Evaluation of nonneutralizing antibodies against factor VIII in severe haemophilia A patients from India Haemophilia A is treated by replacement therapy with factor VIII (FVIII) concentrate. This strategy of treatment is ineffective in some patients due to the development of neutralizing antibodies (NNAs) against FVIII. The inhibitors have been identified to act against the functional domains of FVIII. The presence of NNAs against FVIII has also been identified. There is limited data on the prevalence and significance of NNA in haemophilia. To identify the presence of NNA in severe haemophilia A in our population, patients who were recruited from community-based camps were evaluated for FVIII activity. The patient's samples were further analysed for inhibitor activity with Nijmegen-Bethesda Assay and for NNAs using an in-house ELISA. 312 severe haemophilia patients were analysed for inhibitors and NNA. In-house ELISA picked up antibodies in 56 patients (17.9%). Of these 42 (13.7%) had inhibitory antibodies and in 14 patients (4.5%) there was no evidence of FVIII inhibitory activity. A substantial number of patients with severe haemophilia A have NNA. Continuous long-term follow-up is required in this cohort to evaluate the significance of this observation. Correspondence to Sukesh C. Nair, Department of Transfusion Medicine and Immunohaematology, Christian Medical College, Vellore 632004, Tamil Nadu, India. Tel: +91 416 2283431; e-mail: scnair@cmcvellore.ac.in Received 12 March, 2019 Revised 3 June, 2019 Accepted 9 July, 2019 Copyright © 2019 YEAR Wolters Kluwer Health, Inc. All rights reserved.

Patient characteristics when starting treatment and patterns of treatment in adults with chronic immune thrombocytopenia Asymptomatic patients with primary chronic immune thrombocytopenia (ITP) are not recommended treatment if their platelet counts are above 30 × 109/l. Factors such as age and comorbidities may influence clinical manifestations and should be considered for treatment decisions. The aim of this study was to determine the impact of clinical characteristics for initiation of ITP treatment, and the patterns of ITP treatment given. We performed an observational cohort study in Sweden with information from medical records and National Health Registers. Adults diagnosed with incident primary ITP between years 2009 and 2016 were included. Multinomial logistic regression was used to assess the impact of factors predicting treatment start. Out of 858 patients with chronic ITP from 71 hospitals we identified 585 (68%) with a first ITP treatment. For 537 (92%) corticosteroids were the first choice. The median platelet counts at start of treatment was 12 × 109/l (interquartile range 5–27 × 109/l). The variables predicting treatment start were platelet counts below 20 × 109/l and treatment with antihypertensive drugs. Patients with diabetes were less likely to receive corticosteroids. Severe bleeding occurred in 75 (13%) of the patients. Platelet counts below 20 × 109/l, antihypertensive treatment and bleedings were the strongest predictors of treatment start, diabetes yielded lower odds to start corticosteroid treatment. The majority of the patients had corticosteroids as first treatment while second treatment was diverse. Asymptomatic thrombocytopenia is not considered a reason as such for initiating treatment. In the latter years, splenectomy seemed to occur later in the course of treatment. Correspondence to Charlotta Ekstrand, PhD Student, Department of Medicine, Centre for Pharmacoepidemiology, Karolinska Institutet, SE-171 76 Stockholm, Sweden. Tel: +46 8 517 79314; fax: +46 8 517 79304; e-mail: charlotta.ekstrand@ki.se Received 12 March, 2019 Revised 2 July, 2019 Accepted 27 July, 2019 Supplemental digital content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versions of this article on the journal's website (www.bloodcoagulation.com). Copyright © 2019 YEAR Wolters Kluwer Health, Inc. All rights reserved.

The effect of vitamin D levels on gastrointestinal bleeding in patients with warfarin therapy Upper tract gastrointestinal system (GIS) bleeding is considered as an important cause of morbidity and mortality despite modern and advanced endoscopic interventions. In patients with thrombotic state and vitamin D deficiency, vitamin D analogs and vitamin D receptor activators have been determined as adjunctive anticoagulant treatment in previous studies. However, these studies did not evaluate or reveal the possible bleeding diathesis. In this article, we evaluated the vitamin D status in patients with warfarin treatment and upper tract GIS bleeding. A total of 75 patients with a definite diagnosis of upper tract GIS bleeding who had a treatment of warfarin and current vitamin D measurement; and a total of 75 control patients without any recent or prior GIS bleeding who had a treatment of warfarin and current vitamin D measurement were enrolled to the study. GIS bleeding group had a proportionally higher vitamin D treatment (29.3 vs. 17.3%). In GIS bleeding group, the prevalence of vitamin D level less than 20 ng/ml was significantly lower (66.7 vs. 82.7%; P = 0.024) and the prevalence of vitamin D level at least 30–100 ng/ml was significantly higher (25.3 vs. 10.7%; P = 0.019). According to a subgroup analysis; in patients with a vitamin D level at least 30–100 ng/ml, major bleeding rate was significantly higher compared with other patients. There was not a significant difference regarding mortality between the groups. Our study is the first which represents the possible bleeding effect of elevated vitamin D levels and vitamin D treatment in patients with warfarin treatment. Correspondence to Ümran Keskin, MD, Internal Medicine, Health Sciences University, Haydarpasa Numune Training and Research Hospital, Uskudar, Istanbul, Turkey. Tel: +90 5535451454; e-mail: drumrankeskin@gmail.com Received 27 May, 2019 Revised 16 July, 2019 Accepted 27 July, 2019 Copyright © 2019 YEAR Wolters Kluwer Health, Inc. All rights reserved.

Anticoagulant activity of krait, coral snake, and cobra neurotoxic venoms with diverse proteomes are inhibited by carbon monoxide Background A phenomena of interest is the in vitro anticoagulant effects of neurotoxins found in elapid venoms that kill by paralysis. These enzymes include phospholipase A2 (PLA2), and it has recently been demonstrated that carbon monoxide inhibits the PLA2-dependent neurotoxin contained in Mojave rattlesnake type A venom. The purpose of this investigation was to assess if the anticoagulant activity of elapid venoms containing PLA2 and/or three finger toxins could be inhibited by carbon monoxide. Methods Venoms collected from Bungarus multicinctus, Micrurus fulvius, and five Naja species were exposed to carbon monoxide via carbon monoxide releasing molecule-2 prior to placement into human plasma. Coagulation kinetics were assessed via thrombelastography. Results Compared with plasma without venom addition, all venoms had significant anticoagulant effects, with a 160-fold range of concentrations having similar anticoagulant effects in a species-specific manner. Carbon monoxide significantly inhibited the anticoagulant effect of all venoms tested, but inhibition was not complete in all cases. Conclusion Given that individual neurotoxin activity often depends on intact activity that includes anticoagulant action, it may be possible that carbon monoxide inhibits neurotoxicity. Future investigation is justified to assess such carbon monoxide mediated inhibition with purified neurotoxins in vitro and in vivo. Correspondence to Vance G. Nielsen, MD, The Department of Anesthesiology, University of Arizona College of Medicine, PO Box 245114, 1501 North Campbell Avenue, Tucson, AZ 85724-5114, USA. Tel: +1 520 626 7195; fax: +1 520 626 6943; e-mail: vgnielsen333@gmail.com Received 4 June, 2019 Accepted 29 July, 2019 Copyright © 2019 YEAR Wolters Kluwer Health, Inc. All rights reserved.

Perioperative laboratory monitoring in congenital haemophilia patients with inhibitors: a systematic literature review Although the use of clotting factor concentrates is the mainstay of haemophilia care, the development of inhibitors complicates disease management. Perioperative management of patients with inhibitors is therefore a challenge. A systematic literature review was performed to identify literature reporting on the perioperative monitoring and management of haemophilia. MEDLINE, Embase and Cochrane databases were searched from database inception to 26 March 2018. Recent congress proceedings were also searched. Titles and abstracts, then full texts, were screened for relevance by two reviewers. Quality of included studies was assessed using the Critical Appraisal Skills Programme checklist. Of the 2033 individual entries identified, 86 articles met the inclusion criteria. The identified studies were screened again to find articles reporting perioperative laboratory monitoring in patients with congenital haemophilia A or B, resulting in 24 articles undergoing data extraction. Routine perioperative assay monitoring practices were the most commonly reported (n = 20/24); thrombin generation assay was the least commonly reported (n = 2/24). Other monitoring practices described were factor VII and factor VIII coagulation activity (n = 8/24, n = 5/24, respectively), and thromboelastography or rotational thromboelastometry assessments (n = 3/24). The impact of monitoring on treatment decisions was, however, rarely reported. In conclusion, many methods of perioperative monitoring of haemophilia patients with inhibitors have been identified in this review, yet there is a lack of reporting in larger scale cohort studies. More detailed reporting on the impact of monitoring outcomes on treatment decisions is also needed to share best practice, particularly as new therapeutic agents emerge. Correspondence to Daniel P. Hart, The Royal London Hospital Haemophilia Centre, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London, UK. E-mail: d.hart@qmul.ac.uk Received 7 March, 2019 Revised 30 May, 2019 Accepted 9 July, 2019 Supplemental digital content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versions of this article on the journal's website (www.bloodcoagulation.com). Copyright © 2019 YEAR Wolters Kluwer Health, Inc. All rights reserved.

Thromboelastography testing in mice following blood collection from facial vein and cardiac puncture Blood collection is critical for mouse research studies particularly in hemostatic testing. Cardiac puncture; a standard effective method requires anesthesia and is a terminal procedure while facial vein technique allows multiple collections. Thromboelastography (TEG) is a global hemostasis test, provides a dynamic real-time picture of coagulation. However, TEG experiments in mice require large number of animals and may not allow pre/postinterventions assessment. In this study, we aimed to investigate the feasibility of facial vein sampling for TEG analysis as an alternative to cardiac puncture and examined the impact on coagulation results. Blood samples were obtained from a total of 10 C57BL/6 and CD-1 mice via cardiac puncture and a total of another eight mice of similar strains via facial vein sampling. We compared TEG parameters in both methods using descriptive statistics and the Student t test. Results show no significant difference in any of the TEG parameters between cardiac and facial vein blood indicating the two methods are comparable. Facial vein sampling provides a less costly alternative to cardiac puncture. It is a suitable blood collection method for pre/postinterventions or follow-up studies and it better addresses reduction and refinement goals in mouse studies. A larger study to evaluate the sex or strain and genetic background differences will be valuable. Correspondence to Dr Maha Othman, MD, MSc, PhD, Associate Professor, Department of Biomedical and Molecular Sciences, Queen's University, Boterell Hall, Room 513, Kingston, ON, Canada K7L 3N6. Tel: +1 613 533 6108; fax: +1 613 533 2022; e-mail: othman@queensu.ca Received 5 April, 2019 Revised 17 May, 2019 Accepted 26 June, 2019 Copyright © 2019 YEAR Wolters Kluwer Health, Inc. All rights reserved.

Limitations of prophylactic treatment in patients with hemophilia Prophylaxis with factor VIII concentrates is the cornerstone of treatment for severe hemophilia A. In children, early onset of primary and secondary prophylaxis is the gold standard in most of countries with adequate financial resources. In adults, it is reasonable to continue the prophylactic treatment initiated during childhood to preserve joint functionality. Adults with advanced hemophilic arthropathy can also benefit from tertiary prophylaxis, showing reductions in bleeding episodes, target joints and missed days from work. The long-term evolution of hemophilic patients usually depends on how successfully prophylaxis can prevent bleeding in both children and adults. For prophylaxis to be efficient it is essential to consider the resources available (factor concentrate use and trough levels) and the patient's bleeding phenotype and clinical situation (levels of activity required, presence of chronic synovitis and arthropathy), and to define the annual number of bleeds (particularly into joints) acceptable for each patient. Correspondence to María Fernanda López Fernández, MD, PhD, Hemostasis and Thrombosis Unit, Department of Hematology and Transfusion, Integrated Health Management Directorship, A Coruña, Spain. E-mail: mflopezfer@gmail.com Received 24 June, 2018 Revised 18 March, 2019 Accepted 20 March, 2019 Copyright © 2019 YEAR Wolters Kluwer Health, Inc. All rights reserved.