Fibrinolytic Process Research Articles

IP225. Real-World Outcomes of EKOS Ultrasound-Enhanced Catheter-Directed Thrombolysis for Acute Limb Ischemia

Ultrasound-enhanced catheter-directed thrombolysis (UET) using the EKOS device (EKOS Corporation, Bothell, Wash) for acute peripheral artery ischemia has been purported in clinical trials to accelerate the fibrinolytic process to reduce treatment time and lytic dosage. However, UET performance in a real-world clinical setting has thus far been inadequately described. We performed a retrospective review of all patients undergoing UET for acute limb ischemia at a single institution during a 5-year period. Data collected included patients' demographics, procedural details, and 30-day and 1-year outcomes. Primary outcomes were major bleeding events, limb salvage, and mortality. We identified 32 patients (mean age, 67.4 ± 14.9 years; 25% female) who underwent UET for acute limb ischemia between 2014 and 2018. At presentation, 43.8% were therapeutically anticoagulated. Rutherford acute limb ischemia classification was I in 56.3%, IIa in 25.0%, and IIb in 18.8%. Etiology was thrombosis of native artery in 12.5% of patients, infrainguinal prosthetic bypass in 31.3%, infrainguinal autogenous bypass in 6.3%, and stented native vessel in 50.0%. Mean duration of thrombolytic therapy was 22.2 ± 11.3 hours, and mean tissue plasminogen activator dose was 24.5 ± 15.3 mg. In-line flow to the foot was re-established in 90.6% of patients with a significant improvement in preoperative to postoperative ankle-brachial index (0.31 ± 0.29 vs 0.78 ± 0.34; P < .001) and number of patent tibial runoff vessels (1.31 ± 1.20 vs 1.96 ± 0.86; P < .001; Fig). There was no significant difference in revascularization success between occluded vessel types: native vessel, 75% success; stented vessel, 93.8%, autogenous bypass, 100%, and prosthetic bypass, 90% (P = .48). Major bleeding complications occurred in three patients (9.4%), including two intracranial hemorrhages (6.3%). Limb salvage at 30 days and 1 year was 93.8% and 87.5%, respectively. Ipsilateral reintervention was required in 37.5% of patients within 1 year. Overall mortality was 6.2% at 30 days and 13.5% at 1 year, and mortality was significantly higher in patients with higher Rutherford class (P = .049). UET with the EKOS device demonstrates acceptable real-world outcomes in the treatment of acute limb ischemia. UET is generally safe, with a low rate of associated hemorrhagic complications, and is effective at re-establishing in-line flow to yield high limb salvage rates. However, UET is associated with a high rate of reintervention within 1 year. Further investigation is needed into specific predictors of limb salvage and need for reintervention as well as cost-efficacy of this technology compared with traditional methods.

Use of Viscoelastography in Malignancy-Associated Coagulopathy and Thrombosis: A Review.

The relationship between malignancy and coagulopathy is one that is well documented yet incompletely understood. Clinicians have attempted to quantify the hypercoagulable state produced in various malignancies using common coagulation tests such as prothrombin time, activated partial thromboplastin time, and platelet count; however, due to these tests' focus on individual aspects of coagulation during one specific time point, they have failed to provide clinicians the complete picture of malignancy-associated coagulopathy (MAC). Viscoelastic tests (VETs), such as thromboelastography (TEG) and rotational thromboelastometry (ROTEM), are whole blood analyses that have the advantage of providing information related to the cumulative effects of plasma clotting factors, platelets, leukocytes, and red cells during all stages of the coagulation and fibrinolytic processes. VETs have gained popularity in the care of trauma patients to objectively measure trauma-induced coagulopathy (TIC), but the utility of VETs remains yet unrealized in many other medical specialties. The authors discuss the similarities and differences between TIC and MAC, and propose a mechanism for the hypercoagulable state of MAC that revolves around the thrombomodulin-thrombin complex as it switches between activating the protein C anticoagulation pathway or the thrombin activatable fibrinolysis inhibitor coagulation pathway. Additionally, they review the current literature on the use of TEG and ROTEM in patients with various malignancies. Although limited research is currently available, early results demonstrate the utility of both TEG and ROTEM in the prediction of hypercoagulable states and thromboembolic complications in oncologic patients.

Hexachloronaphthalene as a hemostasis disturbing factor in female Wistar rats – A pilot study

Although Persistent Organic Pollutants (POPs) are some of the most dangerous environmental toxicants, data on their impact on hemostasis are virtually limited. 1,2,3,5,6,7-hexachloronaphthalene (PCN67) seems to be one of the most toxic congeners of polychlorinated naphthalenes (PCNs), which have recently been listed as POPs. The toxic effects of PCNs are similar to other chlorinated aromatics, e.g. polychlorinated dibenzo-p-dioxins (PCDDs), so an impact on hemostasis could not be excluded. Therefore, this study examines, for the first time, if short-term (two and four weeks) exposure of a mixture of hexachloronaphthalene congeners with a PCN67 as a predominant component to female Wistar rats may have an impact on selected hemostasis parameters, such as overall potential and kinetic parameters of clot formation and fibrinolysis; hematology and basic coagulology parameters. It also examines the influence of PCN67 on the stability of erythrocyte membranes. Obtained results indicate that PCN67 may be an important disturbing factor regarding both coagulation and fibrinolysis processes, as well as platelet count. Exposure to PCN67 significantly affected clot formation and lysis processes and diminished fibrinogen concentration after both administration periods. After two weeks of administration, an increased activated partial thromboplastin time (APTT) was noted; after four weeks - decreased platelet count with concomitant increased in mean platelet volume. Moreover, PCN67 may exert adverse effects on the red blood cells membrane stability, which were manifested by a statistically significant increase of red blood cells lysis.

Effect of Lipid-lowering Therapy and Regular Exercise on the Fibrinolytic System in Patients with Metabolic Syndrome

At present, it has been established that in patients with a combination of the main components of the metabolic syndrome (arterial hypertension, dyslipidemia, abdominal obesity, insulin resistance), violations occur in many vital systems, including the hemostasis system. The effect of shortterm (8-week) course of combination of lipid-lowering therapy and dose-related physical loads on the parameters of the fibrinolysis system in patients with metabolic syndrome was studied. It was shown that treatment with a combination of fibrates with physical loads is accompanied by a decrease in the level of fibrinogen and acceleration of fibrinolysis. The use of statins as hypolipidemic agents was accompanied by a reduction in the level of fibrinogen compared with fibrates and activation of fibrinolysis. Obviously, exercise exacerbates the mechanisms of action of hypolipidemic drugs on fibrinogen synthesis and fibrinolysis processes, which causes a favorable effect of their combined use.

Лейцинсодержащие пептиды как перспективные антитромботические вещества

Here we study the gemostatic properties of leucine-containing regulatory oligopeptides Н-Pro-Leu-Pro-OH (PLP) and H-Pro-Leu-Pro-Ala-OH (PLPA) administered orally every 24 hours for 7 days to healthy animals (rats). Standard coagulologic techniques were applied. We demonstrated that 20 hours after the 7th administration of the peptides the anticoagulant activity in the blood plasma was increased by 10.3-30.6% according to the activated partial thromboplastin time data, all types of the fibrinolytic process — total plasma fibrinolytic activity by 16-24%, non-enzymatic fibrinolysis – by 9-11%, enzymatic fibrinolysis – by 28-49%, and the plasmin activity – by 29%. In contrast, the lysis time of the euglobulin fibrin clot (in absence of fibrinolysis inhibitors) was not affected. The activity of tissue plasminogen activator was either unchanged (PLP) or decreased under the effect of PLPA. The fibrinogen concentration either decreased by 18% (PLP), or remained at normal values (PLPA); the activity of factor XIIIa decreased by 17% (PLP) and 10% (PLPA). Both peptides suppressed the platelet aggregation by 31-33.5% (when used as an ADP inductor). The most prominent anticoagulant effect was observerd for the oligopeptide containing the additional alanine residue at the C-terminus. Since both peptided showed varying degrees of complex antiplatelet, anticoagulant, fibrindepolymerization, fibrinolytic activity, and moderate reduction of coagulation factor levels — fibrinogen and factor XIIIa — when administered orally to animals, they can be considered promising antithrombotic agents.

Состояние гемокоагуляции у больных системной красной волчанкой на фоне пульс-терапии

Введение. Ревматические заболевания относятся к процессам, характеризующимся иммунным поражением различных органов и систем с выраженными изменениями сосудистой стенки и системы гемостаза. Цель исследования: изучить влияние высоких доз глюкокортикостероидов (ГКС) и цитостатиков на клинико-гемостазиологические показатели у больных системной красной волчанкой (СКВ). Материалы и методы. Обследовано 18 больных СКВ женщин (средний возраст — 35,6 ± 1,6 года), которые получали пульс-терапию ГКС и цитостатиком циклофосфаном по стандартной схеме. Критерии отбора для проведения пульс-терапии: высокая активность аутоиммунного процесса по шкалам SLAM, SLEDAI, ECLAM, поражение почек и центральной нервной системы. Гемостазиологические показатели исследовали до лечения, на 1-й и 14-й день после пульс-терапии. Для оценки состояния коагуляционного звена гемостаза определяли: активированное частичное тромбопластиновое время, тромбиновое время, протромбиновое время по Квику, содержание фибриногена, растворимых фибрин-мономерных комплексов, Д-димера, активность антитромбина III (АТ-III), протеина С, фибринолитической системы крови. Результаты. У больных СКВ наблюдали активацию коагуляционного гемостаза на фоне снижения активности естественных антикоагулянтов и процессов фибринолиза. На 1-е сутки после пульс-терапии выявлен значительный рост тромбинемии; к 14-у дню большинство показателей плазменного гемостаза возвращались к исходным на фоне снижения активности системы протеина С и роста активности АТ-III. Заключение. Исследование коагулологических параметров следует проводить всем больным СКВ, получающим пульс-терапию, так как существует угроза развития тромбозов у данных пациентов. Полученные данные подтверждают необходимость применения антикоагулянтов при назначении высоких доз ГКС и цитостатиков. Introduction. Rheumatic diseases are characterized by the immune damage of various organs and systems with expressed changes in the vascular wall and hemostasis. Aim: to study the eff ect of high doses of glucocorticosteroids (GCS) and cytostatics on the clinical and hemostasiological parameters in patients with systemic lupus erythematosus (SLE). Materials and methods. We examined 18 women with SLE of (average age — 35.6 ± 1.6 years) who received pulse therapy by GCS and cyclophosphamide according to the standard scheme. Selection criteria for pulse therapy were high activity of the autoimmune process according to SLAM, SLEDAI, ECLAM scales, kidney and central nervous system damage. Hemostasiological parameters were investigated before treatment, on the 1st and 14th days after pulse therapy. To assess the state of coagulation hemostasis, we determined: activated partial thromboplastin time, thrombin time, Quick prothrombin time, fibrinogen content, soluble fi brin-monomer complexes, D-dimer, activity of antithrombin III (AT-III), protein C, fibrinolytic system. Results. In patients with SLE, activation of coagulation hemostasis was observed with the reduction in the activity of natural anticoagulants and fi brinolysis processes. On the 1st day after pulse therapy, a signifi cant thrombinemia growth was detected; by the 14th day, most of plasma hemostatic parameters returned to baseline with reduction of protein C system activity and increasing of AT-III activity. Conclusion. The study of coagulological parameters should be carried out in all patients with SLE who receive pulse therapy, since there is a risk of thrombosis in these patients. The obtained data confirm the need for anticoagulants use in the appointment of high doses of GCS and cytostatics.

Homocysteine-Enhanced Proteolytic and Fibrinolytic Processes in Thin Intraluminal Thrombus and Adjacent Wall of Abdominal Aortic Aneurysm: Study In Vitro.

Homocysteine (Hcy) may affect the pathogenesis of abdominal aortic aneurysms (AAAs) through enhancement of proteolysis and an impaired coagulation/fibrinolysis system. Intensified haemostatic capacity may promote local proteolytic degradation of the aortic wall. This study aimed to examine the effects of Hcy on haemostatic and proteolytic processes in samples of thick and thin fragments of the ILT and underlying walls. Subjects and Methods. Thirty-six patients who underwent AAA surgery were enrolled. Aneurysm tissue sections were incubated with DL-Hcy (100 and 500 μmol/L) in a series of experiments and analyzed for concentration/activity of proteolytic and haemostatic markers by enzyme-linked immunosorbent assay. Results. Incubation of wall underlying thin ILT segments (B) with DL-Hcy resulted in an increase of active MMP-2 levels compared to control tissue (9.54 ± 5.88 versus 7.44 ± 4.48, p=0.011). DL-Hcy also induced t-PA and plasminogen concentration increases in thin thrombus sections (B1) compared to control tissue (respectively: 1.39 ± 1.65 versus 0.84 ± 0.74, p=0.024; 11.64 ± 5.05 versus 10.34 ± 5.52, p=0.018). In contrast, wall adjacent to thick thrombus segments (A) showed decreases in MMP-2 and TF activities compared to control (respectively, 5.89 ± 3.39 versus 7.26 ± 5.49, p=0.046; 67.13 ± 72.59 versus 114.46 ± 106.29, p=0.007). In thick ILT sections (A1), DL-Hcy decreased MMP-2 activity and t-PA and plasminogen concentrations compared to control tissue (respectively, 2.53 ± 2.02 versus 3.28 ± 2.65, p=0.006; 0.67 ± 0.57 versus 0.96 ± 0.91, p=0.021; 9.25 ± 4.59 versus 12.63 ± 9.56, p=0.017). In addition, analysis revealed positive correlations at all sites between activities/concentrations of MMP-2, TF, and PAI-1 measured in control tissues and after incubation with DL-Hcy. Conclusions. These data indicate the potential for excess Hcy to enhance damage of arterial wall in thinner AAA segments as a result of the increased activity of MMP-2 and fibrinolytic factors.

Thromboelastography in the management of snakebite-induced coagulopathy: a case series and literature review.

: North American crotaline envenomations are a significant source of morbidity annually. Envenomation is marked by a constellation of effects that can include severe tissue damage, systemic effects such as anaphylaxis, and coagulopathy with bleeding. Traditionally, static plasma-derived tests such as protime and PTT are used to assess coagulation status. However, other tests are available that provide a whole blood assessment of all portions of coagulation including initiation, propagation, and clot maintenance. One such analytical test is TEG. Minimal data is available regarding the effect of snake venom on TEG results. We report on three cases of North American crotaline envenomation with subsequent TEG analysis. Our results show that TEG analysis does relate the same information as traditional monitoring parameters. Although only in an isolated case, the TEG analysis potentially revealed a fibrinolytic process prior to the same process being apparent with traditional monitoring. Future research is necessary to identify the role of TEG in crotaline snakebite management.

The pathogenesis and available prevention options in patients with diabetic thrombophilia.

Diabetes mellitus (DM), a growing health problem itself, is accompanied by an increased risk of cardiovascular and thrombotic complications. The imbalance between coagulation and fibrinolysis processes observed in patients with diabetes may be defined as diabetic thrombophilia. Several mechanisms are involved in the hypercoagulability state in diabetics, including endothelial cell damage, altered platelet structure and function, increased microparticle formation, different structure of fibrin clots, disturbances in the activity of coagulation factors, fluctuations in the concentrations of fibrinolysis activators and inhibitors, and qualitative changes of proteins due to glycation and oxidation processes. These all are the reasons why DM is the most common cause of acquired thrombophilia. Moreover, diabetes changes the efficacy of certain medications. Results of various trials seem to suggest that thrombolytic drugs are less effective in patients suffering from this disease. The impact of DM on the effectiveness of treatment with acetylsalicylic acid (ASA) remains unclear. Awareness of thrombotic complications in diabetic patients may enable earlier diagnosis and proper therapy.

Hemostasis and Thrombosis in Extreme Temperatures (Hypo- and Hyperthermia).

The delicate biochemistry of coagulation and anticoagulation is greatly affected by deviations from the optimal temperature required for the interactions between various coagulation enzymes, cellular receptors, and intracellular mechanisms. Hyperthermia will lead to a prothrombotic state and, if sufficiently severe such as in heatstroke, a consumption coagulopathy, which will clinically manifest with the simultaneous appearance of intravascular thrombotic obstruction and an increased bleeding tendency. Hypothermia slows down the coagulation process, but as this seems to be adequately balanced by impairment of anticoagulant and fibrinolytic processes, its clinical effects are modest; however, hypothermia may be modestly linked to a somewhat higher risk of localized thrombosis. Restoration of a normal body temperature in patients affected by hyper- or hypothermia is the cornerstone for the management of associated coagulation derangements.

Pharmacological Differentiation of Thrombomodulin Alfa and Activated Protein C on Coagulation and Fibrinolysis In Vitro.

Although thrombomodulin alfa (TM alfa), recombinant human soluble thrombomodulin, exerts antithrombogenic effects through activated protein C (APC), clinical trials suggested that TM alfa has a lower bleeding risk than does recombinant human APC. To address the mechanism explaining this difference, effects of TM alfa and APC on thrombogenic, coagulation, and fibrinolytic processes were compared in vitro. TM alfa and APC inhibited generation of thrombogenic markers, thrombin, and prothrombin fragment F1+2 and prolonged coagulation parameters, activated clotting time (ACT), and activated partial thromboplastin time (APTT). Concentrations of TM alfa effective for thrombin and F1+2 generation inhibition were comparable to those of APC. However, effects of TM alfa on ACT and APTT were clearly weaker than those of APC. TM alfa significantly prolonged clot lysis time (CLT) and decreased LY30, a parameter of degree of fibrinolysis in thromboelastography, whereas APC significantly shortened CLT and increased LY30. These results suggested that while the antithrombogenic effects of TM alfa were similar to those of APC, its anticoagulant effects were lower. In addition, effects of TM alfa were antifibrinolytic, while those of APC were profibrinolytic.

Phenotypic Screening To Discover Novel Chemical Series as Efficient Antihemorrhagic Agents.

In an effort to find novel chemical series as antifibrinolytic agents, we explore α-phenylsulfonyl-α-spiropiperidines bearing different zinc-binding groups (ZBGs) to target those metalloproteinases involved in the fibrinolytic process: MMP3 and MMP10. Surprisingly, all these new chemical series were inactive against these metalloproteinases; however, several new molecules retained the antifibrinolytic activity in a phenotypic functional assay using thromboelastometry and human whole blood. Further optimization led to compound 38 as a potent antifibrinolytic agent in vivo, three times more efficacious than the current standard-of-care (tranexamic acid, TXA) at 300 times lower dose. Finally, in order to decipher the underlying mode-of-action leading to this phenotypic response, an affinity-based probe 39 was successfully designed to identify the target involved in this response: a potentially unknown mechanism-of-action in the fibrinolytic process.

KADAR PENGHAMBAT PENGURAIAN FIBRIN OLEH TROMBIN YANG TERGIATKAN (THROMBIN ACTIVATABLE FIBRINOLYSIS INHIBITOR) DI PASIEN KEGEMUKAN DENGAN DAN TANPA DIABETES MELLITUS

Obesity will cause the development of insulin resistance and endothelial dysfunction through fat metabolism hormones and cytokines products. The metabolic disorders is hallmarked by the decrease of insulin function and will cause disorders of coagulation andfibrinolysis. Thrombin-activatable fibrinolysis inhibitor (TAFI) is a newly found glycoprotein that has a role in the balance of coagulationand fibrinolytic process. The aim of this study was to analyze the TAFI level in obese subjects with and without diabetes mellitus. A crosssectional study was carried out by collecting data from TAFI level measurements at Prodia Clinical Laboratory Jakarta and Makassarduring January-March 2008. The data was analyzed using t test with SPSS version 12.0. From 80 samples that match with the criteria,was found in four (4) diabetes mellitus patients and 76 subjects without diabetes mellitus. The mean TAFI level in obese subjects withdiabetes mellitus was 7.67 ug/ml and in obese subjects without diabetes mellitus was 8.10 ug/ml. The result of t test showed p = 0.58.There was no significant difference between TAFI level in obese subjects with and without diabetes mellitus.

Blood clot parameters: Prejudgment of fibrinolysis in thromboelastography

BackgroundThromboelastography (TEG) is a physical method to simulate the whole process of coagulation and fibrinolysis in the human body environment, and then it also can quickly determine whether patients with hypercoagulable, low coagulation, fibrinolysis or other symptoms. The first step in the diagnosis is based on two parameters: Estimated percentage of lysis (EPL) or the Lysis at 30 min (LY30). These two parameters are used to determine whether the sample has fibrinolysis. However, the determination of LY30 takes a long time, although EPL can reflect real-time fibrinolysis, sometimes the secondary fibrinolysis is not obvious. Materials and methodsWe have an extensive database of results from TEG of fibrinolysis and healthy whole blood (WB). These results were generated using citrated WB, followed by the addition of CaCl2, to initiate clot formation. ResultsAccording to the characteristics of fibrinolysis, a new parameter clot retention time (CRT) was proposed to predict the status of fibrinolysis, and the normal range of the parameters was obtained in this paper. ConclusionIt is essential for the clinician to determine the fibrinolytic and ultimately contribute to the treatment of patients. We believe this parameter will add to the standardization of TEG parameters. The new parameter will also shorten the measurement time of non-fibrinolytic samples, which has definite physiological and pathological significance.

Evaluation of local intra-pleural application of tranexamic acid on postoperative blood loss in lung decortication surgery, a prospective, randomized, double-blind, placebo-controlled study.

Background:During decortication surgery, fibrous peel over the lung was removed to allow expansion of the lung and therefore, wide raw area was created with surface oozing. The phenomenon of fibrinolysis usually activated after such procedure, resulting in increasing the postoperative bleeding. Tranexamic acid is one of antifibrinolytic therapies that could be used topically and to targets directly the source of bleeding and reducing the local activation of the fibrinolytic process and consequently reducing the postoperative bleeding.Patients and Methods:A total of 70 patients underwent lung decortication surgery in Cardiothoracic Surgery Department at Tanta University Hospital from January 2015 to May 2017. Patients were randomly allocated into two groups, Group I (35 patients) receiving 3 g of tranexamic acid in 100 ml of saline solution and Group II (35 patients) receiving 100 ml of saline solution as placebo. At the end of the operation and before closing the chest, in both groups, drug or placebo solution was distributed locally all over the pleural cavity. Comparison between the groups was done regarding the amount of postoperative bleeding, postoperative hemoglobin in the first 24 and 48 h postoperatively, blood transfusion, Intensive Care Unit (ICU) stay, and hospital stay.Results:Both groups were comparable regarding demographic and surgical data. Group I patients had the significantly lesser amount of postoperative blood loss than Group II during the first postoperative 48 h, and hence, the need of postoperative blood transfusion was significantly lower in Group I with better postoperative hemoglobin level than Group II. However, there was no difference in overall ICU and hospital stay.Conclusion:The local intrapleural use of tranexamic acid after decortication surgery of the lung is safe and significantly reduces the amount of postoperative blood loss and in consequence reduces the amount of postoperative blood transfusion.

Synthesis and Biocompatibility Studies of New Iminodiacetic Acid Derivatives.

Background: Iminodiacetic acid (IDA) derivatives can be used as ligands to form complexes with technetium, with potential application as hepatobiliary diagnostic agents. The aim of this study was to synthesize five novel IDA derivatives and to compare their effects on plasma haemostasis with clinically approved ligands for technetium complexation. Methods: The influence of synthesized IDA derivatives on plasma haemostasis was evaluated spectrophotometrically by clot formation and lysis test (CL-test), coagulation assay, Prothrombin Time and Activated Partial Tromboplastin Time. The effects of the tested compounds on erythrocytes were assessed using haemolysis assays, microscopy and flow cytometry studies. Results: Despite their significant influence on the kinetic parameters of the process of clot formation and fibrinolysis, the tested ligands, at potential diagnostic concentrations, did not alter the overall potential of clot formation and lysis (CLAUC). At potential diagnostic concentrations (0.4 μmol/mL) all the tested compounds showed no adverse effects on the membranes of RBCs (Red Blood Cells). Conclusion: IDA derivatives with methoxy substituents in aromatic ring, exert multidirectional effects on plasma haemostasis and should be considered safe as their significant impacts were mostly observed at 4 μmol/mL, which is about 10-fold higher than the theoretical plasma concentrations of these compounds.

Innate immunity: Fibrinolytic and innate systems collide.

Nuclear factor of activated T cells (NFAT) regulates IFNγ production in natural killer cells and controls fibrinolytic processes required to clear skin infections.

Prolonged Hemocoagulase Agkistrodon Halys Pallas Administration Induces Hypofibrinogenemia in Patients with Hematological Disorders: A Clinical Analysis of 11 Patients.

Hemocoagulase Agkistrodon Halys Pallas, a well-established hemostatic agent used in China, has widespread applications in various conditions of bleeding and hemorrhages. Although it presents a low risk of vascular thrombosis, the agent can be easily tolerated and does not show any harmful effects, except allergy. However, long-term use of hemocoagulase may induce hypofibrinogenemia in some patients. Herein, we report 11 patients of prolonged administration of hemocoagulase Agkistrodon, with particular attention to the dynamic changes in their coagulation functions. A marked decline in the level of fibrinogen was observed in patients suffering from hematological disorders, following an extended exposure to hemocoagulase. Nevertheless, the low fibrinogen levels increased after withdrawal of the agent. Since the D-dimer level did not show any significant increase, hemocoagulase inducing lower fibrinogen may be considered to be a process of primary fibrinolysis. In conclusion, we suggested that hemocoagulase must be cautiously used in patients with hematological disorders, because of the potential risk of fibrinolysis; and the coagulation functions should be carefully evaluated during the administration of hemocoagulase.

In vitro and in silico studies on fibrinolytic activity of nattokinase: A clot buster from Bacillus sp.

Nattokinase (NK) is a serine protease enzyme of the subtilisin family. It exhibits a strong fibrinolytic activity. The fibrinolytic enzymes from Bacillus sp. have attracted interest as thrombolytic agents because of their efficiency in the fibrinolytic process including plasmin activation. In the present study, VIT garden soil was collected and subjected to isolation process in order to screen for the NK production. Screening for NK enzyme was performed by radial caseinolytic assay. The production of NK enzyme was done in two different production medium for comparative studies. The NK enzyme was purified by gel permeation chromatography. The activity of the purified NK was checked by clot lysis and casein digestion assay. To investigate the structural basis of NK and fibrinogen interaction and also to identify the best binding mode, molecular dynamics and docking studies were performed. Based on the morphological and biochemical characterization, the isolate was identified as Bacillus sp. The overall purification fold of NK was about 3 with the specific activity of 664U/mg and 9.9% yield. Homogeneity of the purified enzyme was analyzed and confirmed by the single band obtained in SDS-PAGE. Molecular weight of the purified protease was estimated as 25 kDa. Purified NK enzyme exhibited 97% of effective clot lysis activity. The NK was docked in to the knob region of the fibrinogen at its binding site using Dock server. A total of 26 residues of fibrinogen and 29 residues of NK constitute the interface region. However, 9 residues of fibrinogen (THR238, MET264, LYS266, ARG275, THR277, ALA279, ASN308, MET310, and LYS321) and 8 residues of NK (GLY61, SER63, THR99, PHE189, LEU209, TYR217, ASN218, and MET222) are involved in intact binding. A significant amount of NK enzyme was obtained from Bacillus sp. The docking analysis revealed that the NK and fibrinogen adopt an extended binding pattern and interacts with the crucial residues to exhibit their activity.

Anticoagulation in cirrhosis: a new paradigm?

The liver plays a crucial role in coagulation cascade. Global hemostatic process is profoundly influenced by the presence of liver disease and its complications. Patients with cirrhosis have impaired synthesis of most of the factors involved in coagulation and fibrinolysis process due to a reduced liver function and altered platelet count secondary to portal hypertension. Altered routine tests and thrombocytopenia were considered in the past as associated with increased risk of bleeding. These concepts explain both the routine use of plasma and/or platelets transfusion in patients with liver cirrhosis, especially before invasive procedures, and why these patients were considered “auto-anticoagulated”. New recent evidences show that patients with liver cirrhosis have a more complex hemostatic alteration. Despite the presence of altered levels of factors involved in primary hemostasis, coagulation and fibrinolysis, patients with stable cirrhosis have a rebalanced hemostatic, which however can easily be altered by decompensation or infection, both in hemorrhagic or thrombotic direction. Patients with cirrhosis have an increased risk of venous thrombotic events (namely portal vein thrombosis) while bleeding seems to be related to the grade of portal hypertension rather than to a hemostatic imbalance. The use of anticoagulants both as treatment or prophylaxis is safe, reduces the rate of portal vein thrombosis and decompensation, and improves survival. Standard laboratory coagulation tests are unable to predict bleeding and are inadequate for the assessment of hemostatic status in these patients, hence more comprehensive tests are required to guide the management of thrombotic and bleeding complications.