Laboratory Diagnosis of Activated Protein C Resistance and Factor V Leiden.

The factor V Leiden (FVL) polymorphism is known as the most common inherited risk factor for venous thrombosis. In turn, FVL is the leading cause of an activated protein C resistance (APCR) phenotype, in which the addition of exogenous activated protein C to plasma does not result in the expected anticoagulant effect. In the routine laboratory approach to the formal diagnosis of FVL, an initial positive screening plasma-based method for APCR is often performed, and only if needed, this is followed by a confirmatory DNA-based assay for FVL. Multiple methods with accepted sensitivity and specificity for determining an APCR/FVL phenotype are commonly categorized into two separate groups: (1) screening plasma-based assays, including qualitative functional clot-based assays, for APCR, and (2) confirmatory DNA-based molecular assays, entailing several tests and platforms, including polymerase chain reaction-based and non-PCR-based techniques, for FVL. This review will describe the methodological aspects of each laboratory test and prepare suggestions on the indication of APCR and FVL testing and method selection.

Molecular basis of rare congenital bleeding disorders.

Rare bleeding disorders (RBDs), including factor (F) I, FII, FV, FVII, combined FV and FVIII (CF5F8), FXI, FXIII and vitamin-K dependent coagulation factors (VKCF) deficiencies, are a heterogeneous group of hemorrhagic disorder with a variable bleeding tendency. RBDs are due to mutation in underlying coagulation factors genes, except for CF5F8 and VKCF deficiencies. FVII deficiency is the most common RBD with >330 variants in the F7 gene, while only 63 variants have been identified in the F2 gene. Most detected variants in the affected genes are missense (>50% of all RBDs), while large deletions are the rarest, having been reported in FVII, FX, FXI and FXIII deficiencies. Most were located in the catalytic and activated domains of FXI, FX, FXIII and prothrombin deficiencies. Understanding the proper molecular basis of RBDs not only can help achieve a timely and cost-effective diagnosis, but also can help to phenotype properties of the disorders.

The Impact of SARS-CoV-2 Infection on Blood Coagulation and Fibrinolytic Pathways: A Review of Prothrombotic Changes Caused by COVID-19.

The cardinal pathology of coronavirus disease 2019 (COVID-19) is a primary infection of pulmonary tract cells by severe acute respiratory syndrome coronavirus 2, provoking a local inflammatory response, often accompanied by cytokine storm and acute respiratory distress syndrome, especially in patients with severe disease. Systemic propagation of the disease may associate with thrombotic events, including deep vein thrombosis, pulmonary embolism, and thrombotic microangiopathy, which are important causes of morbidity and mortality in patients with COVID-19. This narrative review describes current knowledge of the pathophysiological mechanisms of COVID-19-associated coagulopathy, with focus on prothrombotic changes in hemostatic mediators, including plasma levels of clotting factors, natural anticoagulants, components of fibrinolytic system, and platelets. It will also highlight the central role of endothelial cells in COVID-19-associated coagulopathy. This narrative review discusses also potential therapeutic strategies for managing thrombotic complications. Awareness by medical experts of contributors to the pathogenesis of thrombotic events in COVID-19 is imperative to develop therapeutics not limited to regular anticoagulants. Instituting cooperation among medical personnel and researchers may lessen this novel virus' impact now, and in the event of recurrence.

The demographic and serological characteristics of COVID-19 convalescent plasma donors: Identification of basic criteria for optimal donor selection.

BACKGROUND AND OBJECTIVES: Convalescent plasma has attracted significant attention as a therapeutic option against infectious agents for more than a century. In March 2020, the use of Convalescent COVID-19 plasma (CCP) as a new research drug for COVID-19 treatment was approved by the FDA. The development of SARS-CoV-2 IgG antibodies following infection or vaccination is likely to be essential to provide adequate immunity for the population to halt the COVID19 pandemic. This study aimed to identify the criteria that would be used to determine the most appropriate CCP donors with the highest effective antiviral antibody titers. MATERIALS AND METHODS: In this prospective cohort, univariate analyses and multivariate regression analyses were performed to evaluate the relationship between characteristics of 11949 CCP donors and COVID-19 disease severity prior to donation with antibody titers estimated using ELISA technique and rapid tests. RESULTS: The antibody titer was measured among 8206 (68.7 %) donors. Elderly male and nulliparous female CCP donors who resided in the areas with high load of virus had positive ELISA and rapid test results as well as high levels of SARS-CoV-2 IgG antibodies titer. Moreover, the long hospital stay and elderly donors were the variables associated with high levels of SARS-CoV-2 IgG antibodies. CONCLUSION: This study suggests that nulliparous female and male donors with positive rapid tests who resided in areas with a higher prevalence of SARS-CoV-2, with more than 40 years of age and long hospitalization time can be the preferred donors for CCP donation.

Challenges and concerns of patients with congenital bleeding disorders affected by coronavirus disease 2019.

Coronavirus disease 2019 (COVID-19) is a new medical challenge for all individuals, especially for those with underlying disorders, such as congenital bleeding disorders (CBDs). Therefore, the pandemic might significantly change the behaviour of patients with CBDs and results in some challenges. In the present study, we assessed the main challenges of COVID-19 infection to patients with CBDs. Data were collected from medical files and interviews of patients with CBDs who had COVID-19 infection. Follow-ups were performed on patients who had active severe acute respiratory syndrome coronavirus 2 infection between April and October 2020. All patients were interviewed by an expert in order to collect the pertinent data. Some questions were about patients' preventive behaviors and feelings prior to infection, and some were about the consequences of infection on patients' replacement therapy and bleeding management. Among 25 patients, infection and death of loved ones (n: 7, 28%), and their own (n: 5, 20%) or family members' (n: 1, 4%) infection, and the resulting economic burden (n: 2, 8%) were main concerns. Six patients experienced depression during the pandemic. The pandemic caused all severely affected patients but one (n: 11, 92%) to abandon replacement therapy. However, two received on-demand therapy after exacerbation of their bleeding. Only one (25%) of four patients on prophylaxis received in-home therapy, whereas the others (75%) abandoned prophylaxis. It seems that COVID-19 infection has great consequences on the lives of patients with CBDs, causing some to take dangerous actions, such as abandonment of their treatment. Healthcare systems, and healthcare providers, should have an appropriate strategy for management of patients with CBDs that prevents infection and provides timely replacement therapy.

Standardization of Prothrombin Time/International Normalized Ratio (PT/INR).

The prothrombin time (PT) represents the most commonly used coagulation test in clinical laboratories. The PT is mathematically converted to the international normalized ratio (INR) for use in monitoring anticoagulant therapy with vitamin K antagonists such as warfarin in order to provide test results that are adjusted for thromboplastin and instrument used. The INR is created using two major PT 'correction factors', namely the mean normal PT (MNPT) and the international sensitivity index (ISI). Manufacturers of reagents and coagulometers have made some efforts to harmonizing INRs, for example, by tailoring reagents to specific coagulometers and provide associated ISI values. Thus, two types of ISIs may be generated, with one being a 'general' or 'generic' ISI and others being reagent/coagulometer-specific ISI values. Although these play a crucial role in improving INR results between laboratories, these laboratories reported INR values are known to still differ, even when laboratories use the same thromboplastin reagent and coagulometer. Moreover, ISI values for a specific thromboplastin can vary among different models of coagulometers from a manufacturer using the same method for clot identification. All these factors can be sources of error for INR reporting, which in turn can significantly affect patient management. In this narrative review, we provide some guidance to appropriate ISI verification/validation, which may help decrease the variability in cross laboratory reporting of INRs.

Molecular and clinical profile of type 2 von Willebrand disease in Iran: a thirteen-year experience.

Type 2 von Willebrand disease (VWD) is the most common congenital bleeding disorder, with variable bleeding tendency and a complex laboratory phenotype. In the current study, we report the clinical and molecular profile of a large number of Iranian patients with type 2 VWD. All exons, intron-exon boundaries, and untranslated regions were sequenced by Sanger sequencing for direct mutation detection. All identified mutations were confirmed in family members and by relevant bioinformatics studies. A total of 136 patients with type 2 VWD were diagnosed, including 42 (30.9%), 32 (23.6%), 38 (27.9%), and 24 (17.6%) patients with type 2A, type 2B, type 2M, and type 2N, respectively. Epistaxis (49%), gum bleeding (30.2%), ecchymosis (23.2%), and menorrhagia (16.3%) were the most common clinical presentations, while miscarriage (2.3%) and umbilical cord bleeding (0.8%) were the rarest. Thirty mutations were identified within the VWF gene, nine (30%) being novel, with p.Arg1379Cys (n = 20), p.Val1316Met (n = 13), p.Arg1597Trp (n = 13), p.Arg1374Cys (n = 10), p.Ser1506Leu (n = 10), and p.Arg1308Cys (n = 9) the most common. Type 2 VWD is a hemorrhagic disorder with variable bleeding tendency and a heterogeneous molecular basis in patients in Iran.