Background Coagulopathy is still a serious pattern of coronavirus-19 disease. We aimed to evaluate COVID-19-associated coagulopathy and multiple hemostatic markers in Egyptian patients. In addition, to assess coagulation acute phase reactants and its effect on the outcome. Methods The study included 106 COVID-19 patients, and 51 controls. All patients were positive for COVID-19 infection by nasopharyngeal swab for detection of viral RNA by real-time PCR. In addition to baseline data and radiological findings, the coagulation profile was done with special attention to Fibrinogen, d-dimer, Factor VIII, von Willebrand factor (VWF), Protein C, Protein S, Antithrombin III (ATIII) and Lupus anticoagulant (LA)-1 and 2. Results The results showed significantly higher VWF, d-dimer, and LA1 (screening) and LA2 (confirmation) in patients than a control group. Significantly higher d-dimer FVIII, VWF and LA1-2 were detected in the severe group. ATIII had high diagnostic accuracy in severity prediction. We found a significantly higher international randomized ratio (INR) and VWF among patients with thrombotic events. For prediction of thrombosis; VWF at cutoff > 257.7 has 83.3% sensitivity and 83.3% specificity. Conclusion Patients with COVID-19 infection are vulnerable to different forms of coagulopathy. This could be associated with poor outcomes. d-Dimer is a chief tool in diagnosis, severity evaluation but not thrombosis prediction. Early screening for this complication and its proper management would improve the outcome.

Background Glanzmann's thrombasthenia (GT) and Bernard-Soulier syndrome (BSS) are genetic bleeding syndromes characterized by defects in platelet aggregometry. Although these disorders are classified to be rare, their exact prevalence is still unknown; however, they are more frequent in communities where consanguineous marriages are common. Aim To study platelet surface glycoproteins expression using flow cytometry and to examine their correlation with bleeding severity using International Society of Thrombosis and Hemostasis–Bleeding Assessment Tools (ISTH-BAT) as bleeding score of choice. Patients and methods This case–control study included 51 patients with bleeding disorders recruited from the Department of Pediatric Hematology, Assiut University Hospital, in addition to 36 apparently healthy age- and sex-matched controls. All patients were tested for complete blood count (CBC), prothrombin time, partial thromboplastin time, platelet aggregation, and platelets surface glycoprotein analysis by flow cytometry. ISTH-BAT was used to register bleeding data for patients. Results GT and BSS had some similarities regarding the presentation and bleeding severity, but when CBC, platelet aggregation studies, and flow cytometric analysis were done, differentiation became much easier. GT patients showed a decrease in the expression of CD41 and CD61. Type I GT patients had more bleeding severity than type II and type III. BSS patients showed a decrease in expression of CD42b. There are correlations between the bleeding severity and CD41 in GT, and between the severity and CD42b in BSS. Conclusion Flow cytometric studies of platelet glycoproteins have great values in diagnosing BSS and GT, and further classifying GT cases into its three types. ISTH-BAT is a useful tool when dealing with platelet function disorders and has good sensitivity and ability to determine the severity.

von Willebrand Disease (VWD) is the most prevalent inherited bleeding disorder. Type 1 is the most common form of VWD and results in a partial quantitative deficiency of von Willebrand Factor (VWF). The mechanisms underlying type 1 VWD are still not very well understood although reduced VWF secretion and increased VWF clearance have been implicated in causing VWD. We aimed to characterize novel sequence variants (SV) identified in the VWF gene in type 1 VWD patients recruited through the Zimmerman Program for the Molecular and Clinical Biology of VWD in order to define the underlying mechanism and explore if SV in a particular domain are mechanistically similar. We utilized homozygous expression in human embryonic kidney cells (HEK-293T) to study the effect of VWF SV on VWF secretion, intracellular retention, multimerization, and function. Novel SV have been identified throughout the entire VWF protein. We introduced the following variants into a VWF-mycHis plasmid vector: V86M, W199X, C524Y, M947V, R960P, G994D, C996W, R1204W, Q1353X, E1660X, R1763Q, C2199Y, Q2256H, T2282I, P2524L, A2569E, C2693F, C2701Y, and C2754Y. Sequence variants were confirmed by Sanger sequencing. Variant VWF cDNA is transfected homozygously into HEK-293T cells. The supernatants and cell lysates from 3 independent transfections are collected and analyzed by ELISA for VWF:Ag and VWF binding to collagen type III (VWF:CB). VWF multimer structure is analyzed by SDS-agarose gel electrophoresis and western blotting. The genotype-phenotype patient data is correlated with the data from expression studies to explore a model to predict the impact of the SV on the VWD phenotype. Variants V86M, M947V, R1204W, R1763Q, Q2256H, T2282I, P2524L, A2569E, and C2693F demonstrated secretion comparable to that of wild type (WT)-VWF. In contrast, VWF variants R960P and C2701Y showed reduced VWF secretion (<50% of WT) with increased VWF in the cell lysate. VWF variants W199X, C524Y, G994D, C996W, Q1353X, E1660X, C2199Y, and C2754Y demonstrated a complete absence of secreted VWF. Not unexpectedly, homozygous expression of stop codon variants W199X, Q1353X, and E1660X demonstrated no VWF in the cell lysate. However, non-secreted VWF variants C524Y, G994D, C996W, C2199Y, and C2754Y showed intracellular retention with detectable VWF in the cell lysate. SV occurring at cysteine residues (C524Y, C996W, C2199Y, C2701Y, and C2754Y) all had reduced secretion and increased intracellular retention, consistent with altered conformation leading to increased intracellular chaperone interaction and proteasomal degradation. VWF binding to collagen is dependent on the presence of high molecular weight multimers (HMWM). VWF:CB/VWF:Ag is used to predict multimer structure with VWF:CB/VWF:Ag < 0.7 indicative of loss of the HMWM. VWF variants V86M, M947V, R1763Q, Q2256H, P2524L, C2701Y had VWF:CB/VWF:Ag ≥ 0.7 consistent with normal multimer structure, while variants R960P, R1204W, T2282I, A2569E, and C2693F had VWF:CB/VWF:Ag < 0.7 indicating abnormal multimer structure. 47.3% of the 19 VWF variants studied had normal VWF secretion, 10.5% had reduced secretion with increased intracellular retention, and 26.3% revealed absent secretion with intracellular retention. Variants with a premature stop codon did not synthesize VWF at all. Some SV had normal secretion and multimerization (V86M, M947V, R1204W, R1763Q, Q2256H, T2282I, P2524L, A2569E, and C2693F) implying that the VWD phenotype in these patients results from yet unidentified mechanisms and may not be associated with these SV. Reduced plasma survival is unlikely as these patients had normal VWFpp/VWF:Ag level consistent with normal VWF clearance. Among the VWF variants with normal or decreased secretion, 45.4% had reduced VWF:CB/VWF:Ag consistent with abnormal multimer structure. Heterozygous expression, as observed in the patient, is expected to normalize these multimerization defects. The decreased or absent secretion observed for 52.7% of the variants studied correlates with the patient phenotype, indicating reduced secretion is the mechanism underlying these patients' type 1 VWD phenotype. No domain-specific correlation of VWF secretion or multimer abnormality was observed. In summary, VWF expression studies confirmed the causative nature of many, but not all of the novel sequence variants identified in type 1 VWD subjects in the Zimmerman Program. Disclosures No relevant conflicts of interest to declare.