Von Willebrand Factor Degradation Research Articles

In vitro study on device-induced damage to blood cellular components and degradation of von Willebrand factor in a CentriMag pump-assisted circulation.

High mechanical shear stress (HMSS) generated by blood pumps during mechanical circulatory support induces blood damage (or function alteration) not only of blood cell components but also of plasma proteins. In the present study, fresh, healthy human blood was used to prime a blood circuit assisted by a CentriMag centrifugal pump at a flow rate of 4.5 L/min under three pump pressure heads (75, 150, and 350 mm Hg) for 4 h. Blood samples were collected for analyses of plasma-free hemoglobin (PFH), von Willebrand factor (VWF) degradation and platelet glycoprotein (GP) IIb/IIIa receptor shedding. The extent of all investigated aspects of blood damage increased with increasing cross-pump pressure and duration. Loss of high-molecular-weight multimers (HMWM)-VWF in Loop 2 and Loop 3 significantly increased after 2 h. PFH, loss of HMWM-VWF, and platelet GPIIb/IIIa receptor shedding showed a good linear correlation with mean shear stress corresponding to the three pump pressure heads. HMSS could damage red blood cells, cause pathological VWF degradation, and induce platelet activation and platelet receptor shedding. Different blood components can be damaged to different degrees by HMSS; VWF and VWF-enhanced platelet activation may be more susceptible to HMSS.

Improving our understanding on the clinical role of plasmin-mediated von Willebrand factor degradation.

Von Willebrand factor (VWF) plays a pivotal role in primary hemostasis. A Disintegrin And Metalloproteinase with a ThromboSpondin type 1 motif, member 13 (ADAMTS13) is primarily responsible for cleaving ultra-large VWF multimers into smaller, less adhesive forms. However, plasmin has also been shown to cleave VWF multimers. This proteolytic cleavage of VWF results in a decreased multimer size and, hence, a lower VWF activity. This review aims to present a comprehensive overview of the involvement of plasmin-mediated VWF proteolysis in (micro)thrombosis. Plasmin-mediated VWF proteolysis has been suggested to play a role in various pathologies involving microthrombosis in combination with an imbalance in VWF antigen levels and ADAMTS13 activity, as well as activation of the fibrinolytic system, but quantitative assays to demonstrate this were lacking. Recently, a VHH-based bioassay was developed designed specifically to quantify plasmin-cleaved VWF (cVWF). The novel ELISA assay holds significant promise for gaining further insights into the clinical relevance of plasmin-mediated VWF proteolysis in several pathologies. Furthermore, local plasmin activation at the site of microthrombosis has been shown to be a promising treatment strategy by degrading VWF-rich microthrombi. Plasmin-mediated proteolysis of VWF is observed during microthrombosis; however, it remains unclear whether it impacts disease severity. A novel ELISA method to detect cVWF will improve our understanding of the clinical role of plasmin-mediated VWF degradation.

(1069) - Impact of a Novel Accessory for Left Ventricular Assist Devices on Hemolysis and Degradation of Von Willebrand Factor In Vitro

(1069) - Impact of a Novel Accessory for Left Ventricular Assist Devices on Hemolysis and Degradation of Von Willebrand Factor In Vitro

Shear Forces Induced Platelet Clearance Is a New Mechanism of Thrombocytopenia.

Thrombocytopenia has been consistently described in patients with extracorporeal membrane oxygenation (ECMO) and associated with poor outcome. However, the prevalence and underlying mechanisms remain largely unknown, and a device-related role of ECMO in thrombocytopenia has been hypothesized. This study aims to investigate the mechanisms underlying thrombocytopenia in ECMO patients. In a prospective cohort of 107 ECMO patients, we investigated platelet count, functions, and glycoprotein shedding. In an ex vivo mock circulatory ECMO loop, we assessed platelet responses and VWF (von Willebrand factor)-GP Ibα (glycoprotein Ibα) interactions at low- and high-flow rates, in the presence or absence of red blood cells. The clearance of human platelets subjected or not to ex vivo perfusion was studied using an in vivo transfusion model in NOD/SCID (nonobese diabetic/severe combined Immunodeficient) mice. In ECMO patients, we observed a time-dependent decrease in platelet count starting 1 hour after device onset, with a mean drop of 7%, 35%, and 41% at 1, 24, and 48 hours post-ECMO initiation (P=0.00013, P<0.0001, and P<0.0001, respectively), regardless of the type of ECMO. This drop in platelet count was associated with a decrease in platelet GP Ibα expression (before: 47.8±9.1 versus 24 hours post-ECMO: 42.3±8.9 mean fluorescence intensity; P=0.002) and an increase in soluble GP Ibα plasma levels (before: 5.6±3.3 versus 24 hours post-ECMO: 10.8±4.1 µg/mL; P<0.0001). GP Ibα shedding was also observed ex vivo and was unaffected by (1) red blood cells, (2) the coagulation potential, (3) an antibody blocking VWF-GP Ibα interaction, (4) an antibody limiting VWF degradation, and (5) supraphysiological VWF plasma concentrations. In contrast, GP Ibα shedding was dependent on rheological conditions, with a 2.8-fold increase at high- versus low-flow rates. Platelets perfused at high-flow rates before being transfused to immunodeficient mice were eliminated faster in vivo with an accelerated clearance of GP Ibα-negative versus GP Ibα-positive platelets. ECMO-associated shear forces induce GP Ibα shedding and thrombocytopenia due to faster clearance of GP Ibα-negative platelets. Inhibiting GP Ibα shedding could represent an approach to reduce thrombocytopenia during ECMO.

Accelerated Hemocompatibility Testing of Rotary Blood Pumps.

Ex vivo hemocompatibility testing is a vital element of preclinical assessment for blood-contacting medical devices. Current approaches are resource intensive; thus, we investigated the feasibility of accelerating hemocompatibility testing by standardizing the number of pump exposures in loops of various sizes. Three identical blood loops were constructed, each with a custom-molded reservoir able to facilitate large-volume expansion. Using the HVAD rotary blood pump operating at 5 L·min-1 and 100 mmHg, three test volumes (80, 160, and 320 ml) were circulated for 4000 pump exposures. Blood sampling was performed at individualized intervals every one-sixth of total duration for the assessment of hemolysis and von Willebrand Factor (vWF) degradation. While steady increases in hemolysis (~24 mg·dl-1) were identified in all tests at completion, loop volume was not a primary discriminator. The normalized index of hemolysis did not vary significantly between loops (4.2-4.9 mg·100 L-1). vWF degradation progressively occurred with duration of testing to a similar extent under all conditions. These data support an accelerated approach to preclinical assessment of ex vivo blood damage. Adopting this approach enables: enhanced efficiency for rapid prototyping; reduced ex vivo blood aging, and; greater utility of blood, which is presently limited if 450 ml loops are desired.

Coagulation factor VIII regulates von Willebrand factor homeostasis invivo

BackgroundCoagulation factor VIII (FVIII) and von Willebrand factor (VWF) circulate as a noncovalent complex, but each has its distinct functions. Binding of FVIII to VWF results in a prolongation of FVIII’s half-life in circulation and modulates FVIII’s immunogenicity during hemophilia therapy. However, the biological effect of FVIII and VWF interaction on VWF homeostasis is not fully understood. ObjectivesTo determine the effect of FVIII in VWF proteolysis and homeostasis in vivo. MethodsMouse models, recombinant FVIII infusion, and patients with hemophilia A on a high dose FVIII for immune tolerance induction therapy or emicizumab for bleeding symptoms were included to address this question. ResultsAn intravenous infusion of a recombinant B-domain less FVIII (BDD-FVIII) (40 and 160 μg/kg) into wild-type mice significantly reduced plasma VWF multimer sizes and its antigen levels; an infusion of a high but not low dose of BDD-FVIII into Adamts13+/- and Adamts13-/- mice also significantly reduced the size of VWF multimers. However, plasma levels of VWF antigen remained unchanged following administration of any dose BDD-FVIII into Adamts13-/- mice, suggesting partial ADAMTS-13 dependency in FVIII-augmented VWF degradation. Moreover, persistent expression of BDD-FVIII at ∼50 to 250 U/dL via AAV8 vector in hemophilia A mice also resulted in a significant reduction of plasma VWF multimer sizes and antigen levels. Finally, the sizes of plasma VWF multimers were significantly reduced in patients with hemophilia A who received a dose of recombinant or plasma-derived FVIII for immune tolerance induction therapy. ConclusionOur results demonstrate the pivotal role of FVIII as a cofactor regulating VWF proteolysis and homeostasis under various (patho)physiological conditions.

P36: Filling the Gap: Developing the Penn State University Child Pump

Study: Despite the need for suitable Mechanical Circulatory Support (MCS) for pediatric patients between 10 and 30 kg, there are currently no long-term implantable devices for this patient cohort. In order to address this need, we are developing the Penn State Child Pump, a continuous flow ventricular assist device (cf-VAD) that will remove the risks of device size mismatch and running at suboptimal operating conditions. Methods: A prototype PSU Child Pump was manufactured based on our experiences with adult cf-LVADs and cf-RVADs. The pump size was tailored to the average chest size of the target pediatric patient population (Figure 1A). In-vitro hemolysis was evaluated under three conditions on our small volume mock circulatory loop with bovine blood (Figure 1B): 1) 8300 RPM, ΔP = 60 mmHg, 3.5 LPM; 2) 8200 RPM, ΔP = 20 mmHg, 5.1 LPM; 3) 8400 RPM, ΔP = 70 mmHg, 3.2 LPM. Results: The initial design of the device has an approximate 38% reduction in size compared to our adult cf-VAD. A mock fit study was conducted using a 3D printed model of a 19 kg patient’s thoracic cavity to compare the size of the PSU Child Pump to the HVAD (Medtronic, Minneapolis, MN) (Figure 1A). In-vitro testing revealed the hemolysis levels at conditions 1, 2, and 3 were 0.037 mg/dL, 0.022 mg/dL, and 0.021 mg/dL, respectively (Figure 1C). Conclusions: The PSU Child Pump prototype was constructed and evaluated, yielding initial results indicating promising hemocompatibility under the given testing conditions. Using a computational fluid dynamics-based optimization platform, the PSU Child Pump will be optimized to meet maximum physiological requirements while minimizing the potential for thrombus formation, plasma-free hemoglobin levels, and degradation of von Willebrand factor. The in-silico optimization results will be validated through a series of 30-day in-vivo studies in sheep.

CARD3: Wall Shear Stress Signature of the Axial B-impeller Design with Superior Hemodynamics

Purpose: Ventricular assist devices (VADs) present great promise as both bridge-to-transplant and destination therapies for patients with end-stage heart failure. However, complications—such as hemolysis, gastrointestinal bleeding, and degradation of the von Willebrand factor—stand in the way of their widespread application. High levels of wall shear stress resulting from poor hemodynamics within the bladed flow domain are primarily responsible. Computational fluid dynamics (CFD) software was used to map the hemodynamic performance of the B-impeller—a biologically-inspired, helicoid VAD impeller design—and provide detailed insight into the internal flow structures, leading to the shear stress levels within. Methods: The B-impeller with casing was parametrically modeled using a computer aided design software. The geometry was exported to a CFD software and simulations were subsequently run to assess the impeller hemodynamics with respect to the wall shear stress parameter. Results: The B-impeller was found to have a very low shear stress signature versus typical VAD designs, which exhibit much higher signatures than the defined hemolytic threshold of 400 Pascals. The peak and area-averaged values of shear stress were found to be 304.512 Pa (at a flow rate of 9 L/min) and 39.949 Pa (averaged over multiple input flow rate simulations), respectively. Conclusion: The B-impeller concept shows great potential to reduce the adverse hemolytic events associated with most other turbo VADs, in addition to its ability to generate high rates of flow in a minimally intrusive diameter of 8 mm. This biologically inspired, rim-driven impeller design was found to reduce shear stress levels over competitive designs by virtue of its radical, open architecture. This low shear stress signature translates to lowering rates of hemolysis, bleeding, and von Willebrand factor degradation, thereby improving post-implantation outcomes of patients.

Constructing and evaluating the effect of micron-scale structures on von Willebrand factor damage

The incidence of clinical complication gastrointestinal bleeding has been proved as consequence of von Willebrand factor (VWF) damage after mechanical circulatory support in clinic. Many studies have been conducted to evaluate VWF damage, of which the most studied influencing factors are mechanical factors such as shear stress. However, in addition to mechanical factors, VWF damage may also be affected by interface factors. To address this issue, a roller pump circulation platform was established to investigate the effect of material surface micron-scale structures distribution on VWF damage in flow state. A composite micro-structure combining microngrating and micronpost was designed and constructed on the surface of Si wafer by lithography and reactive ion etching, and detailed characterization of material surfaces was also performed. Then the changes of VWF antigen, VWF ristocetin cofactor activity, and the degradation of high molecular weight VWF on these surfaces were investigated and compared. The results showed that, with the encryption of surface micro-structures arrangement, the material surface tends to be more hydrophobic, which is beneficial to reduce VWF damage. Therefore, in the design of material surface inside the mechanical circulatory support devices, it can be considered to add some surface micro-structures with a certain distribution density to change the hydrophilicity and hydrophobicity, so as to minimize the VWF damage. These results can provide important references for the evaluation of VWF damage caused by interface factors, and aid in designing material surface inside the mechanical circulatory support devices.

ADAMTS13 inhibition to treat acquired von Willebrand syndrome during mechanical circulatory support device implantation

BackgroundAcquired von Willebrand syndrome (aVWS) is common in patients with mechanical circulatory support (MCS) devices. In these patients, the high shear stress in the device leads to increased shear‐induced proteolysis of von Willebrand factor (VWF) by A Disintegrin And Metalloprotease with Thrombospondin type 1 repeats, number 13 (ADAMTS13). As a result, the high molecular weight (HMW) VWF multimers are lost, leading to a decreased VWF function and impaired hemostasis that could explain the bleeding complications that are frequently observed in these patients. To counteract this abnormal VWF degradation by ADAMTS13, we developed a novel targeted therapy, using an anti‐ADAMTS13 monoclonal antibody (mAb) that inhibits the shear‐induced proteolysis of VWF by ADAMTS13. MethodsHuman or bovine blood was circulated through in vitro MCS device systems with either inhibitory anti‐ADAMTS13 mAb 3H9 or 17C7 (20 μg/ml) or control anti‐ADAMTS13 mAb 5C11 or phosphate buffered saline (PBS). VWF multimers and function (collagen binding activity) were determined at different time points. Next, Impella pumps were implanted in calves and the calves were injected with PBS and subsequently treated with mAb 17C7. VWF, ADAMTS13, and blood parameters were determined. ResultsWe demonstrated that blocking ADAMTS13 could prevent the loss of HMW VWF multimers in in vitro MCS device systems. Importantly, our antibody could reverse aVWS in a preclinical Impella‐induced aVWS calf model. ConclusionHence, inhibition of ADAMTS13 could become a novel therapeutic strategy to manage aVWS in MCS device patients.

Loss of pulsatility with continuous-flow left ventricular assist devices and the significance of the arterial endothelium in von-Willebrand factor production and degradation.

Patients on continuous flow ventricular assist devices (CF-VADs) are at high risk for the development of Acquired von-Willebrand Syndrome (AVWS) and non-surgical bleeding. von Willebrand Factor (vWF) plays an essential role in maintaining hemostasis via platelet binding to the damaged endothelium to facilitate coagulation. In CF-VAD patients, degradation of vWF into low MW multimers that are inefficient in facilitating coagulation occurs and has been primarily attributed to the supraphysiological shear stress associated with the CF-VAD impeller. In this review, we evaluate information from the literature regarding the unraveling behavior of surface-immobilized vWF under pulsatile and continuous flow pertaining to: (A) the process of arterial endothelial vWF production and release into circulation, (B) the critical shear stress required to unravel surface bound versus soluble vWF which leads to degradation, and (C) the role of pulsatility in on the production and degradation of vWF. Taken together, these data suggests that the loss of pulsatility and its impact on arterial endothelial cells plays an important role in the production, release, unraveling, and proteolytic degradation of vWF into low MW multimers, contributing to the development of AVWS. Restoration of pulsatility can potentially mitigate this issue by preventing AVWS and minimizing the risk of non-surgical bleeding.

Kinetic and Dynamic Effects on Degradation of von Willebrand Factor.

The loss of high molecular weight multimers (HMWM) of von Willebrand factor (vWF) in aortic stenosis (AS) and continuous-flow left ventricular assist devices (cf-LVADs) is believed to be associated with high turbulent blood shear. The objective of this study is to understand the degradation mechanism of HMWM in terms of exposure time (kinetic) and flow regime (dynamics) within clinically relevant pathophysiologic conditions. A custom high-shear rotary device capable of creating fully controlled exposure times and flows was used. The system was set so that human platelet-poor plasma flowed through at 1.75 ml/sec, 0.76 ml/sec, or 0.38 ml/sec resulting in the exposure time ( texp ) of 22, 50, or 100 ms, respectively. The flow was characterized by the Reynolds number (Re). The device was run under laminar (Re = 1,500), transitional (Re = 3,000; Re = 3,500), and turbulent (Re = 4,500) conditions at a given texp followed by multimer analysis. No degradation was observed at laminar flow at all given texp . Degradation of HMWM at a given texp increases with the Re. Re ( p < 0.0001) and texp ( p = 0.0034) are significant factors in the degradation of HMWM. Interaction between Re and texp , however, is not always significant ( p = 0.73).

Acquired von Willebrand syndrome (AVWS) type 2, characterized by decreased high molecular weight multimers, is common in children with severe pulmonary hypertension (PH).

Emerging evidence suggests that increased degradation of von Willebrand factor and decrease in high molecular weight multimers occurs in patients with pulmonary hypertension (PH). However, the link between acquired von Willebrand Syndrome (AVWS) type 2 and PH remains poorly understood. We retrospectively evaluated the charts of 20 children with PH who underwent bilateral lung transplantation (LuTx) between 2013 and 2022. Von Willebrand variables were determined in 14 of these patients; 11 patients had complete diagnostics including multimer analysis. We confirmed AVWS in 82% of the children studied (9 of 11 patients by multimer analysis). The two remaining patients had suspected AVWS type 2 because of a VWF:Ac/VWF:Ag ratio of <0.7. Platelet dysfunction or suspicion of VWD type 1 were found in two separate patients. All but one of the 14 children with severe PH had a coagulation disorder. Most patients (9 proven, 2 suspected) had AVWS type 2. Notably, 3 of 5 patients (60%) with normal VWF:Ac/VWF:Ag ratio >0.7 had abnormal VWF multimers, indicating AVWS type 2. Hemostatic complications were observed in 4 of 12 (33%) patients with VWS and 3 of 6 (50%) patients without diagnostics and therapy. For children with moderate to severe PH, we recommend systematic analysis of von Willebrand variables, including multimer analysis, PFA-100 and platelet function testing. Awareness of the diagnosis "AVWS" and adequate therapy may help to prevent these patients from bleeding complications in case of surgical interventions or trauma.

Biomarkers of Clot Activation and Degradation and Risk of Future Major Cardiovascular Events in Acute Exacerbation of COPD: A Cohort Sub-Study in a Randomized Trial Population.

Cardiovascular diseases are common in patients with chronic obstructive pulmonary disease (COPD). Clot formation and resolution secondary to systemic inflammation may be a part of the explanation. The aim was to determine whether biomarkers of clot formation (products of von Willebrand Factor formation and activation) and clot resolution (product of fibrin degeneration) during COPD exacerbation predicted major cardiovascular events (MACE). The cohort was based on clinical data and biobank plasma samples from a trial including patients admitted with an acute exacerbation of COPD (CORTICO-COP). Neo-epitope biomarkers of formation and the activation of von Willebrand factor (VWF-N and V-WFA, respectively) and cross-linked fibrin degradation (X-FIB) were assessed using ELISAs in EDTA plasma at the time of acute admission, and analyzed for time-to-first MACE within 36 months, using multivariable Cox proportional hazards models. In total, 299/318 participants had samples available for analysis. The risk of MACE for patients in the upper quartile of each biomarker versus the lower quartile was: X-FIB: HR 0.98 (95% CI 0.65–1.48), VWF-N: HR 1.56 (95% CI 1.07–2.27), and VWF-A: HR 0.78 (95% CI 0.52–1.16). Thus, in COPD patients with an acute exacerbation, VWF-N was associated with future MACE and warrants further studies in a larger population.

Increased cleavage of von Willebrand factor by ADAMTS13 may contribute strongly to acquired von Willebrand syndrome development in patients with essential thrombocythemia

BackgroundPatients with essential thrombocythemia (ET) often experience bleeding associated with acquired von Willebrand syndrome (AVWS) when the platelet count is markedly increased. ObjectiveWe investigated whether von Willebrand factor (VWF) degradation is enhanced in patients with ET. MethodsSeventy patients with ET underwent VWF multimer (VWFM) analysis and measurement of VWF‐related parameters. We calculated the VWFM index, defined as the ratio of intensities of a patient's molecular weight‐categorized VWFMs, and those of a healthy subject's, using densitometric analysis. VWF degradation product (DP) was measured via ELISA using a monoclonal antibody that specifically recognizes Y1605 at the C‐terminal boundary, which is exposed following ADAMTS13‐mediated cleavage of the Y1605‐M1606 bond of the VWF A2 domain. ResultsPatients with higher platelet counts had a significantly reduced high molecular weight (HMW)‐VWFM index and an increased VWF‐DP:VWF antigen (Ag) ratio compared to those with lower platelet counts. On multivariate analysis, the VWF‐DP/VWF:Ag ratio was an independent predictor of the HMW‐VWFM index. Patients who underwent cytoreductive therapy had a significantly higher HMW‐VWFM index and lower VWF‐DP/VWF:Ag ratio than those who did not. Among individual patients, there was also a significant increase in the HMW‐VWFM index and a decrease in the VWF‐DP/VWF:Ag ratio after cytoreductive therapy compared to pre‐therapy values. ConclusionIn patients with ET, an increased platelet count is associated with enhanced cleavage of VWF at the Y1605‐M1606 bond, primarily by ADAMTS13, leading to AVWS. Cytoreductive therapy reduces the platelet count, prevents excessive VWF cleavage, and improves VWFM distributions.

In vitro study of red blood cell and VWF damage in mechanical circulatory support devices based on blood-shearing platform.

Blood damage induced by mechanical circulatory support devices (MCSDs) remains a significant challenge to optimal clinical care. Although researchers have been conducting in vitro studies, the major determinant of blood damage is still unclear. An optimized capillary tube blood-shearing platform with custom designed parts was constructed to investigate the influence of two flow-dependent parameters (shear stress and exposure time) on the shear-induced damage of red blood cells and von Willebrand factor (VWF). Blood samples under different high shear stress and instantaneous exposure time were obtained by changing the flow rate and the length of capillary tube. Plasma free hemoglobin assay and immunoblotting of VWF were then performed on the sheared blood samples. The quantitative correlation between the hemolysis index and the two flow-dependent parameters was found following the power law mathematical model under the flow condition with high shear stress and instantaneous exposure time. The degradation of high molecular weight VWF was not obvious under high shear stress factor. However, the degradation of high molecular weight VWF was found as the result of the accumulation over exposure time under non-physiological shear stress, which was consistent with the different mechanism of VWF damage comparing to red blood cell damage. Compared to peak shear stress, exposure time has a greater effect on both red blood cell and VWF damage. To improve the hemocompatibility of MCSDs, it is more important to avoid regions of slow blood flow with non-physiological shear stress under laminar flow conditions.

Impact of Statins on the Incidence of Gastrointestinal Bleeding Events Among Patients with Continuous Flow Left Ventricular Assist Devices

<h3>Purpose</h3> Patients with continuous flow left ventricular assist devices (CF-LVADs) are at an increased risk of gastrointestinal bleeding (GIB) events due to von Willebrand factor degradation, enhanced angiogenesis and endothelial dysfunction resulting from non-pulsatile flows of the device. LVAD patients also commonly receive statins for the primary or secondary prevention of cardiovascular disease. However, the impact of such therapy on the incidence of GIB is controversial. Importantly, literature regarding the impact of statins on GIB in LVAD patients is lacking. <h3>Methods</h3> This was a single-center, retrospective review of adult patients that underwent CF-LVAD implantation between May 2016 and January 2020. Patients were categorized based on statin use throughout the study period. The primary outcome was the composite of arteriovenous malformation (AVM) confirmed GIB and other major GIB events for up to 1-year post-LVAD implantation. Secondary outcomes included each of the components of the primary outcome, non-clinically relevant GIB, time to GIB, and frequency of GIB. Multivariable Cox regression was utilized to assess association between confounding variables and major GIB. <h3>Results</h3> Of the 123 patients who met inclusion criteria, 66 (54%) received statin therapy during the study period. No difference was observed in the primary outcome of major GIB between the statin and non-statin groups (21.2% vs. 12.3%, p=0.20). Similarly, AVM confirmed GIB (12.1% vs. 5.3%, p=0.20), other major GIB (9.1% vs. 7.0%, p=0.75), and non-clinically relevant GIB (3.0% vs. 1.8%, p=0.68) were similar between the two groups. Using a competing risk modeling, we observed no difference in the development of a major GIB (32.4% vs. 18.7%, p=0.14) or in the frequency of major GIB [1.5 (interquartile range, 1-2) vs. 1 (1-1), p=0.12] within 12 months of device implantation. Multivariable Cox regression revealed that older age and higher baseline creatinine were associated with an increased risk of GIB within 1-year of LVAD implantation (p<0.05 for both). <h3>Conclusion</h3> Among patients with CF-LVADs, there was a non-significant increase in the incidence of major GIB events with the use of statin therapy. Further long-term studies are needed to determine if statin use has a significant impact on the incidence of GIB events among this patient population.

Patient-specific severity of von Willebrand factor degradation identifies patients with a left ventricular assist device at high risk for bleeding

BackgroundContinuous-flow left ventricular assist devices (LVADs) cause an acquired von Willebrand factor (VWF) deficiency and bleeding. Models to risk-stratify for bleeding are urgently needed. We developed a model of continuous-flow LVAD bleeding risk from patient-specific severity of VWF degradation. MethodsIn a prospective, longitudinal cohort study, paired blood samples were obtained from patients (n = 67) with a continuous-flow LVAD before and during support. After 640 ± 395 days, patients were categorized as all-cause bleeders, gastrointestinal (GI) bleeders, or nonbleeders. VWF multimers and VWF clotting function were evaluated to determine bleeding risk. ResultsOf 67 patients, 34 (51%) experienced bleeding, 26 (39%) experienced GI bleeding, and 33 (49%) did not bleed. In all patients, LVAD support significantly reduced high-molecular-weight VWF multimers (P < .001). Bleeders exhibited greater loss of high-molecular-weight VWF multimers (mean ± standard deviation, –10 ± 5% vs –7 ± 4%, P = .008) and reduced VWF clotting function versus nonbleeders (median [interquartile range], –12% [–31% to 4%] vs 0% [–9 to 26%], P = .01). A combined metric of VWF multimers and VWF function generated the All-Cause Bleeding Risk Score, which stratified bleeders versus nonbleeders (86 ± 56% vs 41 ± 48%, P < .001) with a positive predictive value of 86% (95% confidence interval, 66%-95%) and diagnostic odds ratio of 11 (95% confidence interval, 2.9-44). A separate GI Bleeding Risk Score stratified GI bleeders versus nonbleeders (202 ± 114 vs 120 ± 86, P = .003) with a positive predictive value of 88% (64%-97%) and diagnostic odds ratio of 18 (3.1-140). ConclusionsThe severity of loss of VWF multimers and VWF clotting function generated Bleeding Risk Scores with high predictive value for LVAD-associated bleeding. This model may guide personalized antithrombotic therapy and patient surveillance.

Effect of pulsatility on shear-induced extensional behavior of Von Willebrand factor.

Patients with continuous flow ventricular assist devices (CF-VADs) are at high risk for non-surgical bleeding, speculated to associate with the loss of pulsatility following CF-VAD placement. It has been hypothesized that continuous shear stress causes elongation and increased enzymatic degradation of von Willebrand Factor (vWF), a key player in thrombus formation at sites of vascular damage. However, the role of loss of pulsatility on the unravelling behavior of vWF has not been widely explored. vWF molecules were immobilized on the surface of microfluidic devices and subjected to various pulsatile flow profiles, including continuous flow and pulsatile flow of different magnitudes, dQ/dt (i.e., first derivative of flow rate) of pulsatility and pulse frequencies to mimic in vivo shear flow environments with and without CF-VAD support. VWF elongation was observed using total internal reflection fluorescence (TIRF) microscopy. Besides, the vWF level is measured from the patients' blood sample before and after CF-VAD implantation from a clinical perspective. To our knowledge, this work is the first in providing direct, visual observation of single vWF molecule extension under controlled-pulsatile shear flow. Unravelling of vWF (total sample size n~200 molecules) is significantly reduced under pulsatile flow (p<0.01) compared to continuous flow. An increase in the magnitude of pulsatility further reduces unravelling lengths, while lower frequency of pulsatility (20 vs. 60 pulses per min) does not have a major effect on the maximum or minimum unravelling lengths. Evaluation of CF-VAD patient blood samples (n=13) demonstrates that vWF levels decreased by ~40% following CF-VAD placement (p<0.01), which correlates to single-molecule observations from a clinical point of view. Pulsatile flow reduces unfolding of vWF compared to continuous flow and a lower pulse frequency of 20 pulses/minute yielded comparable vWF unfolding to 60 pulses/minute. These findings could shed light on non-surgical bleeding associated with the loss of pulsatility following CF-VAD placement.

Discrete responses of erythrocytes, platelets, and von Willebrand factor to shear

Despite decades of technological advancements in blood-contacting medical devices, complications related to shear flow-induced blood trauma are still frequently observed in clinic. Blood trauma includes haemolysis, platelet activation, and degradation of High Molecular Weight von Willebrand Factor (HMW vWF) multimers, all of which are dependent on the exposure time and magnitude of shear stress. Specifically, accumulating evidence supports that when blood is exposed to shear stresses above a certain threshold, blood trauma ensues; however, it remains unclear how various constituents of blood are affected by discrete shears experimentally. The aim of this study was to expose blood to discrete shear stresses and evaluate blood trauma indices that reflect red cell, platelet, and vWF structure. Citrated human whole blood (n = 6) was collected and its haematocrit was adjusted to 30 ± 2% by adding either phosphate buffered saline (PBS) or polyvinylpyrrolidone (PVP). Viscosity of whole blood was adjusted to 3.0, 12.5, 22.5 and 37.5 mPa·s to yield stresses of 3, 6, 9, 12, 50, 90 and 150 Pa in a custom-developed shearing system. Blood samples were exposed to shear for 0, 300, 600 and 900 s. Haemolysis was measured using spectrophotometry, platelet activation using flow cytometry, and HMW vWF multimer degradation was quantified with gel electrophoresis and immunoblotting. For tolerance to 300, 600 and 900 s of exposure time, the critical threshold of haemolysis was reached after blood was exposed to 90 Pa for 600 s (P < 0.05), platelet activation and HMW vWF multimer degradation were 50 Pa for 600 s and 12 Pa for 300 s respectively (P < 0.05). Our experimental results provide simultaneous comparison of blood trauma indices and thus also the relation between shear duration and magnitude required to induce damage to red cells, platelets, and vWF. Our results also demonstrate that near-physiological shear stress (<12 Pa) is needed in order to completely avoid any form of blood trauma. Therefore, there is an urgent need to design low shear-flow medical devices in order to avoid blood trauma in this blood-contacting medical device field.