Coagulation Activation Research Articles

Dielectric blood coagulometry to evaluate coagulation activity in patients prescribed factor Xa inhibitors undergoing elective surgery: A prospective observational study

Dielectric blood coagulometry to evaluate coagulation activity in patients prescribed factor Xa inhibitors undergoing elective surgery: A prospective observational study

Lipoprotein(a) and prothrombotic effects: Evidence from a genetic association study: Prothrombotic effects of lipoprotein(a).

It is unknown whether lipoprotein(a) [Lp(a)] has prothrombotic effects contributing to its association with the risk of myocardial infarction (MI). In 410,177 participants of UK Biobank, associations of LPA genetic variants and observed Lp(a) concentrations with the risk of venous thromboembolism (VTE) and MI were investigated, stratified by scores of genetic variants influencing coagulation through the thrombin and platelet pathways (denoted as F2/F5 and GUCY1A3 scores, respectively). Risk estimates are expressed as hazard ratio (HR) and 95% confidence interval (95% CI). Neither LPA genetic variants nor observed Lp(a) concentration were associated with the risk of incident VTE (HR per 100 nmol/L higher Lp(a) 1.02, 95% CI 1.00-1.04, p=0.13). In contrast, there was a strong association with the risk of incident MI (HR per 100 nmol/L higher Lp(a) 1.31, 95% CI 1.29-1.33, p<0.001). The F2/F5 score was associated with a stepwise decrease in the risk of VTE, and the GUCY1A3 score with a stepwise decrease in the risk of MI. However, the associations of LPA genetic variants and observed Lp(a) concentrations with the risk of MI were not modified by stratification for either of the coagulation scores. The association between Lp(a) and MI was not modified by genetically determined levels of coagulation activity through the thrombin or platelet pathway. Our findings do not support the notion that the increased risk of MI caused by elevated Lp(a) is due to prothrombotic effects.

Evolution of Coagulation and Platelet Activation Markers After Transcatheter Edge-to-Edge Mitral Valve Repair

Background/Objectives: The recommendations for antithrombotic therapy after transcatheter edge-to-edge mitral valve repair (TEER) are empirical, and the benefit of antiplatelet (APT) or anticoagulation therapy (ACT) remains undetermined. The study sought to investigate the degree and the timing of coagulation and platelet marker activation after TEER. Methods: This was a prospective study including 46 patients undergoing TEER. The markers of coagulation activation, namely prothrombin fragment 1 + 2 (F1 + 2) and thrombin-antithrombin III (TAT), and the markers of platelet activation, namely soluble P-Selectin and soluble CD-40 ligand (sCD40L), were measured at baseline, 24 h, 1 month, and 1 year after TEER. Results: At discharge, 20 (43%) patients received APT (single: 16, dual: 4), 24 (52%) received ACT, and 2 (4%) had both single APT and ACT. Levels of F1 + 2 and TAT significantly increased at 24 h post TEER (both p &lt; 0.001), rapidly returning to baseline levels at 1 month. However, levels of F1 + 2 and TAT remained higher at 1 month in patients without ACT compared to patients with ACT (respectively, 303.1 vs. 148.1 pmol/L; p &lt; 0.001 and 4.6 vs. 3.0 µg/L; p = 0.020), with a similar trend at 1 year. Levels of soluble P-selectin and sCD40L remained stable at all times after TEER (respectively, p = 0.071 and p = 0.056), regardless of the APT. Conclusions: TEER is associated with an acute activation of the coagulation system, with no increase in platelet activation markers. Hence, the use of dual APT is questionable in this population. Our results raise the hypothesis that the optimal antithrombotic therapy after TEER could be short-term ACT over APT. Further larger studies are warranted.

Factor XIII: Driving (Cross-)Links in Hemostasis, Thrombosis, and Disease.

Blood clots are complex structures composed of blood cells and proteins held together by the structural framework provided by an insoluble fibrin network. Factor (F)XIII is a protransglutaminase essential for stabilizing the fibrin network. Activated FXIII(a) introduces novel covalent crosslinks within and between fibrin and other plasma and cellular proteins, and thereby promotes fibrin biochemical and mechanical integrity. These irreversible modifications are also major determinants of clot composition and functional properties. As such, FXIII has central roles in hemostasis and wound healing, thrombosis, and many proinflammatory diseases associated with coagulation activation. FXIII's biochemical properties are as interesting as its biology is unusual, giving rise to unique and still undefined mechanisms. Here we review features underlying FXIII biology, biochemical function, biophysical impact, and (patho)physiologic implications in hemostasis, thrombosis, and disease.

A Review on Therapeutic Potential of Plumbago zeylanica Extracts and Sustainable Production Methods

Herbal medicines have earned a timeless place in healthcare, valued for their biocompatibility and widespread use in treating cancer and related conditions. Plumbago zeylanica, widely recognized as white leadwort or doctorbush, holds a prominent place in traditional medicine systems worldwide, revered for its diverse therapeutic properties. This widespread use, however, coupled with unsustainable harvesting practices, has raised con-cerns about the plant's long-term survival in its natural habitat. Their enduring presence is particularly evident in countries like India, Pakistan, Bangladesh, Sri Lanka, and Australia, where they are deeply rooted in ethnomedical traditions. P. zeylanica stands out as a rich source of bioactive compounds, including naphthoquinones, flavonoids, alkaloids, and ter-penoids, with plumbagin identified as a key driver of its medicinal properties. Plumbagin exhibits a remarkable pharmacological profile, demonstrating significant efficacy as an an-ticancer, antidiabetic, antimalarial, and antimicrobial agent, making it a promising candidate for drug development. The literature reveals that the root and root bark also have a wider claim in traditional medicines against various diseases as a memory enhancer, anti-inflam-matory, wound healing, anti-infertility, blood coagulation, and anti-oxidant activities. This review provides a comprehensive overview of P. zeylanica, encompassing its traditional uses, phytochemical composition, and the pharmacological activities of its key bioactive constituents, with a particular focus on plumbagin. We delve into the scientific evidence supporting its therapeutic applications, highlighting its potential in treating a wide range of diseases. Furthermore, we discuss the progress made in developing sustainable production methods for plumbagin for medicinal purposes from P. zeylanica.

Endothelium Modulates the Prothrombotic Phenotype of Factor V Leiden: Evidence From an Ex Vivo Model.

Clinical expressivity of the thrombophilic factor V Leiden (FVL) mutation is highly variable. Recently, we demonstrated an increased APC (activated protein C) response in asymptomatic FVL carriers compared with FVL carriers with a history of venous thromboembolism (VTE) after in vivo coagulation activation. Here, we further explored this association using a recently developed ex vivo model based on patient-specific endothelial colony-forming cells (ECFCs). ECFCs and citrated plasma were obtained from FVL carriers with previous VTE (VTE+, n=9), FVL carriers without previous VTE (VTE-, n=8), and 7 healthy controls. Coagulation was activated by TF (tissue factor) in defibrinated recalcified plasma added to confluent cell cultures. Thrombin and APC concentration were measured over time, and the respective areas under the curve were calculated. Additionally, inhibition kinetics of exogenously added APC, APC inhibitor levels, and APC sensitivity ratios were measured in plasma. Expression of TM (thrombomodulin) and EPCR (endothelial protein C receptor) on ECFCs was assessed using cell-based ELISAs. In autologous plasma on ECFCs, the APC response (area under the curve APC/area under the curve thrombin) was higher in FVL VTE- than in FVL VTE+ patients (0.138 versus 0.028; P=0.026). APC inhibitor levels, APC inactivation kinetics, and APC sensitivity ratios did not differ between cohorts. Crossover experiments, which combined pooled plasma from FVL VTE- patients with FVL VTE+ ECFCs in the ex vivo model, and vice versa, showed increased APC response rates when FVL VTE- ECFCs were used, regardless of the plasma component. In cell-based ELISAs, TM and EPCR expression showed no significant difference. Although the FVL gene product induces an almost identical APC resistance phenotype in plasma, the endothelial cell-dependent APC response rates differ significantly, with a higher APC response in asymptomatic FVL carriers. Further studies are warranted to elucidate the disease-modulating role of the endothelium in FVL carriers at the molecular level.

Analysis of the sensitization activity of Moringa oleifera leaves protein.

The determination of allergenic proteins in Moringa oleifera leaves, which is the main components of immune activity, has enabled the development of a more effective method for evaluating the activity of extracted Moringa oleifera leaves protein. In this study, the extraction process of Moringa oleifera leaves protein was optimized based on a single factor experiment. The hemagglutination-related properties of Moringa oleifera leaves protein, such as (thermal, acid-base) stability, sugar binding specificity, ion binding characteristics, and hemolytic activity, were detected. The optimal combination of extraction process was: extraction time of 6 h, material-liquid ratio of 1:8, and ammonium sulfate saturation of 60%. The extraction rate of moringa leaf protein under this condition was 14.37 mg/g. The molecular weight of moringa leaf protein was analyzed by SDS-PAGE, and the molecular weight was mainly concentrated around 23 kDa~70 kDa, with the highest content of 35 kDa (major allergen). The study of the hemagglutination characteristics of Moringa oleifera leaves protein revealed that the protein exhibited high stability at temperatures below 60°C, with complete loss of activity occurring at temperatures above 110°C for 20 min. The effect of different pH conditions on the hemagglutination capacity of Moringa oleifera leaves protein was readily discernible. The hemagglutination activity of Moringa oleifera leaves protein was 104 in a pH value from 3.7 to 7.8, and the hemagglutination activity was completely lost at a pH value higher than 11.9. D(+) anhydrous glucose is the specific inhibitory sugar of Moringa oleifera leaves protein lectin. Moringa oleifera leaves protein exhibits hemolytic activity at a concentration of at least 20 mg/mL, and α-methyl-mannoside, galactoside, raffinose and Al3+ can inhibit the hemolysis of Moringa oleifera leaves protein. The present study identified the effects of different factors on the coagulation activity and hemolytic ability of Moringa oleifera leaves protein, thereby providing a theoretical basis for further purification and application of Moringa oleifera lectin. However, it should be noted that the results of the mixture have certain limitations, and further purification of lectin is needed to obtain more targeted research results.

DNA-loaded targeted nanoparticles as a safe platform to produce exogenous proteins in tumor B cells.

The functionalization of nanoparticles (NPs) with an antiCD19 targeting mechanism represents a promising approach for the selective delivery of drugs and nucleic acids into normal and tumor B cells. This strategy has the advantage of minimizing off-target effects by restricting gene delivery to the desired cell population. However, the nanoplatform must guarantee both the local production of the protein and the safety of the treatment to allow an effective therapy with reduced systemic toxicity. In order to ensure a selective delivery of nucleic acids, we developed poly(lactic-co-glycolic acid) (PLGA)-poly(vinyl alcohol) (PVA) NPs loaded with an Enhanced Green Fluorescent Protein (EGFP)-coding plasmid and covalently coated with antiCD19 recombinant antibody as a targeting mechanism. To assess the functionality of the NPs, physicochemical characterization, safety tests, and transfection assay were employed to evaluate the NPs' behavior in vitro and in vivo, in a human/zebrafish lymphoma xenograft model. The results demonstrated that the PLGA-PVA nanoplatform was capable of efficiently encapsulating and releasing the payload. These nanostructures demonstrated a favorable safety profile, as evidenced by the absence of significant cell cytotoxicity, coagulation activation, complement system activation, and the slight activation of endothelial cells and leukocytes. The targeting mechanism facilitated the interaction of NPs with target cells, thereby enhancing their internalization and subsequent exogenous plasmid DNA (pDNA) translation and protein expression. In the human/zebrafish lymphoma xenograft model, no evidence of toxicity was observed, and targeted NPs demonstrated the capacity to enhance exogenous pDNA expression. Our findings provide a rationale for the use of targeted NPs as a DNA delivery system for the local expression of therapeutic proteins.

Efficient production of recombinant human FVII in CHO cells using the piggyBac transposon system.

As an important coagulation factor, activated coagulation factor VII (FVIIa) is mainly used to treat the bleeding of hemophilia patients who have developed inhibitory antibodies against FVIII and FIX conventional treatment. Recombinant human factor VII (rhFVII) produced in mammalian cell lines have been developed as the most important resource of FVIIa. However, cell lines express rhFVII protein derived from an exogenous expression vector at a lower level than most other proteins. In the current study, we have shown efficient rhFVII production in CHO cell lines using piggyBac (PB) transposon system. rhFVII is successfully expressed in fed-batch culture of CHO cells, and the expression of rhFVII up to 100 mg/L. Moreover, the purified secreted rhFVII was determined by SDS-PAGE and Western Blot. The coagulation activity was determined by the chromogenic Activity ELISA kit. In conclusion, this study has demonstrated that the piggyBac transposon system can be used for an efficient production of recombinant FVII.

Real-Time Imaging of Platelet-Initiated Plasma Clot Formation and Lysis Unveils Distinct Impacts of Anticoagulants.

Fibrinolysis is spatiotemporally well-regulated and greatly influenced by activated platelets and coagulation activity. Our previous real-time imaging analyses revealed that clotting commences on activated platelet surfaces, resulting in uneven-density fibrin structures, and that fibrinolysis initiates in dense fibrin regions and extends to the periphery. Despite the widespread clinical use of direct oral anticoagulants (DOACs), their impact on thrombin-dependent activation of thrombin-activatable fibrinolysis inhibitor (TAFI) and fibrinolysis remains unclear. Here, we investigated the effects of different DOACs on the TAFI-mediated inhibition of fibrinolysis. Using human platelet-containing plasma, we performed turbidimetric assays, thrombin generation assays, and confocal laser scanning microscopy to assess the effects of anticoagulants on fibrinolysis. Activated platelets-prolonged plasma clot lysis time, shortened by activated TAFI inhibitor (TAFIaI), positively correlated with the amount of thrombin generated. Rivaroxaban (an activated factor X inhibitor) and dabigatran (a direct thrombin inhibitor) dose-dependently shortened lysis time comparably. The highest concentration of DOACs showed no further shortening of lysis time with TAFIaI. The fibrin network structures initiated by activated platelets and the localization of fluorescently labeled plasminogen were unique for these two drugs. Rivaroxaban maintained an uneven fibrin network but promoted faster plasminogen accumulation and fibrinolysis from outside dense fibrin regions. Conversely, dabigatran resulted in a more even fibrin network, with fibrinolysis starting from the activated platelets and propagating to the periphery. Visualizing and analyzing the patterns of fibrin network formation, plasminogen accumulation, and fibrinolysis provide new insights into the specific impact of anticoagulants on coagulation and fibrinolysis.

Proteases from an Amazonian Mushroom Species: A Mycotechnological Alternative for the Production of Milk Coagulant.

Edible mushrooms have been used as sustainable sources of proteases of industrial interest. The aim of this research was to investigate the influence of different culture media on mycelial growth and the potential of an Amazonian mushroom species, Auricularia fuscosuccinea DPUA 1624, in the biosynthesis of bovine milk coagulant enzymes. The species was cultivated on Sabouraud agar, malt, glucose, and peptone agar, malt extract agar, and glucose and peptone agar, supplemented with yeast extract for mycelial development. Enzyme biosynthesis was evaluated by submerged fermentation. Subsequently, the cultures were incubated at 28°C for 8days. Proteolytic and coagulant activities were determined using 1% azocasein solution and milk powder as substrates, respectively. In the results of radial growth speed of A. fuscosuccinea, the values were significant in the GYP and SAB + YE culture media. However, GYP agar favored the growth and mycelial vigor of A. fuscosuccinea; therefore, this medium was selected to obtain inoculum in the tests. In submerged fermentation, the MGYP medium favored the synthesis of proteases for A. fuscosuccinea and synthesized coagulant proteases in 100% of the media, in which significant activity was observed in SAB + YE. The significant production of coagulant proteases of A. fuscosuccinea was obtained under the following conditions: inoculum size 10%, 8days of fermentation period, and 8days of inoculum age. The results indicate that A. fuscosuccinea DPUA 1624 has potential for use in industrial manufacturing, especially in dairy products.

Comparative Analysis of the Enzymatic, Coagulant, and Neuromuscular Activities of Two Variants of Crotalus durissus ruruima Venom and Antivenom Efficacy.

Background: We compared the enzymatic, coagulant, and neuromuscular activities of two variants (yellow-CDRy and white-CDRw) of Crotalus durissus ruruima venom with a sample of C. d. terrificus (CDT) venom and examined their neutralization by antivenom against CDT venom. Methods: The venoms were screened for enzymatic and coagulant activities using standard assays, and electrophoretic profiles were compared by SDS-PAGE. Neutralization was assessed by preincubating venoms with crotalic antivenom and assaying the residual activity. Results: SDS-PAGE showed that the venoms had similar electrophoretic profiles, with the main bands being phospholipase A2 (PLA2), serine proteinases, L-amino acid oxidase (LAAO), and phosphodiesterase. CDRy venom had the highest proteolytic and LAAO activities, CDRw venom had greater PLA2 and esterolytic activities at the highest quantity tested, and CDT had greater PLA2 activity than CDRy. CDRw and CDT venoms had similar proteolytic and LAAO activities, and CDRy and CDT venoms had comparable esterolytic activity. None of the venoms altered the prothrombin time (PT), but all of them decreased the activated partial thromboplastin time (aPPT); this activity was neutralized by antivenom. The minimum coagulant dose potency was CDRw >> CDRy > CDT. All venoms had thrombin-like activity that was attenuated by antivenom. CDRy and CDRw venoms showed α-fibrinogenolytic activity. All venoms partially cleaved the β-chain. CDRy and CDT venoms caused neuromuscular facilitation (enhanced muscle contractions) followed by complete blockade, whereas CDRw venom caused only blockade. Antivenom neutralized the neuromuscular activity to varying degrees. Conclusions: These findings indicate that while CDR and CDT venoms share similarities, they also differ in some enzymatic and biological activities and in neutralization by antivenom. Some of these differences could influence the clinical manifestations of envenomation by C. d. ruruima and their neutralization by the currently used therapeutic antivenom.

Utility of unfractionated sodium heparin in the prevention of disseminated intravascular coagulation

Introduction: disseminated intravascular coagulation (DIC) is a serious disease characterized by a generalized activation of blood coagulation, with sequelae ranging from thrombus formation to severe bleeding.Objective: to analyze the fundamental alternatives for the management of DIC prevention related to the use of unfractionated heparin.Method: data collection was carried out through a search in the digital databases PUBMED, SCIELO (Scientific Electronic Library Online) and GOOGLE ACADEMIC. The articles were downloaded from the aforementioned platforms, and the Zotero system was used to make the corresponding citations and pertinent references.Results: according to the results achieved, the systematic review allowed to obtain a more detailed view of the studies arising from the data search, related to DIC and the use of unfractionated heparin in its prevention. The importance of research related to this drug was highlighted to improve the understanding and treatment of DIC with the aim of reducing its morbidity and mortality.Conclusions: despite unfractionated heparin presenting significant anticoagulant effects in its clinical utility, mainly in emergency situations due to its rapid action, there are few studies available for a better understanding of the management and functionality of UFH against DIC

Mortality-associated plasma proteome dynamics in a prospective multicentre sepsis cohort.

Sepsis remains a leading cause of mortality in intensive care units. Understanding the dynamics of the plasma proteome of patients with sepsis is critical for improving prognostic and therapeutic strategies. This prospective, multicentre observational cohort study included 363 patients with sepsis recruited from five university hospitals in Germany between March 2018 and April 2023. Plasma samples were collected on days 1 and 4 after sepsis diagnosis, and proteome analysis was performed using mass spectrometry. Classical statistical methods and machine learning (random forest) were employed to identify proteins associated with 30-day survival outcomes. Out of 363 patients, 224 (62%) survived, and 139 (38%) did not survive the 30-day period. Proteomic analysis revealed significant differences in 87 proteins on day 1 and 95 proteins on day 4 between survivors and non-survivors. Additionally, 63 proteins were differentially regulated between day 1 and day 4 in the two groups. The identified protein networks were primarily related to blood coagulation, immune response, and complement activation. The random forest classifier achieved an area under the receiver operating characteristic curve of 0.75 for predicting 30-day survival. The results were compared and partially validated with an external sepsis cohort. This study describes temporal changes in the plasma proteome associated with mortality in sepsis. These findings offer new insights into sepsis pathophysiology, emphasizing the innate immune system as an underexplored network, and may inform the development of targeted therapeutic strategies. European Regional Development Fund of the European Union. The State of North Rhine-Westphalia, Germany.

Pathogenesis and management of sepsis patients with disseminated intravascular coagulation

Sepsis is a life-threatening organ dysfunction caused by dysregulation of the body's response to infection, and is one of the leading causes of death in intensive care units. In sepsis, the systemic inflammatory reaction that occurs activates the coagulation system and increases the consumption of clotting factors, leading to a serious complication of coagulopathy known as disseminated intravascular coagulation (DIC). DIC is a clinicopathological syndrome that is a complication of various diseases characterized by the activation of systemic blood coagulation and intravascular fibrin production, which can cause thrombosis of small- and medium-sized blood vessels and can even cause organ function failure along with the consumption of platelets and coagulation factors, resulting in a clinical picture of bleeding. In 35% of severe sepsis cases, and in shock sepsis DIC causes high mortality, which is associated with increased severity of illness and serious organ failure with poor outcomes. This article aims to summarize the evolving understanding of DIC pathogenesis in patients with sepsis and its implications for current and future management strategies.

IMPROVING THE TECHNOLOGY OF ENRICHED SOFT CHEESE BASED ON THE STUDY OF THE SUITABILITY OF COW'S MILK FOR CHEESE PRODUCTION IN PAVLODAR REGION

This article reflects the results of the conducted research on the cheese suitability of milk from cows of different breeds of the Pavlodar region and the development and improvement of production technology for a new type of enriched soft cheese for functional purposes. At the first stage of the research, a review of special and scientific, technical and patent literature was conducted on the trend in the development of technology for the production of natural rennet soft cheese using raw materials of vegetable origin. Further, in the course of the research work, experimental studies were conducted on the suitability of milk from 3 breeds of cows of the Pavlodar region «Simmental», «Red Steppe» and «Kazakh white-headed» for rennet coagulation. According to the results of the conducted research, cow's milk of the Simmental breed was selected as the main raw material for the production of enriched soft cheese. Then several variants of a mixture of good-quality milk of cows «Simmental» and less good-quality milk of cows of the breed «Krasnaya Stepnaya» were compiled in various ratios (90/10, 80/20, 70/30, 60/40, 50/50) and accordingly, the process of rennet coagulation of selected variants of raw milk using selected starter cultures and rennet was investigated enzymes. According to the results obtained, it was also found that the enzyme «Natural Rennet» and the starter culture «Lactoferm ECO» also have a very high coagulation activity of various milk mixtures. The dynamics of changes in titrated acidity during coagulation of a mixture (60-40 and 50-50) of raw milk from cows of the Simmental breed and less raw milk from cows of the Krasnaya Stepnaya breed reached the maximum index for titrated acidity Further, studies were conducted on the selection and justification of additives of plant origin – as functional components in the formulation of a new type of enriched soft cheese for functional nutrition. According to the results of the research, a milk – vegetable composition (milk – protein clot and vegetable paste) was developed as a functional additive for the production of enriched soft cheese and its concentration in the component composition of enriched soft cheese was determined. As a result, samples of soft cheese with chickpea paste in the amount of 2-4% were selected. It was found that with an increase in the dose of vegetable filler of 6-10% leads to a deterioration in organoleptic parameters, i.e. an overly pronounced taste of the bean component.

Рекомбинантный химозин Camelus dromedarius в экспрессионной системе Pichia pastoris: очистка и анализ ферментативных свойств

Natural chymosin production is an expensive and complex process associated with ethical issues. The article introduces recombinant chymosin Camelus dromedarius (rChn-Cam) isolated from a P. pastoris expression system and optimized for different nutrient media at different zeocin concentrations. The sequence of prochymosin gene was obtained from NCBI BLAST. GS115/his4 P. prastoris served as a producer strain. The pPICZ(alpha) B vector with the AOXI promoter made it possible to construct the expression cassette. The experiment involved methods of genetical engineering and strain cultivation. The recombinant His-Tag-labelled proteins were isolated by the method of metal-affinity chromatography and analyzed using PAG electrophoresis and Western-blot analysis. The molecular weight was determined by MALDI-TOF MS while the concentration was defined spectrophotometrically. The shuttle expression plasmid pPICZ(alpha)B/proCYM_camel_pp_IDT revealed that the cell mass expansion of P. pastoris GS115/his4 (Mut+) should be performed with a preliminary introduction of 0.5% methanol. After the transformation of P. pastoris GS115/his4 and obtaining a strain-producer of P. pastoris/pPICZ(alpha)B/proCYM_camel_pp_IDT, the rate of cell mass gain started to correlate with the zeocin concentrations in two different media. Medium YPD was not fornified and contained 50, 100, and 200 μg/mL zeocin. MediumYPD was fortified with 0.00004% biotin and 1% glycerol and included 50, 100, and 200 μg/mL zeocin. The strain-producer grew better in the medium with a zeocin concentration of 50 μg/mL. The mass of rChn-Cam was 35.673 kDa after isolation and purification. When the pH of the substrate rose from 5.0 to 6.5, the coagulation activity decreased by 24%. The thermal inactivation threshold of rChn-Cam was 40–45°C. The unit of coagulation activity decreased as the zeocin concentration went up. The rChn-Cam concentration was in inverse correlation with the substrate coagulation time. In this research, the rChn-Cam obtained in the expression system of P. prastoris proved to be a good alternative to rChn used in the cheese industry.

Platelet spleen tyrosine kinase is a key regulator of anti-PF4 antibody-induced immunothrombosis.

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a rare but serious prothrombotic adverse event following vaccination with adenovector-based COVID-19 vaccines. Laboratory findings indicate that anti-platelet factor 4 (PF4) immunoglobulin G antibodies are the causing factor for the onset of thromboembolic events in VITT. However, molecular mechanisms of cellular interactions, signaling pathways and involvement of different cell types in VITT antibody-mediated thrombosis are not fully understood. Moreover, uncertainty exists regarding current treatment protocols since the sole anticoagulation was shown to be inefficient to prevent thrombosis progression in severe VITT cases. In this study, we demonstrate that platelet spleen tyrosine kinase (SYK) modulates anti-PF4 VITT-mediated thrombus formation in an ex vivo model of immunothrombosis. Our study showed that the selective inhibition of SYK can abrogate VITT antibody-driven procoagulant platelet formation, activation of plasmatic coagulation as well as platelet-leukocyte interplay. Most importantly, the specific inhibition of SYK in platelets but not in neutrophils prevented VITT antibody-induced multicellular thrombus formation, without perturbing the platelet function. Our findings indicate that the specific targeting of platelet SYK might be a promising therapeutic approach to prevent thrombotic complications in patients with antibody-mediated immunothrombosis.

Effect of Blood Sample Tubes on Diagnosis of Gestational Diabetes in Pregnant Women Undergoing OGTT

Performing a diagnostic oral glucose tolerance test (OGTT) during pregnancy is important for early diagnosis of gestational diabetes mellitus. Hyperglycemia can be detected quickly, and mother and the fetus can be protected. This study aimed to research the relationship between blood sample tubes and the stability of glucose in pregnant women undergoing OGTT. Blood samples were taken at the 120th minute after glucose loading from 20 pregnant volunteers who underwent OGTT (75g). VACUETTE CAT Serum Separator (Clot Activator Tube) was used for serum; VACUETTE FC Mix tube (Na2 EDTA, sodium fluoride, sodium citrate) and VACUETTE FE (Sodium Fluoride/K3 EDTA) were used for plasma. All three of these tubes were centrifuged at three different times i.e. 0. hour, 2. hour, and 4. hour. All samples were centrifuged at 0., 2., and 4. hours, and glucose values were measured. No statistically meaningful difference was found between their glucose values in only VACUETTE FC Mix tubes (p&gt;0.05). However, a statistically meaningful difference was found at these different time intervals in serum and fluoride tubes (p

Inflammation and hemostasis in loiasis before and after ivermectin treatment: A biological pilot cross-sectional study.

We assessed the impact of loiasis and its treatment with ivermectin on hemostasis and inflammation in 38 adults in Cameroon. Participants were divided into four balanced groups based on their Loa loa microfilarial densities. At baseline, eosinophils and platelets increased with microfilarial densities (p<0.001 and p = 0.044, respectively). At day 4 following ivermectin administration, median eosinophils rose from 171/μL to 1,095/μL (p<0.001) and median D-dimers increased from 582 ng/mL to 745 ng/mL (p = 0.024) with a median relative variation of +34.7%. C-reactive protein, fibrinogen, and alpha-1-globulin also increased significantly after treatment. Ivermectin treatment appeared to induce inflammation, coagulation activation and fibrinolysis.