients with ischemic stroke during treatment with oral Xa inhibitors used inappropriate dose regimens. Underdosing was associated with lower functional plasma levels, higher clinical stroke severity and worse functional outcome.

BACKGROUNDTraditionally, white blood cells (WBCs) are collected from buffy coats or freshly drawn blood. However, the increasing demand for peripheral blood mononuclear cells (PBMCs) in the research phases of immunological therapy development makes it necessary to identify alternative sources of these cells.STUDY DESIGN AND METHODSLeukapheresis products are cost intensive and not offered by all blood banks. Therefore, thrombocyte apheresis cassettes (TACs), plateletpheresis waste products, were investigated as a possible low‐cost and easily accessible blood source for research laboratories. The recovery rate, phenotype, and functionality of WBC subsets from TAC are unknown and were investigated in comparison to frequently used blood resources via flow cytometry.RESULTSOn average, TACs provide 30.3 × 106/mL PBMCs, situating themselves between peripheral whole blood (WB; 5.35 × 106/mL) and leukoreduction system chamber (LRSC; 163.9 × 106/mL) yields. Frequencies of CD14, CD3, CD4, CD8, CD56, CD19, and CD11c positive cells in TACs correlate with normal proportions of WBC populations. Stimulation of TAC‐derived PBMCs by lipopolysaccharide (LPS) and resiquimod (R848) showed no significant differences in expression levels of human leukocyte antigen (HLA)‐DR, DQ, DP, and CD86 or cytokine secretion compared to other blood source derived PBMC. Following stimulation with LPS or R848, comparable levels of tumor necrosis factor‐α, interleukin‐10, and interleukin‐1β could be measured between TAC, LRSC, and WB. Additionally, TAC‐derived T cells retained their proliferation capability and were able to produce interferon‐γ following T‐cell receptor stimulation.CONCLUSIONTACs provide a cost‐effective source of viable and functional human blood cells that can readily be used for clinical and laboratory investigations after plateletpheresis preparation.

BACKGROUND:The frequency of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNAemia in blood donors is uncertain. Thus, assays for SARS-CoV-2 RNA detection in blood, validated on commercially available polymerase chain reaction (PCR) systems, are required to allow a good comparability of data.STUDY DESIGN AND METHODS:The cobas SARS-CoV-2 dual-target reverse transcriptase PCR (RT-PCR) assay, licensed for respiratory swab SARS-CoV-2 RNA testing, was validated for detection of viral RNA in blood. For the validation panel, SARS-CoV-2-positive plasma samples were prepared by spiking SARS-CoV-2-positive respiratory specimens in negative human plasma. The 95% limit of detection (LOD95) was determined by probit analysis. For clinical validation, coronavirus disease 2019 (COVID-19) convalescent plasma (CCP) donors and patients with COVID-19 with a severe disease course treated in an intensive care unit (ICU) were included.RESULTS:The validation of the SARS-CoV-2 RT-PCR assay for blood demonstrated high sensitivity and specificity and intra- and inter-assay precision and efficiency. The LOD95 for SARS-CoV-2 RNA was 5.0 genome copies/mL (95% confidence interval [CI], 3.3-12 copies/mL) for target 1 and 4.3 genome copies/mL (95% CI, 2.9-10 copies/mL) for target 2. In a cohort of 39 CCP donors with 66 CCP donations no SARS-CoV-2 RNA in plasma was detected. Screening of 25 blood samples of 19 ICU patients with COVID-19 showed six positive results for SARS-CoV-2 RNA in at least one target of the assay.CONCLUSION:The SARS-CoV-2 RNA assay, only licensed for respiratory swabs, performed on a PCR system for high-throughput testing, showed a good assay performance for blood testing.

SummaryThe blood count is one of the most common tests used for health assessment. In elderly individuals, selection of a ‘healthy’ reference population for laboratory assessment is difficult due to the high prevalence of chronic morbidities, leading to uncertainty regarding appropriate reference intervals. In particular, age‐specific lower haemoglobin reference limits to define anaemia are controversial. Here, we applied a data mining approach to a large dataset of 3 029 904 clinical routine samples to establish blood count reference intervals. We excluded samples from units/specialists with a high proportion of abnormal blood counts, samples from patients with an unknown or decreased estimated glomerular filtration rate, and samples with abnormal test results in selected other analytes. After sample exclusion, 566 775–572 060 samples from different individuals aged 20–100 years were available for analysis. We then used an established statistical algorithm to determine the distribution of physiological test results and calculated age‐ and sex‐specific reference intervals. Our results show substantial trends with age in haematology analytes' reference intervals. Most notably, haemoglobin and red cell counts decline in men with advanced age, accompanied by increases in red cell volume in both sexes. These findings were confirmed in an independent dataset, and suggest an at least partly physiologic cause.

ZusammenfassungIm vergangenen Jahrzehnt kam es in den meisten Industrieländern, auch in Deutschland, zu tiefgreifenden Veränderungen in der klinischen Praxis der Hämotherapie. Die Indikationsstellung zur Erythrozytentransfusion erfolgt immer zurückhaltender. Der Verbrauch von Erythrozytenkonzentraten ist stark rückläufig. Gleichzeitig steigt auch die Zahl der Patienten, bei denen trotz geplanter Operation oder Intervention die präoperative bzw. präinterventionelle Bereitstellung von gekreuzten Blutkonserven unterbleibt. Wir berichten über einen 71-jährigen multimorbiden Patienten mit dilatativer Kardiomyopathie, stark reduzierter kardialer Reserve, Z. n. wenige Wochen zurückliegender Trikuspidalklappenersatz-Operation, laufender Antikoagulation mit Heparin und diagnostizierter Anämie mit Hämoglobinwerten zwischen 7 und 8 g/dl. Bei diesem Patienten kam es nach einer bereits mehrtägigen stationären Behandlung unerwartet zu einem hämorrhagischen Schock infolge einer Forrest-IIb-Blutung aus einem Ulcus duodeni. Im Rahmen der Notfallversorgung stellte sich heraus, dass jede transfusionsvorbereitende Diagnostik versäumt worden war. Die notfallmäßig bereitgestellten, ungekreuzten Erythrozytenkonzentrate der Blutgruppe 0 Rhesus-negativ lösten bei diesem Patienten eine akute hämolytische Transfusionsreaktion aus. Grund war ein zum Transfusionszeitpunkt nicht identifizierter irregulärer Antikörper gegen das Blutgruppenmerkmal s aus dem MNSs-Blutgruppensystem. Akute hämolytische Transfusionsreaktionen außerhalb des AB0-Systems sind eine seltene, dennoch grundsätzlich immer drohende Komplikation der Transfusion ungekreuzter Erythrozytenkonzentrate im lebensbedrohlichen Blutungsnotfall. Der Fall zeigt, dass der Wunsch, eine restriktive Transfusionspraxis zu befolgen, nicht dazu führen darf, im Einzelfall Konstellationen zu übersehen, die selbstverständlich die rechtzeitige Vornahme einer prätransfusionellen Diagnostik mit Blutgruppenbestimmung, Antikörpersuche und Antikörperidentifizierung erfordern. Nach unserer Meinung nimmt die Frequenz vergleichbarer Ereignisse in einem Ausmaß zu, das es nahelegt, solche Fehler im klinischen Behandlungsgeschehen nicht nur in CIRS-Systemen zu bearbeiten, sondern sie auch in den Hämovigilanzsystemen zu erfassen.

The section "Preparative and Therapeutic Hemapheresis" of the German Society for Transfusion Medicine and Immunohematology (DGTI) has reviewed the actual literature and updated techniques and indications for evidence-based use of therapeutic apheresis in human disease. The recommendations are mostly in line with the "Guidelines on the Use of Therapeutic Apheresis in Clinical Practice" published by the Writing Committee of the American Society for Apheresis (ASFA) and have been conducted by experts from the DACH (Germany, Austria, Switzerland) region.

BACKGROUND:Appropriate reference intervals are essential when evaluating laboratory test results. However, establishment of reference intervals is challenging, especially for coagulation screening tests, and uncertainty exists regarding age- and sex-dependency of test results. Data mining of laboratory information systems is an emerging approach to reference interval determination, and we evaluated its applicability to coagulation tests.METHODS:We analyzed measurements of activated partial thromboplastin time (aPTT), prothrombin time (PT), international normalized ratio (INR), thrombin time (TT), and fibrinogen performed during clinical care in the University Hospital Erlangen, Germany (1,778,738 samples from 116,754 adult patients, 45,577-509,859 samples per analyte). We identified the proportion of samples from healthy individuals using an established statistical approach (Reference Limit Estimator), in which the distribution of physiological test results is approximated using a parametrical function, and used for the calculation of reference intervals.RESULTS:We established age- and sex specific reference intervals for aPTT, PT, INR, TT, and fibrinogen, and created batch- and reagent-specific aPTT-reference intervals. Additionally, we evaluated the sensitivity of the established aPTT reference intervals for the detection of factor VIII, IX, XI, XII deficiencies.CONCLUSION:Data mining of laboratory test results allows the creation of age- and sex-reference intervals for coagulation tests that are specific to the examined population, analytical framework, and reagent. This approach can complement conventional methods when establishing reference intervals and improve clinical decision-making based on coagulation tests. The reference intervals established in this study show only minor variation with sex and age, supporting the practice of providing a common reference interval for adult women and men.

Background: Early diagnosis of Alzheimer’s disease (AD) is challenging, and easily accessible biomarkers are an unmet need. Blood platelets frequently serve as peripheral model for studying AD pathogenesis and might represent a reasonable biomarker source. Objective: In the present study, we investigated the potential to differentiate AD patients from healthy controls (HC) based on blood count, platelet morphology, and function as well as molecular markers at the time of first clinical diagnosis. Methods: Blood samples from 40 AD patients and 29 age-matched HC were included for determination of 78 parameter by blood counting, platelet morphometry, aggregometry, flow cytometry (CD62P, CD63, activated fibrinogen receptor), protein quantification of nicotinic acetylcholine receptor α7 (nAChR α7) and caveolin-1 (CAV-1), and miRNA quantification (miR-26b, miR-199a, miR-335). Group comparison between patients and controls was performed in univariate and multivariate statistical analyses. Results: AD patients showed significantly lower aggregation response to ADP and arachidonic acid and significantly decreased CD62P and CD63 surface expression induced by ADP and U46619 compared to HC. Relative nAChR α7 and CAV-1 expression was significantly higher AD platelets than in HC. Multivariate analysis of 63 parameter revealed significant differences between AD patients and healthy controls. The best performing feature model revealed a sensitivity of 96.6%, a specificity of 80.0%, and a positive predictive value of 89.3%. No grouping could be achieved by using single parameter groups. Conclusion: Significant differences between platelet characteristics from AD patients and HC at the time of first clinical diagnosis were observed. The best performing parameter can be used as a blood-based biomarker for AD diagnosis in a multivariate model in addition to the standardized mental tests.

Background and Purpose-In patients with ischemic stroke on therapy with vitamin K antagonists, stroke severity and clinical course are affected by the quality of anticoagulation at the time of stroke onset, but clinical data for patients using direct oral anticoagulants (DOACs) are limited. Methods-Data from our registry including all patients admitted with acute cerebral ischemia while taking oral anticoagulants for atrial fibrillation between November 2014 and October 2017 were investigated. The activity of vitamin K antagonists was assessed using the international normalized ratio on admission and categorized according to a threshold of 1.7. DOAC plasma levels were measured using the calibrated Xa-activity (apixaban, rivaroxaban, and edoxaban) or the Hemoclotassay (dabigatran) and categorized into low (<50 ng/mL), intermediate (50-100 ng/mL), or high (>100 ng/mL). Primary objective was the association between anticoagulant activity and clinical and imaging characteristics. Results-Four hundred sixty patients were included (49% on vitamin K antagonists and 51% on DOAC). Patients on vitamin K antagonists with low international normalized ratio values had higher scores on the National Institutes of Health Stroke Scale and a higher risk of large vessel occlusion on admission. For patients on DOAC, plasma levels were available in 75.6% and found to be low in 49 (27.7%), intermediate in 41 (23.2%), and high in 87 patients (49.2%). Low plasma levels were associated with higher National Institutes of Health Stroke Scale scores on admission (low: 8 [interquartile range, 3-15] versus intermediate: 4 [1-11] versus high: 3 [0-8]; P<0.001) and higher risk of persisting neurological deficits or cerebral infarction on imaging (85.7% versus 75.6% versus 54.0%; P<0.001). Low DOAC plasma levels were an independent predictor of large vessel occlusion (odds ratio, 3.84 [95% CI, 1.80-8.20]; P=0.001). Conclusions-The activity of anticoagulation measured by specific DOAC plasma levels on admission is associated with stroke severity and presence of large vessel occlusion.