Off-target effect of high-dose sildenafil on adenosine 5'- diphosphate and collagen-induced platelet activation through mitogen-activated protein kinase pathway in treated BALB/C mice and in vitro experiments: A preliminary study.

ABSTRACT: Sildenafil, a common over-the-counter pill often self-administered at high doses for erectile dysfunction, has been reported to rarely cause prothrombotic events and sudden cardiac death in a few case reports. Therefore, we investigated the in vitro and in vivo effect of sildenafil treatment and dosage on platelet activation and mitogen-activated protein kinase (MAPK) phosphorylation. BALB/C mice were segregated into four groups, each having four mice each (control, low [3.25 mg/kg], medium [6.5 mg/kg], and high [13 mg/kg] sildenafil), and after the treatment, blood was drawn from each mouse and washed platelets prepared. Washed platelets were incubated with CD41 PE-Cy7 and Phospho-p38 MAPK PE antibodies and analyzed using a flow cytometer for platelet activation and adenosine 5'- diphosphate (ADP)/collagen-induced MAPK phosphorylation. Washed platelets obtained from the venous blood of 18 human volunteers, were incubated with PAC-1 FITC and Phospho-p38 MAPK PE antibodies, and platelet activation (ADP and collagen), followed by flow cytometry analysis. There was a significant increase in both platelet activation as well as MAPK phosphorylation in the presence of collagen in the high-dose (13 mg/kg) sildenafil group (P = 0.000774). Further, increased platelet activation was observed in samples that were treated with high-dose sildenafil as compared to the untreated samples (P < 0.00001). These studies show the risk of prothrombotic episodes in patients on high-dose sildenafil (100 mg), in those with even mild endothelial dysfunction due to ADP, and collagen-induced platelet activation through MAPK phosphorylation, which was not seen in the low-and intermediate-dose cohorts.

Spa2 remodels ADP-actin via molecular condensation under glucose starvation.

Actin nucleotide-dependent actin remodeling is essential to orchestrate signal transduction and cell adaptation. Rapid energy starvation requires accurate and timely reorganization of the actin network. Despite distinct treadmilling mechanisms of ADP- and ATP-actin filaments, their filament structures are nearly identical. How other actin-binding proteins regulate ADP-actin filament assembly is unclear. Here, we show that Spa2 which is the polarisome scaffold protein specifically remodels ADP-actin upon energy starvation in budding yeast. Spa2 triggers ADP-actin monomer nucleation rapidly through a dimeric core of Spa2 (aa 281-535). Concurrently, the intrinsically disordered region (IDR, aa 1-281) guides Spa2 undergoing phase separation and wetting on the surface of ADP-G-actin-derived F-actin and bundles the filaments. Both ADP-actin-specific nucleation and bundling activities of Spa2 are actin D-loop dependent. The IDR and nucleation core of Spa2 are evolutionarily conserved by coexistence in the fungus kingdom, suggesting a universal adaptation mechanism in the fungal kingdom in response to glucose starvation, regulating ADP-G-actin and ADP-F-actin with high nucleotide homogeneity.

The prognostic value of platelet aggregation in patients with sepsis.

BACKGROUND: This study aims to investigate the relevance of platelet aggregation markers, specifically arachidonic acid (AA) and adenosine diphosphate (ADP), in relation to the prognosis of sepsis patients. METHODS: A cohort of 40 sepsis patients was included and stratified, based on their 28-day post-treatment prognosis, into two groups: a survival group (n = 31) and a severe sepsis group (n = 9. Then, their various clinical parameters, including patient demographics, platelet counts (PLT), inflammatory markers, and platelet aggregation rates (PAR) induced by AA and ADP between the two groups, were compared. Long-term health implications of sepsis were assessed using the Acute Physiologic Assessment and Chronic Health Evaluation II (APACHE II) score, and logistic regression analysis was conducted to evaluate the prognostic significance of PAR in sepsis patients. RESULTS: Patients with severe sepsis exhibited significantly elevated levels of procalcitonin (PCT), platelet adhesion rates, and PAR induced by ADP (P < 0.05), but having lower PLT (P < 0.05), compared to those in the survival group. Logistic regression analysis demonstrated that PAR induced by ADP was a protective factor in predicting prognosis in sepsis patients (P < 0.01). CONCLUSIONS: Activation of platelets in sepsis intensifies inflammatory response. Patients with sepsis whose ADP-induced PAR was < 60% displayed significant impairment in platelet aggregation function, and had higher mortality rate. Monitoring ADP-induced PAR is crucial for management of sepsis.

Assessment of platelet functional activity in healthy individuals and patients receiving antiplatelet therapy. Possible inconsistencies between aggregation and flow cytometry tests.

Platelet functional activity was assessed in healthy volunteers (HV, n=92), patients with stable angina pectoris (SA, n=42) and acute coronary syndrome (ACS, n=73), treated with acetylsalicylic acid (ASA) + clopidogrel and ASA + ticagrelor, respectively. In all HV and patients we have compared parameters of platelet aggregation (maximum light transmission and velocity, Tmax and Vmax) and parameters, characterizing exposure of platelet activation markers, evaluated by flow cytometry. HV platelets were activated by 10 µM, 1 µM TRAP, and 20 µM, 5 µM, 2.5 µM ADP; patient platelets were activated by 10 µM TRAP and by 20 µM and 5 µM ADP. Strong and significant correlations between the aggregation and flow cytometry parameters (the r correlation coefficient from 0.4 up to >0.6) most frequently were registered in HV platelet during activation by 1 µM TRAP and in SA patients during platelet activation by 20 µM and 5 µM ADP. However, in many other cases these correlations were rather weak (r < 0.3) and sometimes statistically insignificant. In HV the differences in PAC-1 binding parameters between platelets activated by 10 µM TRAP (the strongest agonist) and all ADP concentrations were negligible (≤ 10%), while CD62P binding (at all ADP concentrations) and LTA parameters for (5 µM and 2.5 µM ADP) were significantly lower (by 40-60%). Antiplatelet therapy in patients decreased all parameters as compared to HV, but to varying extents. For 10 µM TRAP the MFI index for PAC-1 binding (40-50% decrease) and for both ADP concentrations the Tmax values (60-85% decrease) appeared to be the most sensitive in comparison with the other parameters that decreased to a lesser extent. The data obtained indicate a possibility of inconsistency between different LTA and flow cytometry parameters in assessing platelet activity and efficacy of antiplatelet drugs.

Spontaneous Overactivation of Xenopus Frog Eggs Triggers Necrotic Cell Death.

The excessive activation of frog eggs, referred to as overactivation, can be initiated by strong oxidative stress, leading to expedited calcium-dependent non-apoptotic cell death. Overactivation also occurs spontaneously, albeit at a low frequency, in natural populations of spawned frog eggs. Currently, the cytological and biochemical events of the spontaneous process have not been characterized. In the present study, we demonstrate that the spontaneous overactivation of Xenopus frog eggs, similarly to oxidative stress- and mechanical stress-induced overactivation, is characterized by the fast and irreversible contraction of the egg's cortical layer, an increase in egg size, the depletion of intracellular ATP, a drastic increase in the intracellular ADP/ATP ratio, and the degradation of M phase-specific cyclin B2. These events manifest in eggs in the absence of caspase activation within one hour of triggering overactivation. Importantly, substantial amounts of ATP and ADP leak from the overactivated eggs, indicating that plasma membrane integrity is compromised in these cells. The rupture of the plasma membrane and acute depletion of intracellular ATP explicitly define necrotic cell death. Finally, we report that egg overactivation can occur in the frog's genital tract. Our data suggest that mechanical stress may be a key factor promoting egg overactivation during oviposition in frogs.

Glucose Regulation of ß-Cell KATP Channels: Is a New Model Needed?

The canonical model of glucose-induced increase in insulin secretion involves the metabolism of glucose via glycolysis and the citrate cycle, resulting in increased ATP synthesis by the respiratory chain and the closure of ATP-sensitive K+ (KATP) channels. The resulting plasma membrane depolarization, followed by Ca2+ influx through L-type Ca2+ channels, then induces insulin granule fusion. Merrins and colleagues have recently proposed an alternative model whereby KATP channels are controlled by pyruvate kinase, using glycolytic and mitochondrial phosphoenolpyruvate (PEP) to generate microdomains of high ATP/ADP immediately adjacent to KATP channels. This model presents several challenges. First, how mitochondrially generated PEP, but not ATP produced abundantly by the mitochondrial F1F0-ATP synthase, can gain access to the proposed microdomains is unclear. Second, ATP/ADP fluctuations imaged immediately beneath the plasma membrane closely resemble those in the bulk cytosol. Third, ADP privation of the respiratory chain at high glucose, suggested to drive alternating, phased-locked generation by mitochondria of ATP or PEP, has yet to be directly demonstrated. Finally, the approaches used to explore these questions may be complicated by off-target effects. We suggest instead that Ca2+ changes, well known to affect both ATP generation and consumption, likely drive cytosolic ATP/ADP oscillations that in turn regulate KATP channels and membrane potential. Thus, it remains to be demonstrated that a new model is required to replace the existing, mitochondrial bioenergetics-based model.

Competition between physical search and a weak-to-strong transition rate-limits kinesin binding times.

The self-organization of cells relies on the profound complexity of protein-protein interactions. Challenges in directly observing these events have hindered progress toward understanding their diverse behaviors. One notable example is the interaction between molecular motors and cytoskeletal systems that combine to perform a variety of cellular functions. In this work, we leverage theory and experiments to identify and quantify the rate-limiting mechanism of the initial association between a cargo-bound kinesin motor and a microtubule track. Recent advances in optical tweezers provide binding times for several lengths of kinesin motors trapped at varying distances from a microtubule, empowering the investigation of competing models. We first explore a diffusion-limited model of binding. Through Brownian dynamics simulations and simulation-based inference, we find this simple diffusion model fails to explain the experimental binding times, but an extended model that accounts for the ADP state of the molecular motor agrees closely with the data, even under the scrutiny of penalizing for additional model complexity. We provide quantification of both kinetic rates and biophysical parameters underlying the proposed binding process. Our model suggests that a typical binding event is limited by ADP state rather than physical search. Lastly, we predict how these association rates can be modulated in distinct ways through variation of environmental concentrations and physical properties.

Conformations of Bcs1L undergoing ATP hydrolysis suggest a concerted translocation mechanism for folded iron-sulfur protein substrate.

The human AAA-ATPase Bcs1L translocates the fully assembled Rieske iron-sulfur protein (ISP) precursor across the mitochondrial inner membrane, enabling respiratory Complex III assembly. Exactly how the folded substrate is bound to and released from Bcs1L has been unclear, and there has been ongoing debate as to whether subunits of Bcs1L act in sequence or in unison hydrolyzing ATP when moving the protein cargo. Here, we captured Bcs1L conformations by cryo-EM during active ATP hydrolysis in the presence or absence of ISP substrate. In contrast to the threading mechanism widely employed by AAA proteins in substrate translocation, subunits of Bcs1L alternate uniformly between ATP and ADP conformations without detectable intermediates that have different, co-existing nucleotide states, indicating that the subunits act in concert. We further show that the ISP can be trapped by Bcs1 when its subunits are all in the ADP-bound state, which we propose to be released in the apo form.

Structural basis and synergism of ATP and Na+ activation in bacterial K+ uptake system KtrAB.

The K+ uptake system KtrAB is essential for bacterial survival in low K+ environments. The activity of KtrAB is regulated by nucleotides and Na+. Previous studies proposed a putative gating mechanism of KtrB regulated by KtrA upon binding to ATP or ADP. However, how Na+ activates KtrAB and the Na+ binding site remain unknown. Here we present the cryo-EM structures of ATP- and ADP-bound KtrAB from Bacillus subtilis (BsKtrAB) both solved at 2.8 Å. A cryo-EM density at the intra-dimer interface of ATP-KtrA was identified as Na+, as supported by X-ray crystallography and ICP-MS. Thermostability assays and functional studies demonstrated that Na+ binding stabilizes the ATP-bound BsKtrAB complex and enhances its K+ flux activity. Comparing ATP- and ADP-BsKtrAB structures suggests that BsKtrB Arg417 and Phe91 serve as a channel gate. The synergism of ATP and Na+ in activating BsKtrAB is likely applicable to Na+-activated K+ channels in central nervous system.

Thermodynamic analysis of energy coupling by determination of the Onsager phenomenological coefficients for a 3×3 system of coupled chemical reactions and transport in ATP synthesis and its mechanistic implications.

The nonequilibrium coupled processes of oxidation and ATP synthesis in the fundamental process of oxidative phosphorylation (OXPHOS) are of vital importance in biosystems. These coupled chemical reaction and transport bioenergetic processes using the OXPHOS pathway meet >90% of the ATP demand in aerobic systems. On the basis of experimentally determined thermodynamic OXPHOS flux-force relationships and biochemical data for the ternary system of oxidation, ion transport, and ATP synthesis, the Onsager phenomenological coefficients have been computed, including an estimate of error. A new biothermokinetic theory of energy coupling has been formulated and on its basis the thermodynamic parameters, such as the overall degree of coupling, q and the phenomenological stoichiometry, Z of the coupled system have been evaluated. The amount of ATP produced per oxygen consumed, i.e. the actual, operating P/O ratio in the biosystem, the thermodynamic efficiency of the coupled reactions, η, and the Gibbs free energy dissipation, Φ have been calculated and shown to be in agreement with experimental data. At the concentration gradients of ADP and ATP prevailing under state 3 physiological conditions of OXPHOS that yield Vmax rates of ATP synthesis, a maximum in Φ of ∼0.5J(hmgprotein)-1, corresponding to a thermodynamic efficiency of ∼60% for oxidation on succinate, has been obtained. Novel mechanistic insights arising from the above have been discussed. This is the first report of a 3 × 3 system of coupled chemical reactions with transport in a biological context in which the phenomenological coefficients have been evaluated from experimental data.

The tilting motion of the central core reveals the transport mechanism of the sarco/endoplasmic reticulum Ca2+-ATPase.

The sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) transports two Ca2+ ions per ATP hydrolyzed from the cytoplasm to the lumen. However, how the ATP hydrolysis remotely drives the Ca2+ transport is unclear. In the SERCA1a crystal structures, the ATP hydrolysis is accompanied by the notably increasing tilting angle of the central core (CC) and the Ca2+ transport, and the CC tilting angle dramatically decreases in the E2 to E1 transition. We demonstrated that the significantly increasing tilting motion of the CC drove the Ca2+ release in the molecular dynamics simulation of the R836A variant, and the dramatic spontaneous decrease in the CC tilting angle of the E2 state triggers the restart of the SERCA1a's transport cycle. The repulsion between the phosphorylated D351 and the phosphate groups in ADP triggers the release of ADP from the SERCA1a headpiece. We proposed a novel SERCA transport mechanism in which ATP hydrolysis drives a significant tilting motion of the CC, which drives Ca2+ transport and the A domain rotational motion in the E1P-ADP-2Ca2+ to E2P transition. The dramatic spontaneous decrease in the CC tilting angle of the E2 state drives the restart of the transport cycle.

Generation and characterization of antagonistic anti-human CD39 nanobodies.

CD39 is the major enzyme controlling the levels of extracellular adenosine triphosphate (ATP) via the stepwise hydrolysis of ATP to adenosine diphosphate (ADP) and adenosine monophosphate (AMP). As extracellular ATP is a strong promoter of inflammation, monoclonal antibodies (mAbs) blocking CD39 are utilized therapeutically in the field of immune-oncology. Though anti-CD39 mAbs are highly specific for their target, they lack deep penetration into the dense tissue of solid tumors, due to their large size. To overcome this limitation, we generated and characterized nanobodies that targeted and blocked human CD39. From cDNA-immunized alpacas we selected 16 clones from seven nanobody families that bind to two distinct epitopes of human CD39. Among these, clone SB24 inhibited the enzymatic activity of CD39. Of note, SB24 blocked ATP degradation by both soluble and cell surface CD39 as a 15kD monomeric nanobody. Dimerization via fusion to an immunoglobulin Fc portion further increased the blocking potency of SB24 on CD39-transfected HEK cells. Finally, we confirmed the CD39 blocking properties of SB24 on human PBMCs. In summary, SB24 provides a new small biological antagonist of human CD39 with potential application in cancer therapy.

Binding patterns of inhibitors to different pockets of kinesin Eg5.

The kinesin-5 family member, Eg5, plays very important role in the mitosis. As a mitotic protein, Eg5 is the target of various mitotic inhibitors. There are two targeting pockets in the motor domain of Eg5, which locates in the α2/L5/α3 region and the α4/α6 region respectively. We investigated the interactions between the different inhibitors and the two binding pockets of Eg5 by using all-atom molecular dynamics method. Combined the conformational analysis with the free-energy calculation, the binding patterns of inhibitors to the two binding pockets are shown. The α2/L5/α3 pocket can be divided into 4 regions. The structures and binding conformations of inhibitors in region 1 and 2 are highly conserved. The shape of α4/α6 pocket is alterable. The space of this pocket in ADP-binding state of Eg5 is larger than that in ADP·Pi-binding state due to the limitation of a hydrogen bond formed in the ADP·Pi-binding state. The results of this investigation provide the structural basis of the inhibitor-Eg5 interaction and offer a reference for the Eg5-targeted drug design.

AMP-activated protein kinase can be allosterically activated by ADP but AMP remains the key activating ligand.

The AMP-activated protein kinase (AMPK) is a sensor of cellular energy status. When activated by increases in ADP:ATP and/or AMP:ATP ratios (signalling energy deficit), AMPK acts to restore energy balance. Binding of AMP to one or more of three CBS repeats (CBS1, CBS3, CBS4) on the AMPK-γ subunit activates the kinase complex by three complementary mechanisms: (i) promoting α-subunit Thr172 phosphorylation by the upstream kinase LKB1; (ii) protecting against Thr172 dephosphorylation; (iii) allosteric activation. Surprisingly, binding of ADP has been reported to mimic the first two effects, but not the third. We now show that at physiologically relevant concentrations of Mg.ATP2- (above those used in the standard assay) ADP binding does cause allosteric activation. However, ADP causes only a modest activation because (unlike AMP), at concentrations just above those where activation becomes evident, ADP starts to cause competitive inhibition at the catalytic site. Our results cast doubt on the physiological relevance of the effects of ADP and suggest that AMP is the primary activator in vivo. We have also made mutations to hydrophobic residues involved in binding adenine nucleotides at each of the three γ subunit CBS repeats of the human α2ß2γ1 complex and examined their effects on regulation by AMP and ADP. Mutation of the CBS3 site has the largest effects on all three mechanisms of AMP activation, especially at lower ATP concentrations, while mutation of CBS4 reduces the sensitivity to AMP. All three sites appear to be required for allosteric activation by ADP.

Biomimetic nanodrug blocks CD73 to inhibit adenosine and boosts antitumor immune response synergically with photothermal stimulation.

Combination of tumor immunotherapy with photothermal therapy (PTT) is a feasible tactic to overcome the drawback of immunotherapy such as poor immune response. Via triggering the immunogenic cells death (ICD), PTT can stimulate the activity of immune cells, but meanwhile, the level of adenosine is elevated via the CD73-induced decomposition of ATP which is overexpressed accompanying with the PTT process, resulting in negative feedback to impair the immune stimulation. Herein, we developed a novel biomimetic photothermal nanodrug to specifically block CD73 for inhibition of adenosine production and more efficient priming of the suppressive immune microenvironments. The nanodrug, named as AptEM@CBA, is constructed by encapsulation of photothermal agent black phosphorus quantum dots (BPQDs) and selective CD73 inhibitor α, ß-Methyleneadenosine 5'-diphosphate (AMPCP) in chitosan nanogels, which are further covered with aptamer AS1411 modified erythrocyte membrane (EM) for biomimetic camouflage. With AS1411 induced active targeting and EM induced long blood circulation time, the enrichment of the nanodrug tumor sites is promoted. The photothermal treatment promotes the maturation of dendritic cells. Meanwhile, the release of AMPCP suppress the adenosine generation via CD73 blockade, alleviating the impairment of adenosine to dendritic cells and suppressing regulatory T cells, synergically stimulate the activity of T cells. The combination of CD73 blockade with PTT, not only suppresses the growth of primary implanted tumors, but also boosts strong antitumor immunity to inhibit the growth of distal tumors, providing good potential for tumor photoimmunotherapy.

Mitotic spindle positioning protein (MISP) preferentially binds to aged F-actin.

Actin bundling proteins crosslink filaments into polarized structures that shape and support membrane protrusions including filopodia, microvilli, and stereocilia. In the case of epithelial microvilli, mitotic spindle positioning protein (MISP) is an actin bundler that localizes specifically to the basal rootlets, where the pointed ends of core bundle filaments converge. Previous studies established that MISP is prevented from binding more distal segments of the core bundle by competition with other actin-binding proteins. Yet whether MISP holds a preference for binding directly to rootlet actin remains an open question. By immunostaining native intestinal tissue sections, we found that microvillar rootlets are decorated with the severing protein, cofilin, suggesting high levels of ADP-actin in these structures. Using total internal reflection fluorescence microscopy assays, we also found that purified MISP exhibits a binding preference for ADP- versus ADP-Pi-actin-containing filaments. Consistent with this, assays with actively growing actin filaments revealed that MISP binds at or near their pointed ends. Moreover, although substrate attached MISP assembles filament bundles in parallel and antiparallel configurations, in solution MISP assembles parallel bundles consisting of multiple filaments exhibiting uniform polarity. These discoveries highlight nucleotide state sensing as a mechanism for sorting actin bundlers along filaments and driving their accumulation near filament ends. Such localized binding might drive parallel bundle formation and/or locally modulate bundle mechanical properties in microvilli and related protrusions.

Transcriptome analysis of cultured human vascular endothelial cells after γ-ray irradiation and correlation analysis with ATP, ADP, and adenosine as secondary soluble factors.

Vascular endothelial cells serve as barriers between blood components and subendothelial tissue and regulate the blood coagulation-fibrinolytic system. Ionizing radiation is a common physical stimulant that induces a bystander effect whereby irradiated cells influence neighboring cells through signalings, including purinergic receptor signaling, activated by adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), and adenosine as secondary soluble factors. Human vascular endothelial EA.hy926 cells were cultured and irradiated with γ-rays or treated with ATP, ADP, or adenosine under non-toxic conditions. RNA-seq, gene ontology, and hierarchical clustering analyses were performed. The transcriptome analysis of differentially expressed genes in vascular endothelial cells after γ-ray irradiations suggests that the change of gene expression by γ-irradiation is mediated by ATP and ADP. In addition, the expression and activity of the proteins related to blood coagulation and fibrinolysis systems appear to be secondarily regulated by ATP and ADP in vascular endothelial cells after exposure to γ-irradiation. Although it is unclear whether the changes of the gene expression related to blood coagulation and fibrinolysis systems by γ-irradiation affected the increased hemorrhagic tendency through the exposure to γ-irradiation or the negative feedback to the activated blood coagulation system, the present data indicate that toxicity associated with γ-irradiation involves the dysfunction of vascular endothelial cells related to the blood coagulation-fibrinolytic system, which is mediated by the signalings, including purinergic receptor signaling, activated by ATP and ADP.

Metabolomics and network pharmacology reveal the mechanism of antithrombotic effect of Asperosaponin VI.

Dipsaci Radix may possess antithrombotic properties, and one of its primary active ingredients is Asperosaponin VI. However, the antithrombotic effects and pharmacological mechanisms of Asperosaponin VI remain unclear. An in vivo experimental study has demonstrated the antithrombotic activity of Asperosaponin VI. Asperosaponin VI also exhibits anticoagulant properties. Asperosaponin VI significantly hindered collagen adrenergic-induced acute pulmonary thrombosis in mice and enhanced their survival rate. This hinders the formation of acute pulmonary embolisms induced by adenosine diphosphate (ADP) and decreases recovery time. A comprehensive strategy that combines metabolomics, network pharmacology, molecular docking, and experimental validation has the potential to reveal the antithrombotic mechanisms of Asperosaponin VI. Metabolomic evidence suggests that Asperosaponin VI may influence platelet aggregation and the production of anti-inflammatory metabolites through the regulation of pathways such as phenylalanine and arachidonic acid metabolism, thereby inhibiting thrombosis. Network pharmacology identified the pharmacological targets of Asperosaponin VI and indicated that it treats thrombi by partially regulating the signaling pathways related to inflammation and platelet aggregation. Asperosaponin VI showed strong binding affinity for F2, PTPRC, JUN, STAT3, SRC, AKT1. The antiplatelet aggregation activity of Asperosaponin VI was validated based on the metabolomic and network pharmacology results. Asperosaponin VI inhibits platelet aggregation induced by ADP, AA, and collagen. Therefore, Asperosaponin VI exerts antithrombotic effects through antiplatelet aggregation. Therefore, Asperosaponin VI is a promising antithrombotic agent.

A multi-constituent model for assessing the effect of impeller shroud on the thrombosis potential of a centrifugal blood pump.

Centrifugal blood pumps can be used for treating heart failure patients. However, pump thrombosis has remained one of the complications that trouble clinical treatment. This study analyzed the effect of impeller shroud on the thrombosis risk of the blood pump, and predicted areas prone to thrombosis. Multi-constituent transport equations were presented, considering mechanical activation and biochemical activation. It was found that activated platelets concentration can increase with shear stress and adenosine diphosphate(ADP) concentration increasing, and the highest risk of thrombosis inside the blood pump was under extracorporeal membrane oxygenation (ECMO) mode. Under the same condition, ADP concentration and thrombosis index of semi-shroud impeller can increase by 7.3% and 7.2% compared to the closed-shroud impeller. The main reason for the increase in thrombosis risk was owing to elevated scalar shear stress and more coagulation promoting factor-ADP released. The regions with higher thrombosis potential were in the center hole, top and bottom clearance. As a novelty, the findings revealed that impeller shroud can influence mechanical and biochemical activation factors. It is useful for identifying potential risk regions of thrombus formation based on relative comparisons.