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Thrombotic thrombocytopenic purpura (TTP) is an acute, life‐threatening illness with disseminated platelet‐rich thromboses of small vessels that variably presents with the classic clinical “pentad” of microangiopathic hemolytic anemia, thrombocytopenia, fever, altered mental status, and acute kidney injury. Most cases are caused by an acquired autoantibody to ADAMTS13, a metalloproteinase that cleaves large von Willebrand Factor (vWF) multimers. The mainstay of treatment is daily therapeutic plasma exchange (TPE), sometimes with adjunctive pharmacologic immunosuppression. TPE is generally continued until the platelet count is greater than 150 × 103/µL and the lactate dehydrogenase is near normal for 2‐3 consecutive days. Unfortunately, there is no clear guidance for when thrombocytopenia is refractory for a prolonged period of time. The following case describes such a scenario in which consecutive ADAMTS13 activity and inhibitor levels were used to guide the decision to stop treatment with TPE in a patient who failed to recover their platelet count. [ABSTRACT FROM AUTHOR]

Background: Convalescent plasma infusion (CPI) was given to patients with COVID-19 during the early pandemic with mixed therapeutic efficacy. However, the impacts of CPI on the ADAMTS13-von Willebrand factor (VWF) axis and vascular endothelial functions are not known., Objectives: To determine the impacts of CPI on the ADAMTS13-VWF axis and vascular endothelial functions., Methods: Sixty hospitalized patients with COVID-19 were enrolled in the study; 46 received CPI and 14 received no CPI. Plasma ADAMTS13 activity, VWF antigen, endothelial syndecan-1, and soluble thrombomodulin (sTM) were assessed before and 24 hours after treatment., Results: Patients with severe and critical COVID-19 exhibited significantly lower plasma ADAMTS13 activity than the healthy controls. Conversely, these patients showed a significantly increased VWF antigen. This resulted in markedly reduced ratios of ADAMTS13 to VWF in these patients. The levels of plasma ADAMTS13 activity in each patient remained relatively constant throughout hospitalization. Twenty-four hours following CPI, plasma ADAMTS13 activity increased by ∼12% from the baseline in all patients and ∼21% in those who survived. In contrast, plasma levels of VWF antigen varied significantly over time. Patients who died exhibited a significant reduction of plasma VWF antigen from the baseline 24 hours following CPI, whereas those who survived did not. Furthermore, patients with severe and critical COVID-19 showed significantly elevated plasma levels of syndecan-1 and sTM, similar to those found in patients with immune thrombotic thrombocytopenic purpura. Both syndecan-1 and sTM levels were significantly reduced 24 hours following CPI., Conclusion: Our results demonstrate the relative deficiency of plasma ADAMTS13 activity and endothelial damage in patients with severe and critical COVID-19, which could be modestly improved following CPI therapy., (© 2022 The Author(s).)

ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) is also called von Willebrand factor (VWF) cleaving protease (VWFCP). ADAMTS13 acts to cleave VWF multimers and thus reduce plasma VWF activity. In the absence of ADAMTS13 (i.e., in thrombotic thrombocytopenia purpura, TTP), plasma VWF can accumulate, in particular as "ultra-large" VWF multimers, and this can lead to thrombosis. Relative deficiencies in ADAMTS13 can also occur in a variety of other conditions, including secondary thrombotic microangiopathies (TMA). Of contemporary interest, COVID-19 (coronavirus disease 2019) may also be associated with relative reduction of ADAMTS13 and also pathological accumulation of VWF, with this likely contributing to the thrombosis risk seen in affected patients. Laboratory testing for ADAMTS13 can assist in the diagnosis of these disorders (i.e., TTP, TMA), as well as in their management, and can be achieved using a variety of assays. This chapter therefore provides an overview of laboratory testing for ADAMTS13 and the value of such testing to assist the diagnosis and management of associated disorders., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Endosulfan, a persistent organic pollutant, is widely used in agriculture as a pesticide. The aim of the present study was to evaluate the blood toxicity of different doses of endosulfan in Wistar rats. The experimental sample was composed of four groups, a control group that did not receive endosulfan and three endosulfan-exposed groups that respectively received 1, 5, or 10 mg/kg/day (doses below LD), of endosulfan for 21 days. The results showed that endosulfan significantly decreased the prothrombin time (PT) and upregulated the activated coagulation factors VIIa, Xa, and XIIIa; thrombin-antithrombin complex (TAT); and P-selectin. Plasma levels of tissue factor (TF) and malondialdehyde (MDA) were increased in the endosulfan groups. The activated partial thromboplastin time (APTT) and the level of activated coagulation factor IXa showed no obvious changes. Immunohistochemical results showed increased expression of von Willebrand factor (vWF) and the inflammatory cytokine interleukin (IL)-1β in the groups exposed to endosulfan. The pathology and electron microscopy results showed impaired vascular tissue accompanied by the exfoliation of endothelial cells and mitochondrial damage in the endosulfan-exposed groups. In summary, our results suggest that endosulfan damages endothelial cells via oxidative stress and the inflammatory response, leading to the release of TF and vWF into the blood. The TF and vWF in the blood may activate extrinsic coagulation factors and platelets, thus triggering the extrinsic coagulation pathway. There were no obvious effects on the intrinsic coagulation pathway. [ABSTRACT FROM AUTHOR]

Introduction: Atherosclerotic cardiovascular disease remains the leading cause of increased morbidity and mortality observed in chronic kidney disease (CKD) patients. Endothelial dysfunction (ED) is thought to be a key initial event in the development of atherosclerosis. The aim of this study was to evaluate the potential role of hemostatic factors in atherosclerosis, thrombosis and cardiovascular complications in patients suffering from chronic renal disease. Methods: The study was conducted on 50 renal patients divided into two groups of equal size. Group 1 consisted of 25 patients with end-stage renal disease (ESRD) on regular hemodialysis. Group 2 consisted of 25 chronic renal disease patients on conservative treatment. Twenty age- and sex-matched healthy subjects were included in the study to serve as a control group. Thrombomodulin (TM), von Willebrand factor (vWF), tissue plasminogen activator (t-PA), plasminogen activator inhibitor (PAI-1) and hsCRP were assessed. High-resolution B-mode ultrasonography of both the common and internal carotid arteries to measure carotid intima media thickness (CIMT) was performed on all subjects. Results: There were highly significant increases in hsCRP, TM, vWF, tPA and PAI-1 in both patient groups compared to the control group (P<0.01 for all except for TM between group 2 and 3 P<0.05) with significant increase in group 1 compared to group 2 (P<0.01). In addition, there was a highly significant increase in CIMT in both patient groups compared to the control group (P<0.01) with a significant increase in group 1 compared to group 2 (P<0.05). The study revealed significant positive correlation of hemostatic factors (TM, vWf, PAI-1 & t- PA) with creatinine, urea, hsCRP & CIMT. Conclusion: CKD patients have increased risk of atherosclerosis as measured by CIMT, which is used as a surrogate marker of early atherosclerosis and has been shown to be a strong predictor of future myocardial infarction and stroke. They have high levels of TM, vWF, tPA, PAI-1 that correlate with kidney function, hsCRP and CIMT. Therefore, these abnormalities in hemostasis may account for the increased risk of atherothrombosis in these patients. The elevated hsCRP levels and their correlation to hemostatic factors and CIMT might provide an important clue to link a systemic marker of inflammation to atherosclerosis. Further research is required to better understand the procoagulant state in patients with CKD. [ABSTRACT FROM AUTHOR]

Mechanical circulatory support (MCS) is an umbrella term describing the various technologies used in both short- and longterm management of patients with either end-stage chronic heart failure (HF) or acute HF. Most often, MCS has emerged as a bridge to transplantation, but more recently it is also used as a destination therapy. Mechanical circulatory support includes left ventricular assist device (LVAD) or bi-ventricular assist device (Bi-VAD). Currently, 2- to 3-year survival in carefully selected patients is much better than with medical therapy. However, MCS therapy is hampered by sometimes life-threatening complications including bleeding and device thrombosis. Von Willebrand factor (vWF) has two major functions in haemostasis. First, it plays a crucial role in platelet-subendothelium adhesion and platelet-platelet interactions (aggregation). Second, it is the carrier of factor VIII (FVIII) in plasma. Von Willebrand factor prolongs FVIII half-time by protecting it from proteolytic degradation. It delivers FVIII to the site of vascular injury thus enhancing haemostatic process. On one hand, high plasma levels of vWF have been associated with an increased risk of thrombosis. On the other, defects or deficiencies of vWF underlie the inherited von Willebrand disease or acquired von Willebrand syndrome. Here we review the pathophysiology of thrombosis and bleeding associated with vWF. [ABSTRACT FROM AUTHOR]

Objective: To lay the foundation for hemophilia A gene therapy and real-time monitoring of human embryonic stem cells differentiation to endothelial cells, endothelial cell-specific promoters were cloned and its specificity was validated. Methods: A stable method of obtaining endothelial cells from umbilical vein was established. Then the expression of endothelial cell specific marker gene v WF and VE-cadherin were verified via RT-PCR and immunofluorescence. Promoter fragments of v WF and VE-cadherin gene were amplified using PCR. The EGFP expression vector originated from p EGFP-N1. The cloned promoters were substituted for the endogenous CMV promoter. The plasmids: pv WF-1, pv WF-2, p VE-1 and p VE-2 were constructed in total, which were then transfected into HUVECs and h ESCs. In order to find the strongest and most specific promoter, the expression of GFP 48 h after transfection were observed and compared. Results: Endothelial cells were isolated from umbilical vein and the markers of their specific genes were detectable. Four plasmids were confirmed by restriction enzyme digestion and sequencing. HUVECs and h ESCs were transfected by the constructed plasmids, and the GFP expression under fluorescence microscopy showed that p VE-2 promoter was of most specificity and transcription activity. Conclusions: An endothelial cell specific promoter with the highest GFP specificity and activity in endothelial cells was successfully cloned. [ABSTRACT FROM AUTHOR]

Platelet signatures indicating differential dysfunction, hyperactivation, aggregation or adhesion are capable of expressing their characters during the journey of a disease process, and can be utilized as cost effective biomarkers with immense clinical value. Type 2 diabetes mellitus (T2DM) is a major lifestyle disease of contemporary world with progression to diabetes associated cardiovascular diseases (DM-CVD). We identified a few potential biomarkers in platelets of T2DM to analyze the thrombotic risk in diabetes subjects by utilizing flow cytometric quantification with different flurochrome conjugated monoclonal antibodies. Our study describes interesting correlations (p < 0.0001) for different clinical parameters of concurrent threat for vessel occlusion and the status of indices like reactive oxygen species, von Willebrand factor and mitochondrial membrane potential using western blotting and fluorescence techniques. Our study involved 32 T2DM, and 31 T2DM - CVD subjects compared to 29 healthy controls without any history of T2DM or CVD. An altered expression of platelet surface markers P-selectin (CD62p) and GpIIb/IIIa (CD 41/61, PAC1) along with changes in the platelet size due to agonist induced activation contributed to the enhanced thrombotic potential in the patients. This work elucidates the prospect of platelet biomarkers as diagnostic tool to predict cardiovascular risk in DM subjects. [ABSTRACT FROM AUTHOR]

A disintegrin and metalloprotease with thrombospondin motifs 13 (ADAMTS13) is a metalloprotease that regulates von Willebrand factor (VWF) function. ADAMTS13-mediated proteolysis is determined by conformational changes in VWF, but also may depend on its own conformational activation. Kinetic analysis of WT ADAMTS13 revealed ~2.5-fold reduced activity compared with ADAMTS13 lacking its C-terminal tail (MDTCS) or its CUB1-2 domains (WTΔCUB1-2), suggesting that the CUB domains naturally limit ADAMTS13 function. Consistent with this suggestion, WT ADAMTS13 activity was enhanced ~2.5-fold by preincubation with either an anti-CUB mAb (20E9) or VWF D4CK (the natural binding partner for the CUB domains). Furthermore, the isolated CUB1-2 domains not only bound MDTCS, but also inhibited activity by up to 2.5-fold. Interestingly, a gain-of-function (GoF) ADAMTS13 spacer domain variant (R568K/F592Y/R660K/Y661F/Y665F) was ~2.5-fold more active than WT ADAMTS13, but could not be further activated by 20E9 mAb or VWF D4CK and was unable to bind or to be inhibited by the CUB1-2 domains, suggesting that the inhibitory effects of the CUB domains involve an interaction with the spacer domain that is disrupted in GoF ADAMTS13. Electron microscopy demonstrated a "closed" conformation of WT ADAMTS13 and suggested a more "open" conformation for GoF ADAMTS13. The cryptic spacer domain epitope revealed by conformational unfolding also represents the core antigenic target for autoantibodies in thrombotic thrombocytopenic purpura. We propose that ADAMTS13 circulates in a closed conformation, which is maintained by a CUB-spacer domain binding interaction. ADAMTS13 becomes conformationally activated on demand through interaction of its C-terminal CUB domains with VWF, making it susceptible to immune recognition. [ABSTRACT FROM AUTHOR]

Recombinant ADAMTS13 is currently undergoing clinical trials as a treatment for hereditary thrombotic thrombocytopenic purpura, a lethal microvascular condition resulting from ADAMTS13 deficiency. Preclinical studies have also demonstrated its efficacy in treating arterial thrombosis and inflammation without causing bleeding, suggesting that recombinant ADAMTS13 may have broad applicability as an antithrombotic agent. Despite this progress, we currently do not understand the mechanisms that regulate ADAMTS13 activity in vivo. ADAMTS13 evades canonical means of protease regulation because it is secreted as an active enzyme and has a long half-life in circulation, suggesting that it is not inhibited by natural protease inhibitors. Although shear can spatially and temporally activate von Willebrand factor to capture circulating platelets, it is also required for cleavage by ADAMTS13. Therefore, spatial and temporal regulation of ADAMTS13 activity may be required to stabilize von Willebrand factor-platelet strings at sites of vascular injury. This review outlines potential mechanisms that regulate ADAMTS13 in vivo including shear-dependency, local inactivation, and biochemical and structural regulation of substrate binding. Recently published structural data of ADAMTS13 is discussed, which may help to generate novel hypotheses for future research., (© 2022 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals LLC on behalf of International Society on Thrombosis and Haemostasis.)

Von Willebrand factor (VWF) is a glycoprotein synthesized and secreted by vascular endothelial cells and megakaryocytes, found on plasma surface, endothelial cells, and α-granule of platelets. VWF can be interacted with collagen and platelet membrane glycoproteins GPIb and GPIb-IIa and play an important role in platelet adhesion and aggregation. Growing research evidence suggests that VWF also mediates the prevention or protesting of hepatocellular carcinoma (HCC) in chronic hepatitis B (CHB) patients from several clinical studies. While the mechanism of VWF in HCC protection or protest is still unclear, further study is required. This article aims to rationalize the role of VWF in the development of HCC, and the functional domain of VWF in cancer as well as cross-talking with platelets and miRNAs. This article also looks forward to the future development and challenges of VWF research., Competing Interests: The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (© The Author(s), 2022.)

Objectives: This study was aimed at assessing the clinical presentations and laboratory findings among patients diagnosed with vWD at a Saudi tertiary care unit., Methods: This retrospective study included 189 patients with vWD who were followed up in our unit over 4 years. Clinical and laboratory data were collected and analyzed in SPSS., Results: The median age of the study cohort was 30 years (range 11 months-56 years). The cohort had a female preponderance, with 32.30% males and 66.70% females. Bleeding from different sites was observed, mostly from the joints and muscles (23.90%), followed by the mucus membranes (14.60%), genitourinary areas (7.70%), ecchymoses (2.80%), and gastrointestinal areas (2.80%). A total of 48% of participants presented with more than one type of bleeding. A total of 105 (58.01%) participants had type 1; 29 (16.02%) had type 2; and 47 (25.96%) had type 3 vWD. Blood tests indicated the following mean value: hemoglobin, 116 ± 25.60 gm/L; ferritin, 75.80 ± 166.80 μg/L (median 28.5); vWAg, 0.40 ± 0.27IU/ml; and vWD:RCo, 0.32 ± 0.20IU/dL. The partial thromboplastin time was prolonged in 49.20% and normal in 50.80% of participants. Platelet function analysis values were prolonged in 92.90% and normal in 7.10% of participants. Comparative analysis of the O-type and non-O blood type showed that blood type O was significantly correlated with factor VIII (p-value = 0.013), vWF:RCo (p-value = 0.004), and vWF:Ag (p-value = 0.019)., Conclusion: Joint and muscle bleeds were the most common clinical presentations in our cohort. Although type 1 vWD was most prevalent in our cohort, we observed a comparatively higher prevalence of type 3, possibly because of ethnic differences or referral bias. We found a significant difference between O and non-O blood type regarding FVIII and vWF:Ag, and observed a more pronounced difference for vWD activity measuresd by vWF:RCo with blood type O being the systematic factor., (© 2022 [The Author/The Authors].)

The aim of this study was to evaluate the effect of hormone therapy (HT) in the endothelial function of 46,XY disorders of sexual development (DSD) patients with female phenotype. Biochemical and ultrasound measurements were performed in 20 patients at initiation of oral 2 mg 17β-estradiol/1 mg norethisterone acetate, and after 6 months of therapy. Lipid profile, including total cholesterol (TC), LDL, HDL, triglycerides (TG) and Atherogenic Index of Plasma (AIP), as well as levels of VE-Cadherin, E-Selectin, Thrombomodulin and vWf were determined. Ultrasonographic examinations included evaluation of flow-mediated dilatation (FMD) and measurement of Carotid and Femoral Intima Media Thickness (IMT). HT raised HDL (35.4 mg/dl versus 40.1 mg/dl, p = 0.019) while lowering TG (166 mg/dl versus 109 mg/dl, p = 0.026) and AIP (0.24 versus 0.04, p = 0.007). No changes were noted in TC and LDL (215.7 mg/dl versus 192.25 mg/dl and 87.46 mg/dl versus 76.35 mg/dl, respectively). There was significant reduction of VE-Cadherin (4.05 ng/ml versus 2.20 ng/ml, p = 0.002) and E-selectin (73.98 ng/ml versus 56.73 ng/ml, p = 0.004). No change was observed in Thrombomodulin and vWf (11.76 ng/ml versus 13.90 ng/ml and 80.75% versus 79.55%, respectively). FMD improved significantly (5.4% versus 8.15%, p = 0.003), while only carotid bulb IMT decreased significantly (0.65 mm versus 0.60 mm, p = 0.018). Overall, HT was found to improve biochemical and ultrasound markers of endothelial function in 46,XY DSD patients with female phenotype. [ABSTRACT FROM AUTHOR]

Spreading of melanoma is associated with efficient extravasation of circulating tumor cells from the vascular system into distant target organs. This process is accompanied and supported by proinflammatory and procoagulatory conditions. In this study, we analysed the ability of human melanoma cell lines to activate endothelial cells (ECs) in vitro. Some melanoma cells, that is, MV3, were shown to trigger an prompt calcium-fluxdependent, procoagulatory endothelial response that was accompanied by luminal release of ultra-large von Willebrand factor (ULVWF) fibres that were immobilized to the endothelial surface layer. In contrast to MV3-derived supernatant, prolonged treatment of ECs with WM9-derived supernatant mediated a pronounced activation of nuclear factor kappa B (NFκB). NFκB activation in ECs was dependent on both IL-1α and IL-1β secreted from melanoma cells. Melanoma-derived IL-1 mediated an upregulation of proinflammatory cytokines IL-6 and IL-8, the intercellular adhesion molecule-1 (ICAM-1), the vascular cell adhesion molecule-1 (VCAM-1) and the procoagulatory tissue factor (TF) in ECs. Our data show that melanoma cells activate ECs either directly and within seconds or by an IL-1-mediated NFκB activation. Both pathways of EC activation convert the regular repressive function of ECs on inflammation and coagulation to a proinflammatory and procoagulatory surface that supports tumor progression. [ABSTRACT FROM AUTHOR]

This systematic review was designed to summarize the reported valid quantitative evidence on the association between use of von Willebrand factor ( VWF)-containing Factor VIII ( FVIII) concentrates and successful immune tolerance induction ( ITI) in patients with severe haemophilia A. The primary outcome was successful ITI; secondary outcomes were time to success, complications of the inhibitor or ITI and relapse of the inhibitor. A systematic literature search identified 26 randomized controlled trials, registries and cohort studies, evaluating a total of 1284 patients. For a pooled meta-analysis, 13 studies evaluating 382 patients were included. Due to incomplete data we were not able to assign pre- ITI risk categories to all patients for risk factor analysis. The meta-analysis did not demonstrate a difference in the proportion of patients with successful inhibitor eradication between those treated with VWF-containing products and those treated with FVIII concentrates devoid of VWF (relative risk [ RR] 0·70 (95% confidence interval [ CI] 0·52-0·89) and 0·84 (95% CI 0·75-0·93) respectively). Bleeding rate during ITI ranged from 0·00 to 0·85 bleeding episodes per year. The proportion of patients with a relapse of the inhibitor (range 0-20%) was mentioned in four studies that were included in the meta-analysis. The results of this systematic review do not support the idea of a positive effect of VWF-containing products in ITI. [ABSTRACT FROM AUTHOR]

Molecular linkages between von Willebrand factor (VWF) and the alternative complement pathway (AP) have recently been discovered. Endothelial cell (EC)-anchored ultralarge (UL) VWF multimeric strings function as an activating surface for the AP. C3 (in active C3b form) binds to the EC-anchored ULVWF strings, and promotes the assembly of C3bBb (C3 convertase) and C3bBbC3b (C5 convertase). These linkages may help to explain enigmatic clinical problems related to thrombotic microangiopathies, including some cases of refractory thrombotic thrombocytopenic purpura (TTP), TTP associated with onlymild-modest deficiencies of ADAMTS-13, the provocation (or exacerbation) of acute episodes in patients with the atypical hemolytic uremic syndrome, and thrombosis in paroxysmal nocturnal hemoglobinuria. Recent experiments have also demonstrated that complement factor H performs a dual role: participating in regulation of the AP by binding to EC-anchored ULVWF strings; and functioning as a reductase to decrease the size of soluble VWF multimers. [ABSTRACT FROM AUTHOR]

Objective The present study attempts to compare the immunohistochemistry (IHC) of von Willebrand factor (vWf) , endothelial cadherin, Caveolin and endothelial Nitric Oxide Synthase (eNOS) in VasoView Endoscopic Vein Harvesting (EVH) versus traditional Open Vein Harvesting (OVH) techniques for Coronary Artery Bypass Graft (CABG) Surgery performed in Javad al Aemeh Hospital of Mashhad, Iran in 2013,. Methods and materials Forty-seven patients were scheduled for CABG (30 EVH and 17 OVH) among whom patients with relatively same gender and similar age were selected. Three separate two cm vein samples were harvested from each patient's saphenous vein. Each portion was collected from distal, middle and proximal zones of the saphenous vein. The tissues were deparaffinized, and antigen retrieval was done using EZ-retriever followed by an immunohistochemistry evaluation with vWf, e-cadherin, Caveolin and eNOS. In addition, demographic questioner as of Lipid profile, FBS, BMI, and cardiovascular risk factors were collected. Data analyses, including parametric and nonparametric tests were undertaken using the SPSS 16 software. A P value < 0.05 was regarded as statistically significant. Results The mean age of the EVH and OVH groups were 63.76 ± 9.51 and 63.63 ± 8.31 years respectively with no significant difference between them (p = 0.989). In addition, there was no great difference between the EVH and OVH groups in lipid profile, DM, HTN, smoking history, CVA, and valvular dysfunction (P > 0.05). Qualitative report of vWf, e-cadherin, Caveolin and eNOS reveals no significant difference between the EVH and OVH (P > 0.05). Conclusion This study indicates that VasoView EVH technique causes no endothelial damage in comparison with OVH. This study could be a molecular confirmation for the innocuous of EVH technique. [ABSTRACT FROM AUTHOR]

Pharmacologically, vasoactive agents targeting endothelial and/or smooth muscle cells (SMC) are known to cause acute drug-induced vascular injury (DIVI) and the resulting pathology is due to endothelial cell (EC) perturbation, activation, and/or injury. Alteration in EC structure and/or function may be a critical event in vascular injury and, therefore, evaluation of the circulatory kinetic profile and secretory pattern of EC-specific proteins such as VWF and VWFpp could serve as acute vascular injury biomarkers. In rat and dog models of DIVI, this profile was determined using pharmacologically diverse agents associated with functional stimulation/perturbation (DDAVP), pathological activation (lipopolysaccharide [LPS]/endotoxin), and structural damage (fenoldopam [FD], dopamine [DA], and potassium channel opener (PCO) ZD6169). In rats, FD caused moderate DIVI and time-related increase in plasma VWF levels ∼33% while in control rats VWF increased ∼5%. In dogs, VWF levels transiently increased ∼30% when there was morphologic evidence of DIVI by DA or ZD6169. However, in dogs, VWFpp increased >60-fold (LPS) and >6-fold (DDAVP), respectively. This was in comparison to smaller dynamic 1.38-fold (LPS) and 0.54-fold (DDAVP) increases seen in plasma VWF. Furthermore, DA was associated with a dose-dependent increase in plasma VWFpp. In summary, VWF and VWFpp can discriminate between physiological and pathological perturbation, activation, and injury to ECs. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: Through binding to von Willebrand factor (VWF), platelet glycoprotein (GP) Ibα, the major ligand-binding subunit of the GPIb-IX-V complex, initiates platelet adhesion and aggregation in response to exposed VWF or elevated fluid-shear stress. There is little data regarding non-human primate platelet GPIbα. This study cloned and characterized rhesus monkey (Macaca Mullatta) platelet GPIbα. Materials and Methods: DNAMAN software was used for sequence analysis and alignment. N/O-glycosylation sites and 3-D structure modelling were predicted by online OGPET v1.0, NetOGlyc 1.0 Server and SWISS-MODEL, respectively. Platelet function was evaluated by ADP- or ristocetin-induced platelet aggregation. Results: Rhesus monkey GPIbα contains 2,268 nucleotides with an open reading frame encoding 755 amino acids. Rhesus monkey GPIbα nucleotide and protein sequences share 93.27% and 89.20% homology respectively, with human. Sequences encoding the leucine-rich repeats of rhesus monkey GPIbα share strong similarity with human, whereas PEST sequences and N/O-glycosylated residues vary. The GPIbα−binding residues for thrombin, filamin A and 14-3-3ζ are highly conserved between rhesus monkey and human. Platelet function analysis revealed monkey and human platelets respond similarly to ADP, but rhesus monkey platelets failed to respond to low doses of ristocetin where human platelets achieved 76% aggregation. However, monkey platelets aggregated in response to higher ristocetin doses. Conclusions: Monkey GPIbα shares strong homology with human GPIbα, however there are some differences in rhesus monkey platelet activation through GPIbα engagement, which need to be considered when using rhesus monkey platelet to investigate platelet GPIbα function. [Copyright &y& Elsevier]

Abstract: Vascular inflammation plays a key role in the pathogenesis of atherosclerosis. The first step in vascular inflammation is endothelial exocytosis, in which endothelial granules fuse with the plasma membrane, releasing prothrombotic and proinflammatory messenger molecules. The development of cell culture models to study endothelial exocytosis has been challenging because the factors that modulate exocytosis in vitro are not well understood. Here we report a method for studying endothelial exocytosis that optimizes extracellular matrix components, cell density, and duration of culture. Human umbilical vein endothelial cells plated on collagen I-coated plates and cultured in the confluent state for 7–12days in low-serum medium showed robust secretion of von Willebrand factor when stimulated with various agonists. This exocytosis assay is rapid and applicable to high-throughput screening. [Copyright &y& Elsevier]

Abstract: In this study, we investigated the molecular basis of two unrelated Chinese patients with hemostatic disorders. The proband of the first family had severe hemophilia A (HA) coexisting with type 1 von Willebrand disease (VWD) and the proband of the second family had type 2N VWD. Both probands had similar phenotypes, which included joint and mucosal bleeding, very low factor VIII (FVIII) activity (FVIII:C), and moderate reductions in VWF antigen (VWF:Ag) and VWF ristocetin cofactor activity (VWF:Rco), as well as a normal multimeric pattern. One FVIII mutation and three VWF mutations were identified: FVIII p.R446* and VWF heterozygous p.E216K mutations were detected in proband 1 and compound heterozygosity of VWF mutations (p.R816W and c.1911delC) in proband 2. Transient expression studies in HEK293T cells proved that R816W mutation abolished the binding of FVIII to VWF and slightly impaired protein synthesis and secretion; 1911delC mutation mainly impaired VWF protein synthesis and secretion. These results provided insight into the possible pathogenic mechanism of type 2N VWD in Chinese patients carrying these mutations. [Copyright &y& Elsevier]

Background: Thrombosis, attributed to atherosclerosis, is the leading cause of morbidity and mortality in patients with diabetes mellitus. Pathogenesis of atherosclerosis in diabetes mellitus is not entirely clear and conventional risk factors such as smoking, obesity, blood pressure and serum lipids fail to explain fully this excess risk. We set out a cross-sectional study to determine the risk factors and patient attributes that may predispose type 2 Diabetes Mellitus (T2DM) patients to be in hypercoagulable state. Methods: A structured questionnaire was used to capture Age, Sex, duration of diabetes mellitus and knowledge on T2DM of study participants. Body weight and height were also measured and BMI calculated.VWF, Cholesterol, and Glycated haemoglobin were measured in 213 T2DM patients. VWF was used as a proxy marker for hypercoagulability in T2DM patients. Participants with VWF of >2.0 IU/ml plasma concentration were regarded to be in hypercoagulable state. Results: Chi-square analysis revealed that hypercoagulability in T2DM patients was associated with Age (P=0.001), Sex (P=0.000), glycaemic control (P=0.003), duration of diabetes (P=0.000), BMI (P=0.000) and knowledge on type 2 diabetes mellitus (P=0.001). In multivariate analysis after adjusting for confounders, knowledge on T2DM was not independently associated with hypercoagulability AOR being 1.00(CI 95% 0.80-2.20). However Age, sex, glycaemic control, duration of diabetes and BMI were found to be independent risk factors for hypercoagulability in T2DM patients giving AOR of 1.45(95%CI[1.19-3.16]), 4.42 (95% CI [2.77-10.63]), 6.12 (95% CI 2.27-8.36) 5.28(CI 95% 3.01-8.21) and 1.05(95% CI 0.75-2.86) respectively. Conclusion: Age, Sex, poor glycaemic control, duration of diabetes and obesity should be taken into account in the management of T2DM patients as these variables are independent risk factors of hypercoagulability in T2DM patients. [ABSTRACT FROM AUTHOR]

Background: Von Willebrand factor antigen (VWF) is a non-invasive marker for clinically significant portal hypertension (HVPG≥10 mmHg) and confers HVPG-independent prognostic information. While quantification of increased VWF-levels is not relevant in the context of von Willebrand disease, highly elevated VWF may be of clinical significance in ACLD. Thus, we have modified our analytical approach to quantify very high VWF-levels (i.e.,>420%) and investigated their prognostic value., Methods: Patients undergoing HVPG-measurement at the Vienna Hepatic Hemodynamic Lab with evidence of ACLD and information on VWF were considered. Clinical stages (CS) were defined as follows: Probable compensated ACLD (cACLD): LSM≥10kPa&HVPG<6 mmHg; 0: cACLD&6-9 mmHg; 1: cACLD&HVPG≥10 mmHg; 2: bleeding; 3: non-bleeding decompensation; 4: ≥2 decompensations., Results: 124 (16%) of 793 patients had VWF>420%. The proportion of VWF>420% increased with disease severity (probable cACLD-0: 5(4%) vs. 1: 22(10%) vs. 2-4: 97(23%),p ≤ 0.001) as well as across HVPG (<6mmHg: 1(2%) vs. 6-9: 6(6%) vs. 10-15: 17(9%) vs. ≥16: 100(22%),p ≤ 0.001) and MELD (<10: 17(6%) vs. 10-14: 27(10%) vs. ≥15: 79(32%),p ≤ 0.001) strata. In patients with VWF>420%, median VWF was 533 (IQR:466-611)% and VWF was unrelated to HVPG (Spearman's ρ=0.139,p = 0.123), but showed direct correlations of weak/moderate strength with MELD (ρ=0.336,p < 0.001) and CRP (ρ=0.286,p = 0.001). In the subgroup with VWF>420%, VWF was predictive of decompensation/liver-related mortality (VWF per 10%; hazard ratio (HR): 1.02(95% confidence interval (95%CI): 1.01-1.04),p = 0.008, even after adjusting for other factors (VWF per 10%; adjusted HR: 1.02(95%CI: 1.00-1.05),p = 0.031)., Conclusion: The proportion of patients with substantially elevated VWF values steadily increases with disease progression. While VWF is not reflective of HVPG in these patients, it is correlated with hepatic dysfunction and systemic inflammation. Importantly, quantification of high values provides prognostic information., Competing Interests: Declaration of Competing Interest K.P., L.B., R.J.N., J.W., R.P., D.P., P.S., A.F.S., and P.Q. have nothing to declare., (Copyright © 2022. Published by Elsevier Ltd.)

Water-soluble tomato concentrate (WSTC), extracted from mature tomatoes, is the first health product in Europe that has been approved "to help maintain normal platelet activity to maintain healthy blood flow." We hypothesized that WSTC might exert an influence on blood flow shear stress-induced platelet aggregation (SIPA) and in turn maintains healthy blood flow. We used a microfluidic system to measure the effects of WSTC on SIPA in vitro. We also used the strenuous exercise rat model and the κ-carrageenan-induced rat tail thrombosis model to demonstrate the effects of WSTC on blood flow. WSTC significantly inhibited platelet aggregation at pathological high shear rate of 4,000 s -1 and 8,000 s -1 in vitro ( P < 0.01). WSTC reduced the platelet adhesion rate and increased the rolling speed of platelets by inhibiting binding to Von Willebrand Factor (vWF) ( P < 0.01). WSTC reduced the platelet adhesion rate and increased the rolling speed of platelets by inhibiting binding to Von Willebrand Factor (vWF) ( P < 0.05 or P < 0.01). The oral administration of WSTC for 4 weeks in strenuous exercise rats alleviated hyper-reactivity of the platelets and led to a significant reduction in the plasma levels of catecholamine and IL-6. WSTC treatment also led to a reduction in black tail length, reduced blood flow pulse index (PI) and vascular resistance index (RI), and ameliorated local microcirculation perfusion in a rat model of thrombosis. WSTC exerted obvious inhibitory effects on the platelet aggregation induced by shear flow and alleviated the blood flow and microcirculation abnormities induced by an inflammatory reaction., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liu, Xiao, Wang, Wang, Liu, Sun, Zhang, Yin, Liao, You and Zhang.)

Von Willebrand factor (VWF) is a complex glycoprotein found in plasma, composed of disulfide-bond-linked multimers with apparent molecular weights between 500 kDa and 20,000 kDa. After release of VWF from storage granules, it is cleaved in flowing blood by the specific metalloproteinase ADAMTS13, resulting in a highly characteristic cleavage pattern and structure. As the structure of VWF multimers determines diagnosis of von Willebrand disease, which has different sub-types with different multimer- and cleavage patterns, VWF analysis is performed using low-resolution horizontal SDS-agarose gel electrophoresis. However, almost every laboratory uses a different protocol, and all experimental details are rarely, if at all, described. Therefore, the results from similar methods may be substantially different. Here, we present a detailed description of a validated VWF multimer method that we have developed. It has been successfully used for over more than 20 years in quality control of recombinant and plasma-derived VWF drug products, and in preclinical and clinical studies with VWF drug candidates. As most of the published methods, it enables visualization of VWF multimers separated by electrophoresis by immunostaining with a polyclonal anti-human VWF antibody followed by a secondary antibody coupled to alkaline phosphatase. VWF appears as a series of regularly spaced bands on the low and middle molecular weight range of the gel, with an unresolved zone in the high molecular weight (HMW) range, where ultra-large multimers are found. An example is shown below. This low-resolution agarose gel electrophoresis allows the determination of the number of VWF multimers with high reproducibility. Graphical abstract: Example of electrophoretic analysis of multimer structure of four batches of a recombinant VWF drug substance., Competing Interests: Competing interestsHG, PLT and MS are employees of Baxalta Innovations GmbH, a Takeda company, PLT holds relevant Takeda patents and Takeda stock., (Copyright © 2022 The Authors; exclusive licensee Bio-protocol LLC.)

Human induced pluripotent stem cell-derived mesenchymal stem cells (iMSCs) have been believed to be a promising alternative for the stem cell transplantation therapy. The exosomes (Exo) from iMSCs play an important role in several kinds of life activities. The role of exosomes from iMSCs in severe acute pancreatitis (SAP) induced myocardial injury (MI) has not been investigated. The Exo were isolated from iMSCs through differential centrifugation method. The SAP rat model was established with 5% sodium taurocholate injection into the distal end of the bilepancreatic duct. RT-PCR and western blotting were used to measure related gene expression. Masson trichrome and Sirius Red stainings were used to evaluate MI injury. Cardiac function was detected through cardiac ultrasound.Exo promoted cell viability through activating Akt/nuclear factor E2 related factors 2 (Nrf2)/heme oxygenase 1 (HO-1) signaling pathway in vitro . Exo improved MI induced by SAP through activating Akt/Nrf2/HO-1 signaling pathway. Exo improved cardiac function, and suppressed oxidative status in the SAP model. Exo increased the expression of von Willebrand Factor (vWF) and vascular endothelial growth factor (VEGF) through activating Nrf2/HO-1 signaling pathway. Our data indicated that the Exo from iMSCs could improve MI caused by SAP through activating Nrf2/HO-1 axis. These findings firstly unfold the potential application of Exo from iMSCs in treating MI induced by SAP. Abbreviations: LVEF: Left ventricular ejection fraction; LVFS: left ventricular fractional shorten; LVDd: left ventricular end-diastolic diameter; LVDs: left ventricular end-systolic diameter; MI: Myocardial infarction; MSCs: Mesenchymal stem cells; iPSCs: Human-induced pluripotent stem cells; SAP: Severe acute pancreatitis; iMSCs: iPSCs derived VEGF: MSCs; vascular endothelial growth factor; Nrf2: Nuclear factor erythroid 2-related factor; RT-PCR: Real-time polymerase chain reaction; HE: Hematoxylin-eosin; MODS: Multiple organ dysfunction syndrome; PI3K: Phosphatidylinositol 3-kinase; SOD: Superoxide dismutase; FBS: Fetal bovine serum; ECL: Enhanced chemiluminescence; IHC: Immunohistochemistry.

The metastasis and angiogenesis of breast cancer has always been a difficult problem for treatment. It has recently been discovered that Von Willebrand Factor (vWF), in addition to hemostasis, also plays a role in tumor metastasis and angiogenesis. We noticed that besides endothelial cells, breast cancer cells (MDA-MB-231 and MCF-7) could also express vWF. In vitro experiments showed that knocking down vWF inhibited breast cancer cell metastasis. And we found that overexpression of vWF significantly promoted VEGF-A-dependent vascular proliferation in vitro by activating the PI3K/Akt signaling pathway. Further studies indicated that inhibition of PI3K/Akt signaling pathway up-regulated the expression of miR-205-5p, and miR-205-5p could bind to the 3′UTR region of VEGF-A to hinder the production of VEGF-A. Furthermore, when a spontaneous lung metastasis model was established in Balb/c female mice, knockdown of vWF in 4 T1 cells resulted in a decrease in tumor blood vessel density and effectively inhibited lung metastasis, accompanied by a decrease in the expression level of VEGF-A and an increase in the expression level of miR-205-5p. In summary, our findings provide experimental evidence that overexpression of vWF in breast cancer cells down-regulates the expression of miR-205-5p and up-regulates the expression of VEGF-A through the PI3K/Akt signaling pathway, thereby promoting tumor angiogenesis and metastasis. • Knockdown of vWF inhibits the metastasis of breast cancer cells. • Overexpression of vWF promotes VEGF-A-dependent angiogenesis. • vWF regulates angiogenesis through PI3K/Akt/miR-205-5p/VEGF-A signaling pathway. [ABSTRACT FROM AUTHOR]

An increasing number of mutations and polymorphisms have been identified in the von Willebrand factor (VWF) gene of patients with von Willebrand disease (VWD). Most of the sequence alterations are within exon 28, duplicated in the VWF pseudogene on chromosome 22. Genetic recombination causing the gene conversion between the VWF gene and its pseudogene is associated with multiple substitutions in the VWF gene and with VWD. In the present study, VWF gene exon 28 was analyzed in 33 patients with VWD by DNA sequencing. A total of 73% of the patients were heterozygous for p.D1472H, p.V1485L, p.1500A, p.1501F, p.L1503P, and p.S1506L single-nucleotide polymorphisms. Family analysis revealed that the gene conversion occurred between the VWF gene and its pseudogene in 3 patients. Case-control association analysis by Haploview 4.2 did not show an association between the haplotype and VWD. In conclusion, a common exon 28 haplotype in the Turkish population, which might have arisen from the gene conversion events in the founder population, was identified. [ABSTRACT FROM PUBLISHER]

Highlights: [•] Like deafness due to noise, patients with hand arm vibration-syndrome (HAVS) present with numbness due to vibratory impact. [•] Using conventional electrophysiology and quantitative sensory testing, the findings of the present study are consistent with the concept of a sensory damage related to the frequencies of the vibrating devices employed. [•] The frequency of occurrence of a carpal tunnel syndrome in HAVS was not increased. [ABSTRACT FROM AUTHOR]