Kinetic analysis of prothrombinase assembly and substrate delivery mechanisms.

Prothrombinase complex, composed of coagulation factors Xa (FXa) and Va (FVa) is a major enzyme of the blood coagulation network that produces thrombin via activation of its inactive precursor prothrombin (FII) on the surface of phospholipid membranes. However, pathways and mechanisms of prothrombinase formation and substrate delivery are still discussed. Here we designed a novel mathematical model that considered different potential pathways of FXa or FII binding (from the membrane or from solution) and analyzed the kinetics of thrombin formation in the presence of a wide range of reactants concentrations. We observed the inhibitory effect of large FVa concentrations and this effect was phospholipid concentration-dependent. We predicted that efficient FII activation occurred via formation of the ternary complex, in which FVa, FXa and FII were in the membrane-bound state. Prothrombin delivery was mostly membrane-dependent, but delivery from solution was predominant under conditions of phospholipid deficiency or FXa/FVa excess. Likewise, FXa delivery from solution was predominant in the case of FVa excess, but high FII did not switch the FXa delivery to the solution-dependent one. Additionally, the FXa delivery pathway did not depend on the phospholipid concentration, being the membrane-dependent one even in case of the phospholipid deficiency. These results suggest a flexible mechanism of prothrombinase functioning which utilizes different complex formation and even inhibitory mechanisms depending on conditions.

The Potential of Sugarcane Waste-Derived Cellulose Fibres as Haemostatic Agents.

Haemorrhage control during surgery and following traumatic injury remains a critical, life-saving challenge. Cellulose products are already employed in commercially available haemostatic dressings. This work explores sourcing cellulose from sugarcane trash pulp to produce micro- and nanosized fibres with hydroxyl, carboxylic acid, and trimethylamine functional groups, resulting in either positive or negative surface charges. This paper assesses the influence of these fibres on multiple blood clotting parameters in both dispersed solutions and dry gauze applications. In vitro blood clotting studies demonstrated the significant haemostatic potential of cellulose fibres derived from sugarcane waste to initiate clotting. Plasma absorbance assays showed that the 0.25 mg/mL cellulose microfibre dispersion had the highest clotting performance. It was observed that no single property of surface charge, functionality, or fibre morphology exclusively controlled the clotting initiation measured. Instead, a combination of these factors affected clot formation, with negatively charged cellulose microfibres comprising hydroxyl surface groups providing the most promising result, accelerating the coagulation cascade mechanism by 67% compared to the endogenous activity. This difference in clot initiation shows the potential for the non-wood agricultural waste source of cellulose in haemostatic wound healing applications, contributing to the broader understanding of cellulose-based materials' versatility and their applications in biomedicine.

Generation of Rapid and High-Quality Serum by Recombinant Prothrombin Activator Ecarin (RAPClot™).

We recently reported the potential application of recombinant prothrombin activator ecarin (RAPClot™) in blood diagnostics. In a new study, we describe RAPClot™ as an additive to develop a novel blood collection prototype tube that produces the highest quality serum for accurate biochemical analyte determination. The drying process of the RAPClot™ tube generated minimal effect on the enzymatic activity of the prothrombin activator. According to the bioassays of thrombin activity and plasma clotting, γ-radiation (>25 kGy) resulted in a 30-40% loss of the enzymatic activity of the RAPClot™ tubes. However, a visual blood clotting assay revealed that the γ-radiation-sterilized RAPClot™ tubes showed a high capacity for clotting high-dose heparinized blood (8 U/mL) within 5 min. This was confirmed using Thrombelastography (TEG), indicating full clotting efficiency under anticoagulant conditions. The storage of the RAPClot™ tubes at room temperature (RT) for greater than 12 months resulted in the retention of efficient and effective clotting activity for heparinized blood in 342 s. Furthermore, the enzymatic activity of the RAPClot™ tubes sterilized with an electron-beam (EB) was significantly greater than that with γ-radiation. The EB-sterilized RAPClot™ tubes stored at RT for 251 days retained over 70% enzyme activity and clotted the heparinized blood in 340 s after 682 days. Preliminary clinical studies revealed in the two trials that 5 common analytes (K, Glu, lactate dehydrogenase (LD), Fe, and Phos) or 33 analytes determined in the second study in the γ-sterilized RAPClot™ tubes were similar to those in commercial tubes. In conclusion, the findings indicate that the novel RAPClot™ blood collection prototype tube has a significant advantage over current serum or lithium heparin plasma tubes for routine use in measuring biochemical analytes, confirming a promising application of RAPClot™ in clinical medicine.

Injectable and rapidly expandable thrombin-decorated cryogels achieve rapid hemostasis and high survival rates in a swine model of lethal junctional hemorrhage.

Effective therapies are urgently needed to stabilize patients with marginally compressible junctional hemorrhage long enough to get them to the hospital alive. Herein, we report injectable and rapidly expandable cryogels consisting of polyacrylamide and thrombin (AT cryogels) created by cryo-polymerization for the efficient management of lethal junctional hemorrhage in swine. The produced cryogels have small pore sizes and highly interconnected porous architecture with robust mechanical strength. The cryogels exhibit rapid shape memory properties and prove to be resilient against fatigue. These cryogels also show high water/blood absorption capacity, fast blood clotting effect, and enhanced adhesion of red blood cells and platelets in vitro. Further, in vivo, hemostatic efficacy tests in a lethal swine junctional hemorrhage model suggest that treatment with AT cryogels, especially AT-2 cryogels, achieves the least blood loss and the highest survival rate (100 %) compared to currently employed products such as XStat® and combat gauze. The high hemostatic performance of the cryogels may be attributed to highly interconnected porous architecture with small pore size and the use of thrombin as a pro-coagulant agent. Collectively, injectable and rapidly expandable thrombin-decorated polyacrylamide-based cryogels show significant promise as hemostatic material, offering effective management of marginally compressible junctional hemorrhages in prehospital settings.

A novel technique to quantify the kinetics of blood clot contraction based on the expulsion of fluorescently labeled albumin into serum.

BACKGROUND: The platelet-driven contraction or retraction of blood clots has been utilized to obtain blood serum for laboratory studies, but now, in vitro clot contraction assays are used in research laboratories and clinics to assess platelet functionality. The static final extent of clot contraction measured using a clot size or expelled serum volume can be supplemented substantially with a dynamic analysis. OBJECTIVES: To provide a step-by-step protocol for a relatively simple and affordable new automated methodology to follow the kinetics of blood clot contraction, which allows for simultaneous measurements of various samples at a time and requires only a fluorescence plate reader. METHODS: The kinetics of clot contraction in whole blood was assessed by continuously detecting the fluorescence intensity of fluorescein isothiocyanate-albumin added to a blood sample before clotting and expelled into the serum during clot shrinkage. RESULTS: The clots are formed and fluorescence is measured in the wells of a black multiwell plate using a standard plate fluorescent reader. The specificity of this technique for clot contraction has been demonstrated by the strong inhibitory effects of blebbistatin, latrunculin A, and abciximab. To validate the new technique, increased fluorescence intensity in the contracting clots was measured in parallel with a visual decrease in clot size performed with the same blood samples. CONCLUSION: The resulting clot contraction dynamics based on the expulsion of fluorescein isothiocyanate-albumin can be quantified using a number of kinetic parameters as well as a phase kinetics analysis. The advantages and drawbacks of the new technique are discussed.

Whole blood clotting time: Current practices among Thailand National External Quality Assessment Scheme for blood coagulation member laboratories.

INTRODUCTION: The Thai National Guidelines for Hemostatic Laboratory Testing were established in 2018. The guidelines recommend that the 20-min whole blood clotting time (20WBCT) method be used to diagnose/monitor snake bites. The aim of this study was to survey members of the Thailand National External Quality Assessment Scheme (NEQAS) for Blood Coagulation to investigate the use of 20WBCT testing compared between the 2021 post-guideline and 2007 pre-guideline periods. METHODS: In July 2021, questionnaires were sent from the Faculty of Medicine Siriraj Hospital, Mahidol University to 521 Thailand NEQAS for Blood Coagulation member laboratories to survey their WBCT practices. Current WBCT practices were compared with pre-guideline WBCT practices, and chi-square test (x2) was used to test for differences between groups. RESULTS: Ninety-seven (18.6%) of 521 surveys were returned. Seventy-one laboratories (73.2%) reported knowing about 20WBCT from the Thai national guidelines. The reported average frequency of overall WBCT testing in 2021 was 12.4 times/month. The proportion of laboratories that reported using the 20WBCT test increased from 2.0% in 2007 to 46.4% in 2021 (p < 0.001), and the indications for performing WBCT were virtually unchanged from 2007 to 2021. The proportion of laboratories that reported having problems with WBCT testing decreased from 32.7% in 2007 to 16.5% in 2021. CONCLUSION: Despite our findings that almost three-quarters of respondent laboratories reported knowing about 20WBCT testing from the WBCT guidelines, and that WBCT-specific problems decreased significantly from 2007 to 2021, more work and training is needed to improve WBCT guideline dissemination, understanding, and adherence in Thailand.

Antibiofilm and antithrombotic hydrogel coating based on superhydrophilic zwitterionic carboxymethyl chitosan for blood-contacting devices.

Blood-contacting devices must be designed to minimize the risk of bloodstream-associated infections, thrombosis, and intimal lesions caused by surface friction. However, achieving effective prevention of both bloodstream-associated infections and thrombosis poses a challenge due to the conflicting nature of antibacterial and antithrombotic activities, specifically regarding electrostatic interactions. This study introduced a novel biocompatible hydrogel of sodium alginate and zwitterionic carboxymethyl chitosan (ZW@CMC) with antibacterial and antithrombotic activities for use in catheters. The ZW@CMC hydrogel demonstrates a superhydrophilic surface and good hygroscopic properties, which facilitate the formation of a stable hydration layer with low friction. The zwitterionic-functionalized CMC incorporates an additional negative sulfone group and increased negative charge density in the carboxyl group. This augmentation enhances electrostatic repulsion and facilitates the formation of hydration layer. This leads to exceptional prevention of blood clotting factor adhesion and inhibition of biofilm formation. Subsequently, the ZW@CMC hydrogel exhibited biocompatibility with tests of in vitro cytotoxicity, hemolysis, and catheter friction. Furthermore, in vivo tests of antithrombotic and systemic inflammation models with catheterization indicated that ZW@CMC has significant advantages for practical applications in cardiovascular-related and sepsis treatment. This study opens a new avenue for the development of chitosan-based multifunctional hydrogel for applications in blood-contacting devices.

Advances in the cellular biology, biochemistry, and molecular biology of acidocalcisomes.

SUMMARYAcidocalcisomes are organelles conserved during evolution and closely related to the so-called volutin granules of bacteria and archaea, to the acidocalcisome-like vacuoles of yeasts, and to the lysosome-related organelles of animal species. All these organelles have in common their acidity and high content of polyphosphate and calcium. They are characterized by a variety of functions from storage of phosphorus and calcium to roles in Ca2+ signaling, osmoregulation, blood coagulation, and inflammation. They interact with other organelles through membrane contact sites or by fusion, and have several enzymes, pumps, transporters, and channels.

Validation of the time to attain maximal clot amplitude after reaching maximal clot formation velocity parameter as a measure of fibrinolysis using rotational thromboelastometry and its application in the assessment of fibrinolytic resistance in septic patients: a prospective observational study: communication from the ISTH SSC Subcommittee on Fibrinolysis.

BACKGROUND: In sepsis, fibrinolysis resistance correlates with worse outcomes. Practically, rotational thromboelastometry (ROTEM) is used to report residual clot amplitude relative to maximum amplitude at specified times after clot formation clot lysis indices (CLIs). However, healthy individuals can exhibit similar CLIs, thus making it challenging to solely diagnose the low fibrinolytic state. Furthermore, CLI does not include the kinetics of clot formation, which can affect overall fibrinolysis. Therefore, a more nuanced analysis, such as time to attain maximal clot amplitude after reaching maximal clot formation velocity (t-AUCi), is needed to better identify fibrinolysis resistance in sepsis. OBJECTIVES: To evaluate the correlation between the degree of fibrinolytic activation and t-AUCi in healthy or septic individuals. METHODS: Whole blood (n = 60) from septic or healthy donors was analyzed using tissue factor-activated (EXTEM) and nonactivated (NATEM) ROTEM assays. Lysis was initiated with tissue-type plasminogen activator, and CLI and t-AUCi were calculated. Standard coagulation tests and plasma fibrinolysis markers (D-dimer, plasmin-α2-antiplasmin complex, plasminogen activator inhibitor type 1, and plasminogen) were also measured. RESULTS: t-AUCi values decreased with increasing fibrinolytic activity and correlated positively with CLI for different degrees of clot lysis both in EXTEM and NATEM. t-AUCi cutoff value of 1962.0 seconds in EXTEM predicted low fibrinolytic activity with 81.8% sensitivity and 83.7% specificity. In addition, t-AUCi is not influenced by clot retraction. CONCLUSION: Whole-blood point-of-care ROTEM analyses with t-AUCi offers a more rapid and parametric evaluation of fibrinolytic potential compared with CLI, which can be used for a more rapid and accurate diagnosis of fibrinolysis resistance in sepsis.

clotFoam: An open-source framework to simulate blood clot formation under arterial flow.

Blood clotting involves the coupled processes of platelet aggregation and coagulation. Simulating clotting under flow in complex geometries is challenging due to multiple temporal and spatial scales and high computational cost. clotFoam is an open-source software developed in OpenFOAM that employs a continuum model of platelet advection, diffusion, and aggregation in a dynamic fluid environment and a simplified coagulation model with proteins that advect, diffuse, and react within the fluid and with wall-bound species through reactive boundary conditions. Our framework provides the foundation on which one can build more complex models and perform reliable simulations in almost any computational domain.

A Transparent, Tough and Self-Healable Biopolymeric Composites Hydrogel for Open Wound Management.

Modern healthcare engineering requires a wound dressing solution supported by materials with outstanding features such as high biological compatibility, strong mechanical strength, and higher transparency with effective antibacterial properties. Here, we present a unique hydrogel technology consisting of two negatively charged biopolymers and a positively charged synthetic polymer. The interaction between charged polymers through hydrogen bonds has been created, which are revealed in the simulation by density functional theory and Fourier transform infrared spectra of individual polymers and the hydrogel film. The transparent hydrogel film dressings showed excellent stretchability, a higher water swelling ratio (60%), and strong mechanical strength (∼100 MPa) with self-healing abilities (85-90%). The fabricated hydrogel film showed stable blood clots (within 119 ± 15 s) with rapid hemostasis (<2%) properties and effective antibacterial studies against E. coli and S. aureus bacterial strains. In addition, the obtained hydrogel film also showed excellent cell viability on mouse fibroblast cells. With their enormous amenability to modification, these hydrogel films may serve as promising biomaterials for wound dressing applications.

Activation of Piezo1 channels in compressed red blood cells augments platelet-driven contraction of blood clots.

BACKGROUND: Piezo1 is a mechanosensitive cationic channel that boosts intracellular [Ca2+]i. Compression of red blood cells (RBCs) during platelet-driven contraction of blood clots may cause the activation of Piezo1. OBJECTIVES: To establish relationships between Piezo1 activity and blood clot contraction. METHODS: Effects of a Piezo1 agonist, Yoda1, and antagonist, GsMTx-4, on clot contraction in vitro were studied in human blood containing physiological [Ca2+]. Clot contraction was induced by exogenous thrombin. Activation of Piezo1 was assessed by Ca2+ influx in RBCs and with other functional and morphologic features. RESULTS: Piezo1 channels in compressed RBCs are activated naturally during blood clot contraction and induce an upsurge in the intracellular [Ca2+]i, followed by phosphatidylserine exposure. Adding the Piezo1 agonist Yoda1 to whole blood increased the extent of clot contraction due to Ca2+-dependent volumetric shrinkage of RBCs and increased platelet contractility due to their hyperactivation by the enhanced generation of endogenous thrombin on activated RBCs. Addition of rivaroxaban, the inhibitor of thrombin formation, or elimination of Ca2+ from the extracellular space abrogated the stimulating effect of Yoda1 on clot contraction. The Piezo1 antagonist, GsMTx-4, caused a decrease in the extent of clot contraction relative to the control both in whole blood and in platelet-rich plasma. Activated Piezo1 in compressed and deformed RBCs amplified the platelet contractility as a positive feedback mechanism during clot contraction. CONCLUSION: The results obtained demonstrate that the Piezo1 channel expressed on RBCs comprises a mechanochemical modulator of blood clotting that may be considered a potential therapeutic target to correct hemostatic disorders.

Comparing modified Lee and White method against 20-minute whole blood clotting test as bedside coagulation screening test in snake envenomation victims.

Background: Twenty-minute whole blood clotting test (20WBCT) and Modified Lee and White (MLW) method are the most routinely employed bedside tests for detecting coagulopathic snake envenomation. Our study compared the diagnostic utility of MLW and 20WBCT for snakebite victims at a tertiary care hospital in Central Kerala, South India. Methods: This single-center study recruited 267 patients admitted with snake bites. 20WBCT and MLW were performed simultaneously at admission along with the measurement of Prothrombin Time (PT). The diagnostic utility of 20WBCT and MLW was determined by comparing the sensitivity (Sn), specificity (Sp), positive and negative predictive values, likelihood ratios, and accuracy at admission with an INR value > 1.4. Results: Out of 267 patients, 20 (7.5%) patients had VICC. Amongst those who had venom-induced consumption coagulopathy (VICC), MLW was prolonged for 17 patients, (Sn 85% 95% confidence interval [CI]: 61.1-96.0) whereas 20WBCT was abnormal for 11 patients (Sn 55%, 95% CI: 32.04-76.17). MLW and 20WBCT were falsely positive for the same patient (Sp 99.6%, 95% CI: 97.4-99.9%). Conclusion: MLW is more sensitive than 20WBCT to detect coagulopathy at the bedside amongst snakebite victims. However, further studies are necessary for standardizing bedside coagulation tests in snakebite cases.

Clotted blood samples in the neonatal intensive care unit: A retrospective, observational study to evaluate interventions to reduce blood sample clotting.

BACKGROUND: Blood draws for laboratory investigations are essential for patient management in neonatal intensive care units (NICU). When blood samples clot before analysis, they are rejected, which delays treatment decisions and necessitates repeated sampling. AIMS: To decrease the incidence of rejected blood samples taken for laboratory investigation as a result of clotted sample. STUDY DESIGN: This retrospective observational study used routine data on blood draws from preterm infants collected between January 2017 and June 2019 in a 112-cot NICU in Qatar. Quality improvement interventions to reduce the rate of clotted blood samples included: awareness raising and safe sampling workshops with NICU staff, involvement of the neonatal vascular access team, development of a complete blood count (CBC) sample collection pathway, review of sample collection equipment, introducing the Tenderfoot® heel lance, establishment of benchmarks and provision of dedicated blood extraction equipment. RESULTS: First attempt blood draw occurred in 10 706 cases, representing a 96.2% success rate. In 427 (3.8%) cases, the samples were clotted requiring repeat collection. The overall rate of clotted specimens decreased from 4.8% in 2017 and 2018 to 2.4% in 2019, with odds ratios of 1.42 (95% confidence interval [CI] 1.13-1.78, p = .002), 1.46 (95% CI 1.17-1.81, p < .001) and 0.49 (95% CI 0.39-0.63, p < .001), respectively. The majority (87%-95%) of blood samples were by venepuncture using an intravenous (IV) catheter or the NeoSafe™ blood sampling device. Heel prick sampling was the second (2%-9%) most common method. Clotted samples were most frequently associated with needle use, 228 of 427 (53%), and IV cannula, 162 of 427 (38%), with odds ratios of 4.14 (95% CI 3.34-5.13, p < .001) and 3.11 (95% CI 2.51-3.86, p < .001), respectively. CONCLUSIONS: Our interventions over 3 years were associated with reduced rates of sample rejection due to clotting, and this led to improved patient experience through fewer repeated samplings. RELEVANCE TO CLINICAL PRACTICE: The insights gained from this project can help to improve patient care. Interventions that reduce the rate of blood sample rejection by clinical laboratories can lead to economic savings, timelier diagnostic and treatment decisions, and contribute to an improved quality care experience for all critical care patients, irrespective of age, by reducing the need for repeated phlebotomy and the risk of related complications.

clotFoam: An Open-Source Framework to Simulate Blood Clot Formation Under Arterial Flow.

Blood clotting involves the coupled processes of platelet aggregation and coagulation. Simulating clotting under flow in complex geometries is challenging due to multiple temporal and spatial scales and high computational cost. clotFoam is an open-source software developed in OpenFOAM that employs a continuum model of platelet advection, diffusion, and aggregation in a dynamic fluid environment and a simplified coagulation model with proteins that advect, diffuse, and react within the fluid and with wall-bound species through reactive boundary conditions. Our framework provides the foundation on which one can build more complex models and perform reliable simulations in almost any computational domain.

Primary hemostasis dysfunctions and bleeding risk in newly diagnosed acute myeloid leukemia.

BACKGROUND: Acute myeloid leukaemia carries the risk of complications associated with dysfunctions in haemostasis system. The purpose of this study was to investigate the factors associated with the risk of bleeding in patients with newly diagnosed acute myeloid leukaemia (AML). METHODS: This study involved the methods of immunoenzymatic analysis and classical coagulation studies. The number of biochemical parameters important for establishing coagulative dysfunction in acute myeloid leukaemia was determined, the main ones being the level of von Willebrand factor, the Ristocetin-cofactor activity of von Willebrand factor and factor VIII activity, prothrombin time, platelet count, and fibrinogen concentration. RESULTS: According to the results of the present study, the reduced activity of von Willebrand factor in patients with AML was associated with severe bleeding. The authors observed an increase in the number of platelets count in patients with AML who experienced haemorrhages compared to patients with no bleeding signs. The study also established an increase in the concentration of fibrinogen in cancer patients, compared to the control sample. Symptoms and quantitative indicators for diagnosing the severity of haemorrhagic syndrome were grouped. The authors considered the advantages and disadvantages of many therapeutic preparations and focussed on specific markers of activated haemorrhage-predicting platelets. CONCLUSION: Further studies concern the search for effective markers and therapeutic approaches to minimize haemorrhagic syndrome. The results were statistically processed using the functions ANOVA, t test, CORREL, determination of the value of reliability, and mean square deviation.

Potential benefits of using chitosan and silk fibroin topical hydrogel for managing wound healing and coagulation.

Background & Objectives: The intricate process of wound healing involves replacing the cellular or tissue structure that has been destroyed. In recent years various wound dressings were launched but reported several limitations. The topical gel preparations are intended for certain skin wound conditions for local action. Chitosan-based hemostatic materials are the most effective in halting acute hemorrhage, and naturally occurring silk fibroin is widely utilized for tissue regeneration. So, this study was conducted to evaluate the potential of chitosan hydrogel(CHI-HYD) and chitosan silk fibroin hydrogel (CHI-SF-HYD) on blood clotting and wound healing. Methods: Hydrogel was prepared using various concentrations of silk fibroin with guar gum as a gelling agent. The optimized formulations were evaluated for visual appearance, Fourier transforms infrared spectroscopy (FT-IR), pH, spreadability, viscosity, antimicrobial activity, HR-TEM analysis, ex vivo skin permeation, skin irritation, stability studies, and in vivo studies by using adult male Wistar albino rats. Results: Based on the outcome of FT-IR, no chemical interaction between the components was noticed. The developed hydrogels exhibited a viscosity of 79.2 ± 4.2 Pa.s (CHI-HYD), 79.8 ± 3.8 Pa.s (CHI-SF-HYD), and pH of 5.87 ± 0.2 (CHI-HYD), 5.96 ± 0.1 (CHI-SF-HYD). The prepared hydrogels were sterile and non-irritant to the skin. The in vivo study outcomes show that the CHI-SF-HYD treated group has significantly shortened the span of tissue reformation than other groups. This demonstrated that the CHI-SF-HYD could consequently accelerate the regeneration of the damaged area. Interpretation & Conclusion: Overall, the positive outcomes revealed improved blood coagulation and re-epithelialization. This indicates that the CHI-SF-HYD could be used to develop novel wound-healing devices.

A kaolin/calcium incorporated shape memory and antimicrobial chitosan-dextran based cryogel as an efficient haemostatic dressing for uncontrolled hemorrhagic wounds.

Increased mortalities associated with uncontrolled and excessive bleeding is still of paramount concern in the clinics, caregivers and military medics. Herein, we designed a shape memory cryogel based on chitosan (C) and functionalized-dextran (D), incorporated with Kaolin (K) and calcium (Ca2+) as haemostatic agents. The developed cryogel (CDKCa) exhibits a uniform interconnected porous architecture with profound fluid absorption ability, rapid blood clotting, stable clot formation and good antibacterial activity. The CDKCa elucidates significantly less clotting time (~30 s; in-vitro) and increased aggregation and activation of platelets/red blood cells in comparison to the control groups and commercial dressings (Axiostat and QuikClot). The developed CDKCa also significantly reduced the aPTT and PT values by ~58 % and 31 % respectively, leading to the activation of intrinsic and extrinsic coagulation cascades. The CDKCa cryogel displays enhanced mechanical stability, flexibility and a good shape memory, a property quintessential to cease uncontrolled bleeding in irregular and non-compressible wounds. Further, the Kaolin and Ca2+ incorporated shape memory CDKCa cryogel demonstrates a rapid blood coagulation and stable clot formation in different compressible and non-compressible rat liver and femur hemorrhagic models. In summary, the endorsed results of CDKCa suggest that the design, fabrication and excellent clotting ability may attribute to high haemostatic efficiency of CDKCa dressing and have a great potential to prevent uncontrollable hemorrhages.

Fabrication of a Chitosan-Based Wound Dressing Patch for Enhanced Antimicrobial, Hemostatic, and Wound Healing Application.

Wounds are a serious life threat that occurs in daily life. The complex cascade of synchronized cellular and molecular phases in wound healing is impaired by different means, involving infection, neuropathic complexes, abnormal blood circulation, and cell proliferation at the wound region. Thus, to overcome these problems, a multifunctional wound dressing material is fabricated. In the current research work, we have fabricated a wound dressing polymeric patch, with poly(vinyl alcohol) (PVA) and chitosan (Cs) incorporated with a photocatalytic graphene nanocomposite (GO/TiO2(V-N)) and curcumin by a gel casting method, that focuses on multiple stages of the healing process. The morphology, swelling, degradation, moisture vapor transmission rate (MVTR), porosity, light-induced antibacterial activity, hemolysis, blood clotting, blood abortion, light-induced biocompatibility, migration assay, and drug release were analyzed for the polymeric patches under in vitro conditions. PVA/Cs/GO/TiO2(V-N)/Cur patches have shown enhanced wound healing in in vivo wound healing experiments on Wister rats. They show higher collagen deposition, thicker granulation tissue, and higher fibroblast density than conventional dressing. A histological study shows excellent re-epithelialization ability and dense collagen deposition. In vitro and in vivo analysis confirmed that PVA/Cs/GO/TiO2(V-N) and PVA/Cs/GO/TiO2(V-N)/Cur patches enhance the wound healing process.

Different modeling approaches in the simulation of extrinsic coagulation factor X activation: Limitations and areas of applicability.

Proteolytic reactions on the phospholipid membrane surface, so-called "membrane-dependent" reactions, play central role in the process of blood clotting. One particularly important example is FX activation by the extrinsic tenase (VIIa/TF). Here we constructed three mathematical models of FX activation by VIIa/TF: (A) a homogeneous "well-mixed" model, (B) a two-compartment "well-mixed" model, (C) a heterogeneous model with diffusion, to investigate the impact and importance of inclusion of each complexity level. All models provided good description of the reported experimental data and were equivalently applicable for <40 µM of phospholipids. Model C provided better predictions than A, B in the presence of TF-negative phospholipid microparticles. Models predicted that for high TF surface density (STF ) and FX deficiency the FX activation rate was limited by the rate of FX binding to the membrane. For low STF and excess of FX the reaction rate was limited by the tenase formation rate. The analysis of the substrate delivery pathways revealed that FX bound to VIIa/TF predominantly from solution for STF >2.8 × 10-3  nmol/cm2 and from the membrane for lower STF . We proposed the experimental setting to distinguish between the collision-limited and non-collision-limited binding. The analysis of models in flow and non-flow conditions revealed that the model of a vesicle in flow might be substituted by model C in the absence of the substrate depletion. Together, this study was the first which provided the direct comparison of more simple and more complex models. The reaction mechanisms were studied in a wide range of conditions.