Intrinsic Coagulation Pathway Research Articles

#435 Endothelial activation and inflammation in chronic kidney disease (CKD): role of neutrophil extracellular traps (NETs)

Abstract Background and Aims Advanced chronic kidney disease (CKD), especially in stages 4-5, amplifies cardiovascular disease (CVD) risk by around 50%. This heightened susceptibility is linked to a 1.5 to 3-fold increase in venous thromboembolism incidence, driven by coagulation dysregulation and heightened inflammation. During inflammation, neutrophils can release extracellular traps (NETs), that was described contribute to endothelial damage, through adhesion molecule elevation, and tissue factor (TF) secretion, exacerbating thrombosis. Additionally, NETs interact with von Willebrand Factor (vWF) and factor XII, further amplifying thrombus formation and the activation of the intrinsic coagulation pathway. The aim of this study was to observe whether NETs may contribute to increased CVD risk in CKD patients by inducing endothelial activation and thromboinflammatory responses to a larger extent compared to NETs from healthy donors. Method For that, neutrophils were isolated from healthy donors’ and CKD patients’ (stages 3-5 HD) peripheral blood, and NETs were collected by stimulating neutrophils with PMA for 3.5 hours. Human aortic endothelial cells (HAoECs) were subsequently exposed to NETs for 3 to 24 hours, followed by gene expression analysis with a focus on inflammation, endothelial activation, and prothrombotic markers. Additionally, protein analysis of the supernatant was conducted using ELISA. All experiments were performed under ethical protocol EK 225-21 from the University Hospital of Aachen. Results Exposure to both healthy NETs as well as the ones isolated from CKD patients resulted in a notable upregulation of endothelial activation (VCAM-1), prothrombotic (TF), and inflammatory (IL-6) gene expression within 3 hours, without difference between healthy and CKD conditions. On the other hand, when examining the cellular supernatant, a significantly higher increase in vWF secretion was observed upon treatment with CKD-NETs compared to healthy NETs. Furthermore, while secretion of the inflammatory cytokine IL-6 presented a comparable increase upon treatment with NETs derived from healthy and CKD neutrophils, IL-1β secretion was markedly enhanced after CKD-NET treatment in comparison to healthy NETs. Conclusion In summary, our findings suggest that NETs exert a significant influence on endothelial cells, contributing to increased thromboinflammatory activation. Notably, CKD-NETs evoked a more rapid impact on the endothelium, accompanied by increased secretion of IL-1β and vWF, and underlying mechanisms will be further investigated.

Anti-Xa/APTT discrepancy in heparinized ImpellaTM-supported cardiogenic shock patients can predict mortality

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Fonds Wetenschappelijk Onderzoek Flanders. Background Meticulous management of unfractionated heparin (UFH) is important to avoid complications in patients receiving percutaneous mechanical circulatory support (pMCS) for cardiogenic shock (CS). Whereas the anti-Xa assay directly measures UFH activity, the activated partial thromboplastin time (APTT) remains the most frequently used test. The APTT covers the intrinsic and common coagulation pathway and can be influenced by various confounding (laboratory) factors during critical illness, unrelated to the patient’s (anti)coagulation state. Purpose We investigated a CS population supported with isolated left-sided Impella™ and UFH-anticoagulation. UFH-levels were guided by anti-Xa with APTT in parallel. Here, we focus on those samples with anti-Xa in-range and discrepantly prolonged parallel APTT-measurements. We differentiated the APTT-prolongation according to reduced fibrinogen levels and/or prolonged INR (factor XI/XII consumption vs. concomitant conditions) and correlated this with mortality during Impella™-support. Methods Between April-2017 and June-2022, all CS patients with isolated left-sided ImpellaTM-support (>24h) and UFH anticoagulation monitored by anti-Xa with APTT in parallel were assessed at two high-volume tertiary institutions. In-range anti-Xa samples (between 0.20-0.30IU/mL or 0.31-0.50IU/mL based on the patient’s profile) were selected. In-range APTT values were considered between 40-55s and 55-80s respectively. Samples with prolonged APTT as compared to the in-range anti-Xa were allocated in 4 groups based on fibrinogen (<1.5g/dL or ≥ 1.5g/dL) and INR (<1.5 or ≥1.5) on the same day as the sample with prolonged APTT. Mortality of patients with normal fibrinogen ànd INR was then compared to those with abnormal fibrinogen and/or INR via a Chi² test. Results Ninety-three ImpellaTM-runs (77 men, 83%) with a median age of 56 years (IQR 46-66) were analyzed. Median support was 6 days (IQR 4-12). Among the 1914 in-range anti-Xa samples, parallel APTT was prolonged in 468 samples (24%) across 59 patients. 80 Samples from 2 patients were excluded due to absence of fibrinogen and/or INR measurements on the same day. The mortality in patients with prolonged APTT but normal fibrinogen levels and normal INR (N=29; due to FXI/FXII-consumption) was significantly lower compared to the remaining 28 patients who showed samples with reduced fibrinogen levels and/or prolonged INR (10% vs. 32%; p=0.043). Conclusions Fibrinogen-levels and INR uncover the underlying etiology in discrepant APTT-prolongation as compared to an in-range anti-Xa value in a CS population during left-sided ImpellaTM support. In patients with prolonged APTT and reduced fibrinogen (<1.5 g/dL) and/or prolonged INR (≥1.5), the risk of mortality is significantly elevated. This is due to concomitant conditions with high mortality (liver failure, sepsis, DIC,…). We advocate to monitor such patients carefully and treat them in a patient/disease tailored approach.

Sulfated polysaccharides from the viscera of Mustelus shark: Characterization and antioxidant, anticoagulant and anti-proliferative activities

In the present study, sulfated polysaccharides (SPs) from smooth hound shark were extracted using different precipitation agents, the cetylpyridinium chloride or ethanol, named SHSP-I and SHSP-II, respectively. The UV–visible scan demonstrated the absence of nucleic acids or proteins in the extracted SHSPs. The antioxidant, anticoagulant and anti-proliferative activities were also investigated. Results showed that SHSPs displayed reducing and scavenging capacities, as shown by the oxygen radical absorbance capacity, the total antioxidant capacity and the hydroxyl radical-scavenging activities. Moreover, extracted SPs were able to prolong the activated partial thromboplastin time (aPTT), the prothrombin time (PT) and the thrombin time (TT) and they reduced the fibrinogen level (FL), without inducing erythrocytes hemolysis, which indicated that they inhibited the intrinsic and extrinsic coagulation pathways and thrombin-mediated fibrin formation. The SHSP-I was more effective on prolonging the blood clotting time by more than 120 s at 1 mg/ml in the aPTT and about 70 s and 60 s at 2.5 mg/ml in the PT and TT, respectively, compared to the SHSP-II. The anti-proliferative activity of SHSPs was tested against the growth of K562 human myelogenous leukemia cells and Caco-2 colon cancer cells. Data showed that both polysaccharides have a dose-dependent inhibitory effect, where SHSP-I exhibited the strongest activity against the tested cells and the K562 was more sensitive than Caco-2 line cells to the action of SHSPs. The overall data suggested that SPs from shark viscera could be useful to be used as nutraceutical agents.

The hemostatic performance and mechanism of palygorskite with structural regulate by oxalic acid gradient leaching

Palygorskite (Pal) is a naturally available one-dimensional clay mineral, featuring rod-shaped morphology, nanoporous structure, permanent negative charges as well as abundant surface hydroxyl groups, exhibiting promising potential as a natural hemostatic material. In this study, the hemostatic performance and mechanisms of Pal were systematically investigated based on the structural regulate induced by oxalic acid (OA) gradient leaching from perspectives of structure, surface attributes and ion release. In vitro and in vivo hemostasis evaluation showed that Pal with OA leaching for 1 h exhibited a superior blood procoagulant effect compared with the raw Pal as well as the others leached for prolonging time. This phenomenon might be ascribed to the synergistic effect of the intact nanorod-like morphology, the increase in the surface negative charge, the release of metal ions (Fe3+ and Mg2+), and the improved blood affinity, which promoted the intrinsic coagulation pathway, the fibrinogenesis and the adhesion of blood cells, thereby accelerating the formation of robust blood clots. This work is expected to provide experimental and theoretical basis for the construction of hemostatic biomaterials based on clay minerals.

IMPACT OF CONTACT SYSTEM ON CARDIOVASCULAR DISEASES

Contact and intrinsic coagulation pathway components are involved in pathological thrombus formation. In addi­tion to coagulation, contact system activation is involved in anti/profibrinolytic and inflammatory processes that can contribute to the atherosclerotic and thrombotic en­vironment, leading to cardiovascular diseases. This lite­rature review concerns the role of the contact system on cardiovascular diseases and accentuates the potential of coagulation factors FXII (FXII) and FXI (FXI) as targets for antithrombotic therapy.

Superior hemostatic and wound-healing properties of tetrastigma polysaccharide

Hemostatic materials that allow fast and convenient hemostasis are urgently needed. However, obtaining an ideal hemostatic agent that exhibits biocompatibility, biodegradability, and hemostatic efficacy remains challenging. Recent studies demonstrate that morphological characteristics of hemostatic materials impact their efficacy. This study reports on the importance of morphological modification of traditional Chinese medicine plant polysaccharides. The hemostatic properties of Three-leaf Qing polysaccharides, optimized in particle (SYQ-P) and sponge (SYQ-S) forms, were investigated in vitro and in vivo. SYQ-S and SYQ-P exhibit excellent biodegradability and biocompatibility, thereby improving fibroblast cell proliferation. The hemostatic mechanism involves stable blood clot formation through intrinsic coagulation pathways. In a lethal mouse liver defect bleeding model, blood loss and clotting time were lower with polysaccharides use than with commercial hemostatic drugs. SYQ-S demonstrates instant water-triggered expansion and high absorption capacity, while the sponge-like morphological characteristics effectively improve blood component concentrations. With efficient hemostasis and excellent biocompatibility, SYQ-S dramatically accelerated wound healing in a full-thickness skin defect model in vivo. Thus, SYQ-S effectively promotes the formation of granulation tissue, collagen deposition, and angiogenesis, which ultimately accelerates wound healing. Our work highlights the design of natural Chinese medicine polysaccharides and presents a promising hemostatic agent for widespread clinical application.

The Prediction of Milk Whey Extract (MWE) Bioactive Compounds Based on Proximate Analysis and its Effects on Blood Coagulation: A New Approach

The present work demonstrates the milk whey extraction, proximate analysis, and biochemical characterization of bioactive compounds in MWE. Effects of MWE on plasma coagulation and platelet aggregation. Proximate analysis of MWE was done according to AOAC. Whey protein banding was confirmed in 12%, 15%, and 18% SDS-PAGEs. A quantitative analysis of bioactive compounds was done. The anticoagulant effects of MWE were tested using plasma recalcification time in both PRP and PPP, further confirmed by PT and APTT assays and in tail bleeding assays at concentrations of 0-100 µg. The non-toxic property of MWE was screened by edema, hemorrhage, and direct hemolytic activities. MWE proximate analysis showed the presence of both micro- and macronutrients. Qualitative analysis confirmed the presence of proteins and carbohydrates. A similar protein-banding pattern was observed in 12%, 15%, and 18% SDS-PAGE’s in both reduced and non-reduced conditions. MWE prolonged the clotting time of human citrated plasma, both PRP and PPP, against the control 210 sec to 770 sec, suggesting its anticoagulant property. MWE delayed the clot progress of only the APTT ratio (8.99 ± 0.09), and PT was not altered, suggesting its role in an intrinsic pathway of blood coagulation. MWE exhibited antiplatelet activity in PRP against ADP and epinephrine-induced platelet aggregation. The percentage of inhibition was 74% and 100% for ADP and epinephrine, respectively. Interestingly, MWE exhibits nontoxic properties, it does not cause hemolysis, hemorrhage, or edema. Milk whey extraction and studies confirm bioactive compounds, and these exhibit antithrombotic, antiplatelet, and non-toxic properties. Hence, MWE not only acts as a good nutritive source but also may prove tobe a therapeutic bioactive compound in the management of cardiovascular disease.

Factor XI: structure, function and therapeutic inhibition.

Arterial and venous thromboembolism is a major medical concern that requires therapeutic anticoagulation in various medical fields to prevent its drastic consequences. Despite significant advances in anticoagulant therapy, thrombosis remains a leading cause of morbidity and mortality worldwide. Traditional anticoagulants like heparin and vitamin K antagonists (VKAs) have shown efficacy in preventing and treating thrombosis but come with an inherent risk of bleeding due to their non-specific inhibition of multiple coagulation factors. Subsequent direct oral anticoagulants (DOACs), targeting specific factors such as Xa or thrombin, demonstrated improved safety profiles compared to VKAs, yet bleeding remains a concern. Accordingly, research is focused on developing anticoagulants with improved safety profiles. A safer class of anticoagulants would have broad appeal. The intrinsic pathway of coagulation, involving factor XI (FXI), has attracted attention as a potential target for safer anticoagulants. Preclinical studies and epidemiological data indicate that FXI deficiency or inhibition protects against thrombosis with minimal bleeding. Current research involves evaluating various FXI-directed strategies, and phase 2 studies have shown promising results in orthopedic surgery, atrial fibrillation, end-stage renal disease (ESRD), myocardial infarction, and ischemic stroke. Several agents, such as antisense oligonucleotides, monoclonal antibodies, small synthetic molecules, natural peptides, and aptamers, have been developed to inhibit FXI at different stages, offering potentially safer alternatives to traditional anticoagulants. However, the optimal balance between preventing thrombosis and the risk of bleeding associated with FXI inhibitors requires validation through extensive phase 3 clinical trials using definite clinical endpoints. Several of such trials are currently underway or planned to define the role of FXI inhibitors in clinical practice and determine the most suitable FXI inhibitor for each specific indication. The current review highlightsthe rationale behind developing FXI inhibitors, presenting the most advanced agents in development, summarizing completed clinical trials, and discussing ongoing research efforts.

A macromolecular cross-linked alginate aerogel with excellent concentrating effect for rapid hemostasis

Alginate-based materials present promising potential for emergency hemostasis due to their excellent properties, such as procoagulant capability, biocompatibility, low immunogenicity, and cost-effectiveness. However, the inherent deficiencies in water solubility and mechanical strength pose a threat to hemostatic efficiency. Here, we innovatively developed a macromolecular cross-linked alginate aerogel based on norbornene- and thiol-functionalized alginates through a combined thiol-ene cross-linking/freeze-drying process. The resulting aerogel features an interconnected macroporous structure with remarkable water-uptake capacity (approximately 9000 % in weight ratio), contributing to efficient blood absorption, while the enhanced mechanical strength of the aerogel ensures stability and durability during the hemostatic process. Comprehensive hemostasis-relevant assays demonstrated that the aerogel possessed outstanding coagulation capability, which is attributed to the synergistic impacts on concentrating effect, platelet enrichment, and intrinsic coagulation pathway. Upon application to in vivo uncontrolled hemorrhage models of tail amputation and hepatic injury, the aerogel demonstrated significantly superior performance compared to commercial alginate hemostatic agent, yielding reductions in clotting time and blood loss of up to 80 % and 85 %, respectively. Collectively, our work illustrated that the alginate porous aerogel overcomes the deficiencies of alginate materials while exhibiting exceptional performance in hemorrhage, rendering it an appealing candidate for rapid hemostasis.

Purification of an Acidic Polysaccharide with Anticoagulant Activity from the Marine Sponge Sarcotragus spinosulus.

Thromboembolic conditions are the most common cause of death in developed countries. Anticoagulant therapy is the treatment of choice, and heparinoids and warfarin are the most adopted drugs. Sulphated polysaccharides extracted from marine organisms have been demonstrated to be effective alternatives, blocking thrombus formation by inhibiting some factors involved in the coagulation cascade. In this study, four acidic glycan fractions from the marine sponge Sarcotragus spinosulus were purified by anion-exchange chromatography, and their anticoagulant properties were investigated through APTT and PT assays and compared with both standard glycosaminoglycans and holothurian sulphated polysaccharides. Moreover, their topographic localization was assessed through histological analysis, and their cytocompatibility was tested on a human fibroblast cell line. A positive correlation between the amount of acid glycans and the inhibitory effect towards both the intrinsic and extrinsic coagulation pathways was observed. The most effective anticoagulant activity was shown by a highly charged fraction, which accounted for almost half (about 40%) of the total hexuronate-containing polysaccharides. Its preliminary structural characterization, performed through infrared spectroscopy and nuclear magnetic resonance, suggested that it may consist of a fucosylated chondroitin sulphate, whose unique structure may be responsible for the anticoagulant activity reported herein for the first time.

Anti-hemostatic, antithrombotic, and chemical profiles of a curly-leaf variety of Petroselinum crispum (Apiaceae), a food and medicinal aromatic herb

Thrombosis is currently among the major causes of morbidity and mortality in the World. New prevention and therapy alternatives have been increasingly sought in medicinal plants. In this context, we have been investigating parsley, Petroselinum crispum (Mill.) Nym, an aromatic herb with two leaf varieties. We report here the in vitro, in vivo, and ex vivo anti-hemostatic and antithrombotic activities of a parsley curly-leaf variety. Aqueous extracts of aerial parts (PCC-AP), stems (PCC-S), and leaves (PCC-L) showed significant in vitro antiplatelet activity. PCC-AP extract exhibited the highest activity (IC50 2.92 mg/mL) when using ADP and collagen as agonists. All extracts also presented in vitro anticoagulant activity (APTT and PT) and anti-thrombogenic activity. PCC-S was the most active, with more significant interference in the factors of the intrinsic coagulation pathway. The oral administration of PCC-AP extract in rats caused a greater inhibitory activity in the deep vein thrombi (50%; 65 mg/kg) than in arterial thrombi formation (50%; 200 mg/kg), without cumulative effect after consecutive five-day administration. PCC-AP extract was safe in the induced bleeding time test. Its anti-aggregating profile was similar in ex vivo and in vitro conditions but was more effective in the extrinsic pathway when compared to in vitro results. Apiin and coumaric acid derivatives are the main compounds in PCC-AP according to the HPLC-DAD-ESI-MS/MS profile. We demonstrated for the first time that extracts from different parts of curly parsley have significant antiplatelet, anticoagulant, and antithrombotic activity without inducing hemorrhage, proving its potential as a source of antithrombotic compounds.

Adverse effects of microparticles on transfusion of stored red blood cell concentrates

BackgroundSystemic and pulmonary coagulopathy and inflammation are important characteristics of transfusion-related acute lung injury (TRALI). Whether microparticles that accumulate in transfused red blood cell concentrates (RBCs) have proinflammatory and procoagulant potential and contribute to adverse reactions of RBC transfusions is unclear. AimTo investigate the ability of microparticles in stored RBCs to promote thrombin generation and induce human pulmonary microvascular endothelial cell (HMVEC) activation and damage. MethodsThe number and size of microparticles were determined by flow cytometric and nanoparticle tracking analyses, respectively. Thrombin generation and the intrinsic coagulation pathway were assayed by a calibrated automated thrombogram and by measuring activated partial thromboplastin time (aPTT), respectively. The expression of ICAM-1 and the release of cytokines by endothelial cells were detected by flow cytometric analyses. HMVEC damage was assessed by incubating lipopolysaccharide-activated endothelial cells with MP-primed polymorphonuclear neutrophils (PMNs). ResultsThe size of the microparticles in the RBC supernatant was approximately 100–300 nm. Microparticles promoted thrombin generation in a dose-dependent manner and the aPTT was shortened. Depleting microparticles from the supernatant of RBCs stored for 35 days by either filtration or centrifugation significantly decreased the promotion of thrombin generation. The expression of ICAM-1 on HMVECs was increased significantly by incubation with isolated microparticles. Furthermore, microparticles induced the release of interleukin-6 (IL-6) and interleukin-8 (IL-8) from HMVECs. Microparticles induced lipopolysaccharide-activated HMVEC damage by priming PMNs, but this effect was prevented by inhibiting the PMNs respiratory burst with apocynin. ConclusionMicroparticles in stored RBCs promote thrombin generation, HMVEC activation and damage which may be involved in TRALI development.

The pathogenesis of cancer-associated thrombosis.

Patients with cancer have a higher risk of venous thromboembolism (VTE), including deep vein thrombosis (DVT) and pulmonary embolism (PE), compared to the general population. Cancer-associated thrombosis (CAT) is a thrombotic event that occurs as a complication of cancer or cancer therapy. Major factors determining VTE risk in cancer patients include not only treatment history and patient characteristics, but also cancer type and site. Cancer types can be broadly divided into three groups based on VTE risk: high risk (pancreatic, ovarian, brain, stomach, gynecologic, and hematologic), intermediate risk (colon and lung), and low risk (breast and prostate). This implies that the mechanism of VTE differs between cancer types and that specific VTE pathways may exist for different cancer types. This review summarizes the specific pathways that contribute to VTE in cancer patients, with a particular focus on leukocytosis, neutrophil extracellular traps (NETs), tissue factor (TF), thrombocytosis, podoplanin (PDPN), plasminogen activator inhibitor-1 (PAI-1), the intrinsic coagulation pathway, and von Willebrand factor (VWF).

Anticoagulation with osocimab in patients with kidney failure undergoing hemodialysis: a randomized phase 2 trial

Individuals with kidney failure undergoing hemodialysis are at elevated risk for thromboembolic events. Factor (F) XI, which is in the intrinsic pathway of coagulation, is emerging as an attractive target for new anticoagulants that may be safer than existing agents. Osocimab—an inhibitory FXIa antibody—is a potential treatment option for such patients. We conducted a phase 2b, double-blind, placebo-controlled trial, in which 704 participants (448 male, 256 female) with kidney failure undergoing hemodialysis were randomized to receive lower- or higher-dose osocimab or placebo. In total, 686 participants (436 male, 250 female) received treatment for ≤18 months (planned minimal treatment period of 6 months). The co-primary outcomes were clinically relevant bleeding (a composite of major and clinically relevant nonmajor bleeding) and a composite of the incidence of moderate, severe or serious adverse events. Clinically relevant bleeding occurred in 16/232 (6.9%) and 11/224 (4.9%) participants who received lower- and higher-dose osocimab, respectively, and in 18/230 participants (7.8%) who received a placebo. For the composite adverse event endpoint, incidences were 51%, 47% and 43% in the lower-dose osocimab, higher-dose osocimab and placebo groups, respectively. These results suggest that osocimab is associated with a low risk of bleeding and is generally well tolerated in this population; findings that require confirmation in larger trials. ClinicalTrials.gov identifier, NCT04523220.

Bioactive glass incorporated dressing matrix for rapid hemostatic action with antibacterial activity

Uncontrolled bleeding stands as a leading cause of preventable death in both civilian trauma and military battlefield scenarios. Existing hemostatic dressings like HemCon, Celox, and QuickClot, often lack instant hemostasis at the bleeding site and may have biodegradability and exothermic issues. To address the above in this communication, we have synthesized a hemostatic glass (acronym AlBG, CaO-SiO2 system, incorporated with Al2O3 and ZnO) by sol gel route and carried out detail physicochemical characterizations, e.g., XRD, FESEM, FTIR, BET and particle size analysis etc. The AlBG of particle size range 140–253 nm was incorporated into the nonwoven surgical cotton gauze to obtain AlBGscg. In vitro biological assays including cytocomtability, hemocompatibility assays of the above coated gauze was undertaken using NIH3T3 of which, the later showed optimum hemocompatibility with <6 % lysis of red blood cells. Potent antibacterial action on both gram positive and gram negative strains were obtained with significant zone of inhibition of 17 ± 0.34 mm and 16 ± 0.56 mm, respectively. Importantly, significant reduction of the clotting time (31.81 ± 0.12 % for P time and 33.3 ± 0.23 % for aPTT) compared to the control group, indicated activation of both intrinsic and extrinsic coagulation pathways. Scanning electron microscope (SEM) images exhibited an enhanced in vitro blood clot formation in case of AlBGscg in comparison to the control (uncoated surgical cotton gauze), visually. Additionally, a noticeable reduction of ∼30 % in the time required for the blood clot formation, in case of AlBGscg, was observed. The in vivo hemostatic potential of AlBGscg was evaluated in Wistar rats through femoral artery injury and was compared with commercially available surgical cotton gauze. The results demonstrated substantial reduction in the time required for hemostasis (approximately 50 % reduction), a lower amount of blood loss (7.33 % with AlBGscg compared to 22.31 % with surgical cotton gauze), and importantly, there were no incidents of re-bleeding observed with AlBGscg, highlighting its promise as a candidate for effective hemostasis.

The intrinsic coagulation pathway plays a dominant role in driving hypercoagulability in ANCA-associated vasculitis

AbstractThe risk of a venous thrombotic event (VTE) is increased in patients with antineutrophil cytoplasmic antibody (ANCA)–associated vasculitis (AAV); however, a detailed understanding of the underlying mechanisms of hypercoagulability is limited. We assessed prospectively different coagulation parameters in 71 patients with active AAV at baseline and after 6 months of follow-up. D-dimers and fibrinogen were increased in most patients at presentation and remained elevated in half of the patients. Particularly, thrombin-antithrombin (T:AT) complex and activated coagulation factors in complex with their natural inhibitors of the intrinsic coagulation pathway (ie, activated FXII:C1 esterase inhibitor [FXIIa:C1Inh], FXIa:AT, and FXIa:alpha1-antitrypsin [FXIa:α1AT]) were profoundly elevated in patients at baseline. Thrombin formation was dominantly correlated with coagulation factors of the intrinsic pathway (ie, FXIIa:AT, FXIa:AT, FXIa:α1AT, and FXIa:C1Inh) compared to the extrinsic pathway (ie, FVIIa:AT). Hypercoagulability correlated with higher disease activity, ANCA levels, C-reactive protein, serum creatinine, and proteinuria. VTEs were observed in 5 out of 71 (7%) patients within 1 month (interquartile range, 1-5) after inclusion. Baseline T:AT levels were significantly higher in patients with VTE than in those without VTE (P = .044), but other clinical or laboratory markers were comparable between both groups. Hypercoagulability is dominantly characterized by activation of the intrinsic coagulation pathway and elevated D-dimers in active AAV. The driving factors of hypercoagulability are yet to be studied but are most likely related to an interplay of increased disease activity, vascular inflammation, and endothelial damage. Future targets for intervention could include inhibitors of the intrinsic coagulation pathway and compounds specifically reducing the hyperinflammatory state.

Structural Characterization and Anticoagulant Activities of a Keratan Sulfate-like Polysaccharide from the Sea Cucumber Holothuria fuscopunctata.

A sulfated polysaccharide (AG) was extracted and isolated from the sea cucumber H. fuscopunctata, consisting of GlcNAc, GalNAc, Gal, Fuc and lacking any uronic acid residues. Importantly, several chemical depolymerization methods were used to elucidate the structure of the AG through a bottom-up strategy. A highly sulfated galactose (oAG-1) and two disaccharides labeled with 2,5-anhydro-D-mannose (oAG-2, oAG-3) were obtained from the deaminative depolymerized product along with the structures of the disaccharide derivatives (oAG-4~oAG-6) identified from the free radical depolymerized product, suggesting that the repeating building blocks in a natural AG should comprise the disaccharide β-D-GalS-1,4-D-GlcNAc6S. The possible disaccharide side chains (bAG-1) were obtained with mild acid hydrolysis. Thus, a natural AG may consist of a keratan sulfate-like (KS-like) glycosaminoglycan with diverse modifications, including the sulfation types of the Gal residue and the possible disaccharide branches α-D-GalNAc4S6S-1,2-α/β-L-Fuc3S linked to the KS-like chain. Additionally, the anticoagulant activities of the AG and its depolymerized products (dAG1-9) were evaluated in vitro using normal human plasma. The AG could prolong activated partial thromboplastin time (APTT) in a dose-dependent manner, and the activity potency was positively related to the chain length. The AG and dAG1-dAG3 could prolong thrombin time (TT), while they had little effect on prothrombin time (PT). The results indicate that the AG could inhibit the intrinsic and common coagulation pathways.

Chitosan nonwoven fabric composited calcium alginate and adenosine diphosphate as a hemostatic bandage for acute bleeding wounds

Acute bleeding following accidental injury is a leading cause of mortality. However, conventional hemostatic bandages impede wound healing by inducing excessive blood loss, dehydration, and adherence to granulation tissue. Strategies such as incorporating active hemostatic agents and implementing chemical modifications can augment the properties of these bandages. Nevertheless, the presence of remote thrombosis and initiators may pose risks to human health. Here, a hemostatic bandage was developed by physically combined chitosan nonwoven fabric, calcium alginate sponge, and adenosine diphosphate. The presented hemostatic bandage not only exhibits active and passive mechanisms for promoting clotting but also demonstrates excellent mechanical properties, breathability, ease of removal without causing damage to the wound bed or surrounding tissues, as well as maintaining an optimal moist environment conducive to wound healing. In vitro evaluation results indicated that the hemostatic bandage possesses favorable cytocompatibility with low levels of hemolysis. Furthermore, it effectively aggregates various blood cells while activating platelets synergistically to promote both extrinsic and intrinsic coagulation pathways. In an in vivo rat model study involving liver laceration and femoral artery injury scenarios, our developed hemostatic bandage demonstrated rapid clot formation capabilities along with reduced blood loss compared to commercially available fabrics.

Robust adhesive nanocomposite sponge composed of citric acid and nano clays modified cellulose for rapid hemostasis of lethal non-compressible hemorrhage

Massive bleeding control plays the main role in saving people's lives in emergency situations. Herein, modified cellulose-based nanocomposite sponges by polydopamine (PDA) and laponite nano-clay was developed to sturdily deal with non-compressible lethal severe bleeding. PDA accomplishes supreme adhesion in the bleeding site (∼405 kPa) to form strong physical barrier and seal the position. Sponges super porous (∼70 % porosity) and super absorbent capacity (48 g blood absorbed per 1 g sponge) by concentrating the blood cells and platelets provides the requirements for primary hemostasis. Synergistically, the nanocomposite sponges' intelligent chemical structure induces hemostasis by activation of the XI, IX, X, II and FVII factors of intrinsic and extrinsic coagulation pathways. Excellent hemostatic performance of sponges in-vitro was assessed by RBC accumulation (∼100 %), blood clotting index (∼10 %), platelet aggregation/activation (∼93 %) and clotting time. The nanocomposite sponges depicted super performance in the fatal high-pressure non-compressible hemorrhage model by reducing of >2, 15 and 3 times in the bleeding amount at New Zealand rabbit's heart and liver, and rat's femoral artery bleeding models, respectively compared to commercial hemostatic agents (Pvalue˂0.001). The in-vivo host response results exhibited biosafety with no systemic and significant local inflammatory response by hematological, pathological and biochemical parameters assessments.

Structural characterization of a water-soluble acidic polysaccharide CSP-IV with potential anticoagulant activity from fruit pulp of Clausena lansium (Lour.) Skeels Guifei

Four main water-soluble wampee fruit pulp polysaccharides, named CSP-I, CSP-II, CSP-III and CSP-IV, were isolated from Clausena lansium (Lour.) Skeels Guifei, therein CSP-IV content was higher than the others. All components possess certain anticoagulant activity demonstrated by prolonged activated partial thromboplastin time, especially CSP-IV, which suggests that CSP-IV plays anticoagulant effect through disturbing intrinsic coagulation pathway. The wampee polysaccharide CSP-IV with Mw of 510.1 kDa was mainly composed of Gal, Ara and GalA. Backbone of CSP-IV contains Gal, Ara and GalA, two kinds of side chains contain one monosaccharide Gal or Ara, both branch on Gal residue of backbone. CSP-IV has no the conformation of triple helix demonstrated by Congo red test. These results showed that CSP-IV is an acidic polysaccharide with potential anticoagulant activity via targeting intrinsic coagulation pathway.