Value of cervical electrical impedance spectroscopy to predict spontaneous preterm delivery in asymptomatic women: the ECCLIPPx prospective cohort study.

OBJECTIVES: Preterm birth (PTB) accounts for two-thirds of deaths of structurally normal babies and is associated with substantial lifetime healthcare costs. Prevention of PTB remains limited by the modest accuracy of prediction methods, namely transvaginal ultrasound (TVS) cervical length (CL) measurement and quantitative cervicovaginal fetal fibronectin (FFN) estimation. We report the first substantive study detailing the predictive performance of a cervical probe device based on electrical impedance spectroscopy (EIS) for PTB - the EleCtriCaL Impedance Prediction of Preterm birth by spectroscopy of the cervix (ECCLIPPx) study. We aimed to compare the accuracy of cervical EIS-based prediction of spontaneous PTB with that of prediction using TVS-CL and FFN in asymptomatic women in the mid-trimester. METHODS: We studied asymptomatic women with a singleton pregnancy at 20-22 weeks' and 26-28 weeks' gestation. EIS was performed using a Sheffield Mark 5.0 device that makes measurements in the frequency range 76 Hz to 625 kHz using a small probe housing tetrapolar electrodes. TVS-CL and FFN were also measured. The associations of cervical EIS, TVS-CL and FFN with spontaneous delivery before 37 weeks and before 32 weeks were determined by multivariate linear and non-linear logistic regression analysis. Areas under the receiver-operating-characteristics curves (AUC) plots of sensitivity against specificity were used to compare the predictive performance of all parameters, both in isolation and in combination. RESULTS: Of the 365 asymptomatic women studied at 20-22 weeks who were not receiving treatment, 29 had spontaneous PTB, 14 had indicated PTB and 322 had term birth. At the higher frequencies assessed, cervical EIS predicted spontaneous PTB before 37 weeks with an AUC of 0.76 (95% CI, 0.71-0.81), compared with AUCs of 0.72 (95% CI, 0.66-0.76) for TVS-CL and 0.62 (95% CI, 0.56-0.72) for FFN. Combining all three assessments improved the prediction of spontaneous PTB before 37 weeks (AUC, 0.79 (95% CI, 0.74-0.83)) compared with TVS-CL and FFN alone. Incorporating a history of spontaneous PTB (defined as previous mid-trimester miscarriage or spontaneous PTB (14 to < 37 weeks)) into the cervical EIS prediction model improved the accuracy of prediction of spontaneous PTB before 37 weeks (AUC, 0.83 (95% CI, 0.78-0.87)) and before 32 weeks (AUC, 0.86 (95% CI, 0.82-0.90)). CONCLUSIONS: Mid-trimester cervical EIS assessment predicts spontaneous PTB. Larger confirmatory studies investigating its potential clinical utility and to inform effective preventive interventions are required. © 2020 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

The 2020 Canadian Cardiovascular Society/Canadian Heart Rhythm Society Comprehensive guidelines for the management of Atrial Fibrillation

The Canadian Cardiovascular Society (CCS) atrial fibrillation (AF) guidelines program was developed to aid clinicians in the management of these complex patients, as well as to provide direction to policy makers and health care systems regarding related issues. The most recent comprehensive CCS AF guidelines update was published in 2010. Since then, periodic updates were published dealing with rapidly changing areas. However, since 2010 a large number of developments had accumulated in a wide range of areas, motivating the committee to complete a thorough guideline review. The 2020 iteration of the CCS AF guidelines represents a comprehensive renewal that integrates, updates, and replaces the past decade of guidelines, recommendations, and practical tips. It is intended to be used by practicing clinicians across all disciplines who care for patients with AF. The Grading of Recommendations, Assessment, Development and Evaluations (GRADE) system was used to evaluate recommendation strength and the quality of evidence. Areas of focus include: AF classification and definitions, epidemiology, pathophysiology, clinical evaluation, screening and opportunistic AF detection, detection and management of modifiable risk factors, integrated approach to AF management, stroke prevention, arrhythmia management, sex differences, and AF in special populations. Extensive use is made of tables and figures to synthesize important material and present key concepts. This document should be an important aid for knowledge translation and a tool to help improve clinical management of this important and challenging arrhythmia.

Le programme de lignes directrices de la Société canadienne de cardiologie (SCC) en matière de fibrillation auriculaire (FA) a été élaboré pour aider les cliniciens à prendre en charge ces patients complexes, ainsi que pour orienter les décideurs politiques et les systèmes de soins de santé sur des questions connexes. La dernière édition complète des lignes directrices de la SCC en matière de FA a été publiée en 2010. Depuis lors, des mises à jour périodiques ont été publiées, traitant de domaines en évolution rapide. Cependant, en 2020, un grand nombre de développements s'y étaient ajoutés, couvrant un large éventail de domaines, ce qui a motivé le comité à créer une refonte complète des lignes directrices. L'édition 2020 des lignes directrices de la SCC en matière de FA représente un renouvellement complet qui intègre, met à jour et remplace les lignes directrices, les recommandations et les conseils pratiques des dix dernières années. Elle est destinée à être utilisée par les cliniciens praticiens de toutes les disciplines qui s'occupent de patients souffrant de FA. L'approche GRADE (Gradation des Recommandations, de l'Appréciation, du Développement et des Évaluations) a été utilisée pour évaluer la pertinence des recommandations et la qualité des résultats. Les domaines d'intérêt incluent : la classification et les définitions de la FA, son épidémiologie, sa physiopathologie, l'évaluation clinique, le dépistage de la FA, la détection et la gestion des facteurs de risque modifiables, l'approche intégrée de la gestion de la FA, la prévention des accidents vasculaires cérébraux, la gestion de l'arythmie, les différences entre les sexes et la FA dans des populations particulières. Des tableaux et figures ont été largement utilisés pour synthétiser les éléments importants et présenter les concepts clés. Ce document devrait représenter une aide importante pour l'intégration des connaissances et un outil pour aider à améliorer la gestion clinique de cette arythmie importante et difficile à traiter.

Surgical Outcome and Oncological Survival of Osteofibrous Dysplasia-Like and Classic Adamantinomas: An International Multicenter Study of 318 Cases.

BACKGROUND: Osteofibrous dysplasia-like adamantinoma (OFD-AD) and classic adamantinoma (AD) are rare, neoplastic diseases with only limited data supporting current treatment protocols. We believe that our retrospective multicenter cohort study is the largest analysis of patients with adamantinoma to date. The primary purpose of this study was to describe the disease characteristics and evaluate the oncological outcomes. The secondary purpose was to identify risk factors for local recurrence after surgical treatment and propose treatment guidelines. METHODS: Three hundred and eighteen confirmed cases of OFD-AD and AD for which primary treatment was carried out between 1985 and 2015 were submitted by 22 tertiary bone tumor centers. Proposed clinical risk factors for local recurrence such as size, type, and margins were analyzed using univariable and multivariate Cox regression analysis. RESULTS: Of the 318 cases, 128 were OFD-AD and 190 were AD. The mean age at diagnosis was 17 years (median, 14.5 years) for OFD-AD and 32 years (median, 28 years) for AD; 53% of the patients were female. The mean tumor size in the OFD-AD and AD groups combined was 7.8 cm, measured histologically. Sixteen percent of the patients sustained a pathological fracture prior to treatment. Local recurrence was recorded in 22% of the OFD-AD cases and 24% of the AD cases. None of the recurrences in the OFD-AD group progressed to AD. Metastatic disease was found in 18% of the AD cases and fatal disease, in 11% of the AD cases. No metastatic or fatal disease was reported in the OFD-AD group. Multivariate Cox regression analysis demonstrated that uncontaminated resection margins (hazard ratio [HR] = 0.164, 95% confidence interval [CI] = 0.092 to 0.290, p < 0.001), pathological fracture (HR = 1.968, 95% CI = 1.076 to 3.600, p = 0.028), and sex (female versus male: HR = 0.535, 95% CI = 0.300 to 0.952, p = 0.033) impacted the risk of local recurrence. CONCLUSIONS: OFD-AD and AD are parts of a disease spectrum but should be regarded as different entities. Our results support reclassification of OFD-AD into the intermediate locally aggressive category, based on the local recurrence rate of 22% and absence of metastases. In our study, metastatic disease was restricted to the AD group (an 18% rate). We advocate wide resection with uncontaminated margins including bone and involved periosteum for both OFD-AD and AD. LEVEL OF EVIDENCE: Prognostic Level IV. See Instructions for Authors for a complete description of levels of evidence.

Prevalence of undiagnosed atrial fibrillation in elderly individuals and potential cost-effectiveness of non-invasive ambulatory electrocardiographic screening: The ASSERT-III study.

BACKGROUND: In approximately 10% of patients with implanted pacemakers or defibrillators, previously unrecognized atrial fibrillation (AF) is detected within 3 months. It is unknown whether elderly patients without implanted devices have a similar prevalence of undiagnosed AF using non-invasive ECG monitoring, and if this approach to screening in this population is cost-effective. METHODS: Individuals ≥80 years old attending outpatient clinics without a history of AF and with hypertension and one additional risk factor underwent 30 days of continuous ECG monitoring with an option for an additional 30 days of monitoring if no AF was detected. The primary outcome was AF ≥ 6 min. Cost-effectiveness to prevent stroke was estimated using a Markov model based on observed AF detection rates and data from the published literature. RESULTS: Among 129 patients enrolled, 100 initiated monitoring for an average duration of 36 ±â€¯21 days. The proportion of patients that completed at least 30 days of monitoring was 59%. Average age was 84 ±â€¯3 years and mean CHA2DS2-VASc score was 4.5 ±â€¯1.2. AF ≥ 6 min was documented in 14%, ≥6 h in 8%, and ≥24 h in 3%. One week of monitoring costed $50,000 per quality-adjusted life-year-gained, 30 days and 60 days of monitoring costed $70,000 and $84,000, respectively. CONCLUSIONS: Continuous non-invasive ECG monitoring is feasible in elderly patients. Undiagnosed AF is present in many elderly individuals, with 1 in 7 having episodes lasting ≥6 min. One week of monitoring may be cost-effective for stroke prevention in this population.

Design and rationale of the atrial fibrillation occurring transiently with stress (AFOTS) incidence study.

BACKGROUND: Atrial fibrillation (AF) is often detected for the first time in patients hospitalized for medical illness or non-cardiovascular surgery. AF occurring transiently with stress (AFOTS) describes this manifestation of AF, which may either be the result of a non-cardiac stressor, or existing paroxysmal AF that was not previously detected. Current estimates of AFOTS incidence are imprecise: ranging from 1 to 44%, owing to the marked heterogeneity in patient populations, identification and methods used to detect AFOTS. METHODS: The prospective, two-centre epidemiological AFOTS Incidence study will enroll 250 consecutive participants without a history of AF but with at increased risk of AF (Age ≥ 65 or >50 with one risk factor for AF) admitted to intensive care units (ICUs) for medical illness or non-cardiac surgery. Upon admission, participants will wear an ECG patch monitor that will remain in place for 14 days, or until discharge from hospital. Patients' consent to participation is deferred for up to 72 h after admission. The primary endpoint is the incidence of AF lasting ≥30 s. The study is powered to detect an AF incidence of 17% ±â€¯5%. RESULTS: We conducted a vanguard feasibility study, and 55 participants have completed participation. The median duration of monitoring was seven days. AF was detected by the clinical team in 8 participants (14%; 95% Confidence Interval 7-26%). CONCLUSIONS: The AFOTS Incidence study will employ a systematic and highly sensitive protocol for detecting AFOTS in medical illness and non-cardiac surgery ICU patients. This study is feasible and will provide a reliable estimate of the true incidence of AFOTS in this population.

Phospholamban cardiomyopathy: a Canadian perspective on a unique population.

INTRODUCTION: Phospholamban cardiomyopathy is an inherited cardiomyopathy, characterised by a defect in regulation of the sarcoplasmic reticulum Ca2+ pump, often presenting with malignant arrhythmias and progressive cardiac dysfunction occurring at a young age. METHODS: Phospholamban R14del mutation carriers and family members were identified from inherited arrhythmia clinics at 13 sites across Canada. Cardiac investigations, including electrocardiograms, Holter monitoring (premature ventricular complexes, PVCs), and imaging results were summarised. RESULTS: Fifty patients (10 families) were identified (median age 30 years, range 3-71, 46% female). Mutation carriers were more likely to be older, have low-voltage QRS, T­wave inversion, frequent PVCs, and cardiac dysfunction, compared to unaffected relatives. Increasing age, low-voltage QRS, T­wave inversion, late potentials, and frequent PVCs were predictors of cardiac dysfunction (p < 0.05 for all). Older carriers (age ≥45 years) were more likely to have disease manifestations than were their younger counterparts, with disease onset occurring at an older age in Canadian patients and their Dutch counterparts. DISCUSSION: Among Canadian patients with phospholamban cardiomyopathy, clinical manifestations resembled those of their Dutch counterparts, with increasing age a major predictor of disease manifestation. Older mutation carriers were more likely to have electrical and structural abnormalities, and may represent variable expressivity, age-dependent penetrance, or genetic heterogeneity among Canadian patients.

Anti-C1 domain antibodies that accelerate factor VIII clearance contribute to antibody pathogenicity in a murine hemophilia A model.

Essentials Inhibitor formation remains a challenging complication of hemophilia A care. The Bethesda assay is the primary method used for determining bleeding risk and management. Antibodies that block factor VIII binding to von Willebrand factor can increase FVIII clearance. Antibodies that increase clearance contribute to antibody pathogenicity. SUMMARY: Background The development of neutralizing anti-factor VIII (FVIII) antibodies remains a challenging complication of modern hemophilia A care. In vitro assays are the primary method used for quantifying inhibitor titers, predicting bleeding risk, and determining bleeding management. However, other mechanisms of inhibition are not accounted for in these assays, which may result in discrepancies between the inhibitor titer and clinical bleeding symptoms. Objectives To evaluate FVIII clearance in vivo as a potential mechanism for antibody pathogenicity and to determine whether increased FVIII dosing regimens correct the associated bleeding phenotype. Methods FVIII-/- or FVIII-/- /von Willebrand factor (VWF)-/- mice were infused with anti-FVIII mAbs directed against the FVIII C1, C2 or A2 domains, followed by infusion of FVIII. Blood loss via the tail snip bleeding model, FVIII activity and FVIII antigen levels were subsequently measured. Results Pathogenic anti-C1 mAbs that compete with VWF for FVIII binding increased the clearance of FVIII-mAb complexes in FVIII-/- mice but not in FVIII-/- /VWF-/- mice. Additionally, pathogenic anti-C2 mAbs that inhibit FVIII binding to VWF increased FVIII clearance in FVIII-/- mice. Anti-C1, anti-C2 and anti-A2 mAbs that do not inhibit VWF binding did not accelerate FVIII clearance. Infusion of increased doses of FVIII in the presence of anti-C1 mAbs partially corrected blood loss in FVIII-/- mice. Conclusions A subset of antibodies that inhibit VWF binding to FVIII increase the clearance of FVIII-mAb complexes, which contributes to antibody pathogenicity. This may explain differences in the bleeding phenotype observed despite factor replacement in some patients with hemophilia A and low-titer inhibitors.

Are there increased periprocedural complications with the MRI-conditional Medtronic Revo SureScan Pacing System? : A meta-analysis.

BACKGROUND: The use of magnetic resonance imaging (MRI)-conditional permanent pacemakers has increased significantly. In this meta-analysis, we examine the safety of MRI-conditional pacing systems in comparison with conventional systems. METHODS: An electronic search was performed using major databases, including studies that compared the outcomes of interest between patients receiving MRI-conditional pacemakers (MRI group) versus conventional pacemakers (control group). RESULTS: Six studies (5 retrospective and 1 prospective non-randomised) involving 2,118 adult patients were identified. The MRI-conditional pacemakers, deployed in 969 patients, were all from a single manufacturer (Medtronic Pacing System with 5086 leads). The rate of pacemaker lead dislodgement (atrial and ventricular) was significantly higher in the MRI group (3% vs. 1%, OR 2.47 (95% CI 1.26; 4.83), p = 0.008). The MRI group had a significantly higher rate of pericardial complications (2% vs. 1%, OR 4.23 (95% CI 1.18; 15.10), p = 0.03) and a numerically higher overall complication rate in comparison with the conventional group (6% vs. 3%, OR 2.02 (95% CI 0.88; 4.66), p = 0.10) but this was not statistically significant. CONCLUSIONS: In this meta-analysis, the rates of pacemaker lead dislodgement and pericardial complications were significantly higher with the Medtronic MRI-conditional pacing system.

Identification of aggregates in therapeutic formulations of recombinant full-length factor VIII products by sedimentation velocity analytical ultracentrifugation.

Essentials Factor VIII inhibitors are the most serious complication in patients with hemophilia A. Aggregates in biopharmaceutical products are an immunogenic risk factor. Aggregates were identified in recombinant full-length factor VIII products. Aggregates in recombinant factor VIII products are identified by analytical ultracentrifugation. SUMMARY: Background The development of inhibitory anti-factor VIII antibodies is the most serious complication in the management of patients with hemophilia A. Studies have suggested that recombinant full-length FVIII is more immunogenic than plasma-derived FVIII, and that, among recombinant FVIII products, Kogenate is more immunogenic than Advate. Aggregates in biopharmaceutical products are considered a risk factor for the development of anti-drug antibodies. Objective To evaluate recombinant full-length FVIII products for the presence of aggregates. Methods Advate, Helixate and Kogenate were reconstituted to their therapeutic formulations, and subjected to sedimentation velocity (SV) analytical ultracentrifugation (AUC). Additionally, Advate and Kogenate were concentrated and subjected to buffer exchange by ultrafiltration to remove viscous cosolvents for the purpose of measuring s20,w values and molecular weights. Results The major component of all three products was a population of ~7.5 S heterodimers with a weight-average molecular weight of ~230 kDa. Helixate and Kogenate contained aggregates ranging from 12 S to at least 100 S, representing ≈ 20% of the protein mass. Aggregates greater than 12 S represented < 3% of the protein mass in Advate. An approximately 10.5 S aggregate, possibly representing a dimer of heterodimers, was identified in buffer-exchanged Advate and Kogenate. SV AUC analysis of a plasma-derived FVIII product was confounded by the presence of von Willebrand factor in molar excess over FVIII. Conclusions Aggregate formation has been identified in recombinant full-length FVIII products, and is more extensive in Helixate and Kogenate than in Advate. SV AUC is an important method for characterizing FVIII products.

A discontinuous autoinhibitory module masks the A1 domain of von Willebrand factor.

Essentials The mechanism for the auto-inhibition of von Willebrand factor (VWF) remains unclear. Hydrogen exchange of two VWF A1 fragments with disparate activities was measured and compared. Discontinuous residues flanking A1 form a structural module that blocks A1 binding to the platelet. Our results suggest a potentially unified model of VWF activation. Click to hear an ISTH Academy presentation on the domain architecture of VWF and activation by elongational flow by Dr Springer SUMMARY: Background How von Willebrand factor (VWF) senses and responds to shear flow remains unclear. In the absence of shear flow, VWF or its fragments can be induced to bind spontaneously to platelet GPIbα. Objectives To elucidate the auto-inhibition mechanism of VWF. Methods Hydrogen-deuterium exchange (HDX) of two recombinant VWF fragments expressed from baby hamster kidney cells were measured and compared. Results The shortA1 protein contains VWF residues 1261-1472 and binds GPIbα with a significantly higher affinity than the longA1 protein that contains VWF residues 1238-1472. Both proteins contain the VWF A1 domain (residues 1272-1458). Many residues in longA1, particularly those in the N- and C-terminal sequences flanking the A1 domain, and in helix α1, loops α1ß2 and ß3α2, demonstrated markedly reduced HDX compared with their counterparts in shortA1. The HDX-protected region in longA1 overlaps with the GPIbα-binding interface and is clustered with type 2B von Willebrand disease (VWD) mutations. Additional comparison with the HDX of denatured longA1 and ristocetin-bound longA1 indicates the N- and C-terminal sequences flanking the A1 domain form cooperatively an integrated autoinhibitory module (AIM) that interacts with the HDX-protected region. Binding of ristocetin to the C-terminal part of the AIM desorbs the AIM from A1 and enables longA1 binding to GPIbα. Conclusion The discontinuous AIM binds the A1 domain and prevents it from binding to GPIbα, which has significant implications for the pathogenesis of type 2B VWD and the shear-induced activation of VWF activity.

Physical activity and motivational predictors of changes in health behavior and health among DM2 and CAD patients.

This study tested a physical activity intervention and the self-determination theory (SDT) process model of health-behavior change and health among 108 adult patients with both diabetes mellitus type 2 (DM2) and coronary artery disease (CAD). Patients were randomly assigned to an organized physical activity intervention group (led by instructors) or a non-physical activity control group. At baseline and after 12 months, we measured the following: needs satisfaction, autonomous and controlled motivation for physical activity, perceived competence for physical activity and blood sugar testing, physical activity and blood sugar testing, body weight, glucose control (HbA1c), and self-perceptions of general health and vitality. The intervention produced, as hypothesized, significant changes in all study variables in favor of the experimental group (Cohen's d effect sizes: 0.23-0.72), except the non-significant result for controlled motivation and body weight. The data supported the SDT process model, in which the effect of the intervention significantly predicted indirect changes in behavior and health through motivation variables. Considering the moderate to large effects on increases in motivation, behavior, and health, promoting organized physical activity programs that are perceived as need-supportive may have important health implications for patients with DM2 and CAD.

Anti-factor VIII antibodies in brothers with haemophilia A share similar characteristics.

INTRODUCTION: The development of neutralizing antibodies (inhibitors) against coagulation factor VIII (FVIII) is currently the most serious complication for patients with haemophilia A undergoing FVIII replacement therapy. Several genetic factors have been acknowledged as risk factors for inhibitor development. AIM: To analyze the influence of genetic factors on the nature of the humoral immune response to FVIII in eight brother pairs with inhibitors. METHODS: The domain specificity of FVIII-specific IgG was analysed by antibody binding to FVIII fragments and homologue-scanning mutagenesis (HSM). The FVIII-specific IgG subclasses were measured by direct ELISA. RESULTS: Of the 16 patient analysed with both methods, 12 had A2- and 13 had C2-specific IgG. The presence of A1-, A3- or C1-specific IgG was identified in nine of 14 patients analysed by HSM. IgG1, IgG2 and IgG4 subclasses contributed to the anti-FVIII IgG response, and the amount of FVIII-specific IgG1 (r = 0.66) and IgG4 (r = 0.69) correlated significantly with inhibitor titres. Patients with high concentrations of total anti-FVIII IgG (r = 0.69) or high inhibitor titres (r = 0.52) had a high proportion of FVIII-specific IgG4. Statistical analysis revealed trends/evidence that the subclass distribution (P = 0.0847) and domain specificity to HC/LC (P = 0.0883) and A2/C2 (P = 0.0011) of anti-FVIII IgG were more similar in brothers compared to unrelated subjects. CONCLUSION: Overall, our data provide a first hint that anti-FVIII IgG characteristics are comparable among haemophilic brothers with inhibitors. Whether genetic factors also influence the nature of patients' antibodies needs to be confirmed in a larger study population.

Characterization of a genetically engineered mouse model of hemophilia A with complete deletion of the F8 gene.

UNLABELLED: ESSENTIALS: Anti-factor VIII (FVIII) inhibitory antibody formation is a severe complication in hemophilia A therapy. We genetically engineered and characterized a mouse model with complete deletion of the F8 coding region. F8(TKO) mice exhibit severe hemophilia, express no detectable F8 mRNA, and produce FVIII inhibitors. The defined background and lack of FVIII in F8(TKO) mice will aid in studying FVIII inhibitor formation. BACKGROUND: The most important complication in hemophilia A treatment is the development of inhibitory anti-Factor VIII (FVIII) antibodies in patients after FVIII therapy. Patients with severe hemophilia who express no endogenous FVIII (i.e. cross-reacting material, CRM) have the greatest incidence of inhibitor formation. However, current mouse models of severe hemophilia A produce low levels of truncated FVIII. The lack of a corresponding mouse model hampers the study of inhibitor formation in the complete absence of FVIII protein. OBJECTIVES: We aimed to generate and characterize a novel mouse model of severe hemophilia A (designated the F8(TKO) strain) lacking the complete coding sequence of F8 and any FVIII CRM. METHODS: Mice were created on a C57BL/6 background using Cre-Lox recombination and characterized using in vivo bleeding assays, measurement of FVIII activity by coagulation and chromogenic assays, and anti-FVIII antibody production using ELISA. RESULTS: All F8 exonic coding regions were deleted from the genome and no F8 mRNA was detected in F8(TKO) mice. The bleeding phenotype of F8(TKO) mice was comparable to E16 mice by measurements of factor activity and tail snip assay. Similar levels of anti-FVIII antibody titers after recombinant FVIII injections were observed between F8(TKO) and E16 mice. CONCLUSIONS: We describe a new C57BL/6 mouse model for severe hemophilia A patients lacking CRM. These mice can be directly bred to the many C57BL/6 strains of genetically engineered mice, which is valuable for studying the impact of a wide variety of genes on FVIII inhibitor formation on a defined genetic background.

Multi-centre variability of ISCEV standard ERGs in two normal adults.

BACKGROUND: The reproducibility of an individual's full-field ERG between centres has not previously been investigated. METHODS: ERGs were recorded using both silver thread and skin electrodes from the same two normal adult subjects at 15 UK centres using routine, local protocols and a highly standardised, 'ISCEV-specified' protocol matching the values specified in the ISCEV standard; where the ISCEV standard allows options, a single value was chosen. RESULTS: Inter-ocular differences were small, and amplitudes were smaller for skin than silver thread electrodes. No centre produced outlying data points, and ERGs across all 15 centres were remarkably similar. Amplitude variability was less for local protocols (using LED flashes) than for the ISCEV-specified protocol using xenon flashes (22 vs. 24 %, p = 0.01), but peak time variability was less for the ISCEV-specified protocol (6.1 vs. 7.4 %, p = 0.001). Only the DA 0.01 ERG correlated with photometric variability. The bifidity of the DA 3 a-wave doubled its peak time variability compared with the DA 10 a-wave. CONCLUSIONS: Inter-centre amplitude variability was typically within clinically significant thresholds, suggesting that inter-centre variability with suitable standardisation may not add more to total variability than inter-subject variability. Variability improvements gained by the tighter specifications of the ISCEV-specified protocol were possibly more than lost due to imprecisions of xenon flashtubes. Peak time variability was far lower than amplitude variability, corresponding with acceptable variability of biochemical assays. These results represent a vindication of the existence of an ERG standard and suggest that further standardisation would lend itself to greater reproducibility of ERGs worldwide.

Reduced-dose warfarin or interrupted warfarin with heparin bridging for pacemaker or defibrillator implantation: a randomized trial.

BACKGROUND: Perioperative management with reduced-dose warfarin is of potential interest by eliminating the need for bridging while still maintaining a degree of anticoagulation. The outcomes of this regimen have not been well determined. METHODS: In a randomized controlled trial we compared two regimens for management of anticoagulation with warfarin in patients with implantation of a pacemaker or defibrillator. Half dose of warfarin for 3-6 days, depending on the baseline international normalized ratio (INR), before surgery aiming at an INR of ≤ 1.7 was compared with interrupted warfarin for 5 days with preoperative bridging with low-molecular-weight heparin (LMWH) at therapeutic dose for 2.5 days. Main safety outcome was pocket hematoma. Secondary outcomes were major bleeding, thromboembolism - all within 1 month, days of hospitalization and number of patients requiring correction of INR with vitamin K. RESULTS: The study was planned for 450 patients but it was discontinued prematurely due to a change in practice. Pocket hematoma occurred in 4 of 85 patients (5%) randomized to the bridged regimen and in 3 of 86 patients (3%) randomized to reduced-dose warfarin. One pocket hematoma in each group was severe. There were no major hemorrhages or thromboembolism within the 1-month window. Duration of hospitalization was similar in the two groups. Correction of INR the day before surgery with vitamin K had to be used for significantly more patients in the reduced-dose warfarin group (41%) than in the bridged regimen group (6%). CONCLUSION: The reduced-dose warfarin regimen appeared to have similar safety after device implantation as interrupted warfarin with preoperative LMWH bridging. Due to premature discontinuation no firm conclusion can be drawn. The reduced-dose warfarin regimen often failed to achieve the intended preoperative INR. ClinicalTrials.gov Identifier: NCT 02094157.

Lack of recombinant factor VIII B-domain induces phospholipid vesicle aggregation: implications for the immunogenicity of factor VIII.

Factor VIII (FVIII) is a multidomain blood plasma glycoprotein. Activated FVIII acts as a cofactor to the serine protease factor IXa within the membrane-bound tenase complex assembled on the activated platelet surface. Defect or deficiency in FVIII causes haemophilia A, a severe hereditary bleeding disorder. Intravenous administration of plasma-derived FVIII or recombinant FVIII concentrates restores normal coagulation in haemophilia A patients and is used as an effective therapy. In this work, we studied the biophysical properties of clinically potent recombinant FVIII forms: human FVIII full-length (FVIII-FL), human FVIII B-domain deleted (FVIII-BDD) and porcine FVIII-BDD bound to negatively charged phospholipid vesicles at near-physiological conditions. We used cryo-electron microscopy (Cryo-EM) as a direct method to evaluate the homogeneity and micro-organization of the protein-vesicle suspensions, which are important for FVIII therapeutic properties. Applying concurrent Cryo-EM, circular dichroism and dynamic light scattering studies to the three recombinant FVIII forms when bound to phospholipid vesicles revealed novel properties for their functional, membrane-bound state. The three FVIII constructs have similar activity, secondary structure distribution and bind specifically to negatively charged phospholipid membranes. Human and porcine FVIII-BDD induce strong aggregation of the vesicles, but the human FVIII-FL form does not. The proposed methodology is effective in characterizing and identifying differences in therapeutic recombinant FVIII membrane-bound forms near physiological conditions, because protein-containing aggregates are considered to be a factor in increasing the immunogenicity of protein therapeutics. This will provide better characterization and development of safer and more effective FVIII products with implications for haemophilia A treatment.

Epitope mapping of inhibitory antibodies targeting the C2 domain of coagulation factor VIII by hydrogen-deuterium exchange mass spectrometry.

BACKGROUND: The development of anti-factor VIII antibodies (inhibitors) is a significant complication in the management of patients with hemophilia A, leading to significant increases in morbidity and treatment cost. Using a panel of mAbs against different epitopes on FVIII, we have recently shown that epitope specificity, inhibitor kinetics and time to maximum inhibition are more important than inhibitor titer in predicting responses to FVIII and the combination of FVIII and recombinant FVIIa. In particular, a subset of high-titer inhibitors responded to high-dose FVIII, which would not be predicted on the basis of their inhibitor titer alone. Thus, the ability to quickly map the epitope spectrum of patient plasma with a clinically feasible assay may fundamentally change how clinicians approach the treatment of high-titer inhibitor patients. OBJECTIVES: To map the epitopes of anti-FVIII mAbs, three of which are classic inhibitors and one of which is a non-classic inhibitor, by the use of hydrogen-deuterium exchange coupled with mass spectrometry (HDX-MS). METHODS: The binding epitopes of four mAbs targeting the FVIII C2 domain were mapped with HDX-MS. RESULTS: The epitopes determined with HDX-MS are consistent with those obtained earlier through structural characterization and antibody competition assays. In addition, classic and non-classic inhibitor epitopes could be distinguished by the use of a limited subset of C2 domain-derived peptic fragments. CONCLUSION: Our results demonstrate the effectiveness and robustness of the HDX-MS method for epitope mapping, and suggest a potential role of rapid mapping of FVIII inhibitor epitopes in facilitating individualized treatment of inhibitor patients.

Anticoagulation management in patients with mechanical heart valves having pacemaker or defibrillator insertion.

BACKGROUND: In patients with a high risk for stroke and having invasive procedures with a high risk for bleeding it is unclear how anticoagulant therapy should be managed. METHODS: We reviewed data from all patients with mechanical heart valves, who had elective insertion or replacement of pacemaker or implantable cardioverter defibrillator (ICD) during the past 8years at our hospital. Data on anticoagulant treatment, pocket hematoma and thromboembolic complications were captured. RESULTS: Of the 111 patients reviewed, 68 (61%) had a mechanical valve in the mitral position with or without other valves replaced and 43 (39%) had a mechanical valve only in the aortic position. Fifty-nine (53%) were undergoing replacement for their device. Six patients received a tapered warfarin regimen and 102 received preoperative bridging anticoagulation of whom 12 also received postoperative bridging. One stroke occurred 40days after pacemaker replacement in a patient with mitral mechanical valve and without postoperative bridging. Six patients (5.5%) developed pocket hematoma without a significant association to postoperative bridging, type of mechanical valve or to type of device. Predictors for pocket hematoma appeared to be replacement surgery (odds ratio 12.5; 95% confidence interval [CI], 0.69-228) and an international normalized ratio of 1.5 or higher on the day of surgery (odds ratio 8.4; 95% CI, 0.96-68.1). CONCLUSION: We found a low risk for stroke in the absence of postoperative bridging. For patients with device replacement surgery reversal of the anticoagulant effect at the time of procedure might reduce the risk for pocket hematoma, but this requires prospective evaluation including the risk of thromboembolism.