Fibrinolytic Research Articles

Clot-Targeted Nanogels for Dual-Delivery of AntithrombinIII and Tissue Plasminogen Activator to Mitigate Disseminated Intravascular Coagulation Complications.

Disseminated intravascular coagulation (DIC) is a pathologic state that follows systemic injury and other diseases. Often a complication of sepsis or trauma, DIC causes coagulopathy associated with paradoxical thrombosis and hemorrhage. DIC upregulates the thrombotic pathways while simultaneously downregulating the fibrinolytic pathways that cause excessive fibrin deposition, microcirculatory thrombosis, multiorgan dysfunction, and consumptive coagulopathy with excessive bleeding. Given these opposing disease phenotypes, DIC management is challenging and includes treating the underlying disease and managing the coagulopathy. Currently, no therapies are approved for DIC. We have developed clot-targeted therapeutics that inhibit clot polymerization and activate clot fibrinolysis to manage DIC. We hypothesize that delivering both an anticoagulant and a fibrinolytic agent directly to clots will inhibit active clot polymerization while also breaking up pre-existing clots; therefore, reversing consumptive coagulopathy and restoring hemostatic balance. To test this hypothesis, we single- and dual-loaded fibrin-specific nanogels (FSNs) with antithrombinIII (ATIII) and/or tissue plasminogen activator (tPA) and evaluated their clot preventing and clot lysing abilities in vitro and in a rodent model of DIC. In vivo, single-loaded ATIII-FSNs decreased fibrin deposits in DIC organs and reduced blood loss when DIC rodents were injured. We also observed that the addition of tPA in dual-loaded ATIII-tPA-FSNs intensified the antithrombotic and fibrinolytic mechanisms, which proved advantageous for clot lysis and restoring platelet counts. However, the addition of tPA may have hindered wound healing capabilities when an injury was introduced. Our data supports the benefits of delivering both anticoagulants and fibrinolytic agents directly to clots to reduce the fibrin load and restore hemostatic balance in DIC.

Hypercoagulability based on thromboelastography is common in dogs undergoing adrenalectomy.

Thromboelastography (TEG) is a whole blood assay that yields global assessment of hemostasis, as it evaluates clot time, strength, and kinematics of clot formation and lysis. The main objective was to describe preoperative TEG findings in dogs that had an adrenalectomy performed and, secondarily, to describe TEG findings in the dogs with or without hyperadrenocorticism (HAC). 30 dogs that had preoperative TEG and adrenalectomy performed. Medical records between 2018 and 2022 were reviewed. Signalment, diagnostic data, and perioperative treatment were abstracted. 53% (16/30) of the dogs were hypercoagulable, and none were hypocoagulable. Based on histopathology, 6 of 9 dogs with adenocarcinoma were hypercoagulable, 4 of 8 with pheochromocytoma were hypercoagulable, and 6 of 10 with adenoma were hypercoagulable. None of the 3 dogs with other histopathologic diagnoses or combinations of diagnoses (adrenocortical hyperplasia, poorly differentiated sarcoma, and both adrenocortical adenocarcinoma and pheochromocytoma) were hypercoagulable. Of the 14 dogs tested preoperatively for HAC, 4 of 8 HAC dogs were hypercoagulable and 2 of 6 non-HAC dogs were hypercoagulable. The present report describes for the first time TEG findings for dogs undergoing adrenalectomy and suggests that the majority of dogs with adrenal neoplasia are hypercoagulable based on TEG results.

Coagulation and fibrinolytic markers offer utility when distinguishing between benign and malignant gallbladder tumors: A cross-sectional study

BackgroundThe metabolic or proliferative abnormalities that are characteristic of tumor cells can lead to abnormal fibrinolysis or coagulation system activity, with certain tumors exhibiting hypercoagulability or existing in a fibrinolytic state. However, the utility of biomarkers of coagulation and fibrinolysis when seeking to differentiate between benign gallbladder disease and malignant gallbladder tumors remains uncertain. MethodsThis study included a total of 81 patients with benign gallbladder polyps and 94 patients with malignant gallbladder tumors. Pre-biopsy or pretreatment levels of PT, APTT, FIB, D-dimer, FDP, PLT, PIC, TAT, TM, and t-PAIC from these patients were compared using Mann-Whitney tests. The baseline data of the patients were analyzed using chi-square tests, and the diagnostic utility of these biomarkers in distinguishing between benign and malignant gallbladder lesions was evaluated using ROC curves, and Spearman correlation analysis was employed to assess the correlation between these indicators and tumor parameters. ResultsThe average age of malignant gallbladder tumor group was higher than benign gallbladder polyp group. And the base line analysis showed that there was a statistic difference in age, history of smoking, drinking, biliary tract disease, BMI of over weight between these two groups. In patients with malignant gallbladder tumors, FIB, D-dimer, FDP, PIC, TAT, TM, and t-PAIC levels were significantly elevated relative to those in patients affected by benign gallbladder polyp. The AUC for FIB, D-dimer, and FDP was 0.8469, 0.6514, 0.5950, while for PIC, TAT, TM, t-PAIC and four biomarker combined diagnosed was 0.8455, 0.6554, 0.7130, 0.6806, and 0.8859. Among these, TM was associated with the vascular invasion of tumor patients; TAT and t-PAIC were associated with neural invasion; D-dimer and FDP were related to the maximum tumor diameter; and FDP had a certain correlation with the tumor stage. ConclusionsIn gallbladder tumor patients, conventional coagulation metrics like FIB, D-dimer, and FDP, as well as newer thrombotic indicators such as PIC, TAT, TM, and t-PAIC, were obviously increased. Correlations with tumor parameters suggested their potential as biomarkers to distinguish benign from malignant gallbladder growths.

Production of Nattokinase from Bacillus amyloliquefaciens MRS18: A Bacterial Strain Isolated from Fermented Beans.

Nattokinase (NK) is a naturally occurring fibrinolytic protease enzyme obtained from the traditional Japanese food called Natto and has several uses in the pharmaceutical and medical industries. Nowadays, the most often used thrombolytic agent in the clinical field is NK, in part because it is less expensive than other thrombolytic medicines. The objective of this study is to investigate the screening, isolation and characterization of the NK enzyme-producing Bacillus strain from fermented Soya beans. The sample of fermented soya beans were tested for the presence of fibrinolytic protease- producing bacteria, followed by the screening, extraction, characterization and clot lysis assays. A total of three isolates were screened for caseinolytic activities by casein hydrolysis assay. Out of those isolates, MRS18 was found to be potent in producing the enzyme proteinase. To determine the taxonomy and phylogeny of these isolates, biochemical and molecular characterization has been carried out. Bacillus amyloliquefaciens MRS18 has been found with the highest caseinolytic activity. The clot lysing ability of the potent strain Bacillus amyloliquefaciens was found to be 61.7% after 120 min, and on further purification, by ammonium sulphate precipitation method, the lysis percentage was found to be 656% after 120 min. From the results of the present study, we concluded that Bacillus amyloliquefaciens isolated from the fermented soya beans produced an NK enzyme that exhibits immense potential to lyse blood clots.

Packaging of supplemented urokinase into alpha granules of in vitro–grown megakaryocytes for targeted nascent clot lysis

AbstractFibrinolytics delivered into the general circulation lack selectivity for nascent thrombi, reducing efficacy and increasing the risk of bleeding. Urokinase-type plasminogen activator (uPA) transgenically expressed within murine platelets provided targeted thromboprophylaxis without causing bleeding but is not clinically feasible. Recent advances in generating megakaryocytes prompted us to develop a potentially clinically relevant means to produce “antithrombotic” platelets from CD34+ hematopoietic stem cell–derived in vitro–grown megakaryocytes. CD34+ megakaryocytes internalize and store in alpha granules (α-granules) single-chain uPA (scuPA) and a plasmin-resistant thrombin-activatable variant (uPAT). Both uPAs colocalized with internalized factor V (FV), fibrinogen and plasminogen, low-density lipoprotein receptor–related protein 1 (LRP1), and interferon-induced transmembrane protein 3, but not with endogenous von Willebrand factor (VWF). Endocytosis of uPA by CD34+ megakaryocytes was mediated, in part, via LRP1 and αIIbβ3. scuPA-containing megakaryocytes degraded endocytosed intragranular FV but not endogenous VWF in the presence of internalized plasminogen, whereas uPAT-megakaryocytes did not significantly degrade either protein. We used a carotid artery injury model in nonobese diabetic-severe combined immunodeficiency IL2rγnull (NSG) mice homozygous for VWFR1326H (a mutation switching binding VWF specificity from mouse to human glycoprotein Ibα) to test whether platelets derived from scuPA- or uPAT-megakaryocytes would prevent thrombus formation. NSG/VWFR1326H mice exhibited a lower thrombotic burden after carotid artery injury compared with NSG mice unless infused with human platelets or megakaryocytes, whereas intravenous injection of uPA-megakaryocytes generated sufficient uPA-containing human platelets to lyse nascent thrombi. These studies describe the use of in vitro–generated megakaryocytes as a potential platform for delivering uPA or other ectopic proteins within platelet α-granules to sites of vascular injury.

Green Synthesis of Silver Nanoparticles using Coriandrum sativum and Murraya koenigii Leaf Extract and its Thrombolytic Activity.

Plants have been used for ages in traditional medicine, and it is exciting to perceive how recent research has recognized the bioactive compounds liable for their beneficial effects. Green synthesis of metal nanoparticles is a hastily emergent research area in nanotechnology. This study describes the synthesis of silver nanoparticles (AgNPs) using Coriandrum sativum and Murraya koenigii leaf extract and its thrombolytic activity. The aim of the study was to determine the clot lysis activity of Coriandrum sativum and Murraya koenigii synthesized silver nanoparticles. Leaves of Coriandrum sativum and Murraya koenigii were collected. Methanolic extraction of the plant sample was done through a Soxhlet extractor. The methanolic extract obtained from both the leaves was subjected to GC-MS analysis. The synthesized NPs from leaf extracts were monitored for analysis, where the typical X-ray diffraction pattern and its diffraction peaks were identified. 3D image of the NPs was analysed by Atomic Force Microscopy. The surface charge of nanoparticles was identified by Zeta potential. The Clot lysis activity of Coriandrum sativum and Murraya koenigii synthesized silver nanoparticles were analysed by the modified Holmstorm method. The thrombolytic property of the methanolic extract of plants Coriandrum sativum showed clot lysis activity at 2.5 mg/mL with 45.99% activity, and Murraya koenigii extract with 66.56% activity. The nanoparticles (Nps) from Coriandrum sativum showed clot lysis activity at 2.5 mg/mL with 58.29% activity, and NPs from Murraya koenigii with 54.04% activity. Coriandrum sativum in GC-MS exhibited 3 peaks, whereas Murraya koenigii extract showed five peaks with notable bioactive compounds. These NPs were further used for biomedical applications after being fixed by an organic encapsulation agent. The present research reveals the usefulness of Coriandrum sativum and Murraya koenigii for the environmentally friendly manufacture of silver nanoparticles.

Targeting tissue factor pathway inhibitor with concizumab to improve hemostasis in patients with Glanzmann thrombasthenia: an in vitro study

BackgroundGlanzmann thrombasthenia (GT) is caused by an inherited defect of platelet αIIbβ3 integrin. Concizumab, a monoclonal antibody specific for tissue factor pathway inhibitor, abolishes its anticoagulant effect. ObjectivesTo evaluate the in vitro ability of concizumab to improve hemostasis in GT. MethodsThe effects of concizumab were evaluated in whole blood or platelet-rich plasma from GT patients (n = 5-9) using a thrombin generation assay, rotational thromboelastometry (ROTEM), a global fibrinolytic capacity assay, and a flow chamber assay (Total Thrombus formation Analysis System). Washed platelets (WPs) and 20 nM recombinant activated factor VII (rFVIIa) were included for comparison. ResultsThe lag time in the thrombin generation assay was significantly longer (+85%; P < .0001) in GT patients than in controls. WPs, rFVIIa, and concizumab each significantly improved thrombin generation profiles. The ROTEM clotting time (CT) was significantly longer in GT patients than in controls (677 seconds vs 523 seconds; P = .03). However, CT improved after adding WPs, rFVIIa, or concizumab. Under flow, occlusive thrombi were present in all healthy controls after 10 minutes, whereas platelet-fibrin depositions were not seen in GT patients. Subocclusive or occlusive thrombi formed when GT blood was mixed with WPs, rFVIIa, or concizumab. Clots in GT platelet-rich plasma were more susceptible to fibrinolysis and were improved by WPs, rFVIIa, or concizumab. ConclusionConcizumab enhanced thrombin generation, decreased the ROTEM CT, improved thrombus formation under flow, and reduced clot lysis. Our results demonstrate the potential of concizumab for subcutaneous prophylaxis in GT patients.

Membrane-stabilizing and clot lysis activities of (±) citronellal: In-vitro studies

Numerous plants and their phytochemicals have been identified as having potential medicinal values and are used to treat different ailments worldwide. (±) Citronellal (CTL) is a monoterpene phytochemical with potential therapeutic benefits including anti-inflammatory, antioxidant, antifungal, and antibacterial actions. AimThe present study aimed to evaluate the membrane-stabilizing and clot-lysis activities of CTL through in vitro studies. For this, we performed hypotonic solution-induced erythrocyte lysing and human blood clot lysis methods to check the membrane-stabilizing and clot lysis capacities of CTL using acetylsalicylic acid (ASA) and streptokinase (STK) as standards, respectively. CTL exhibited concentration-dependent membrane-stabilizing activity with an IC50 = 28.83 ± 1.19 µg/mL. In the clot lysis test, CTL also showed a concentration-dependent clot lysing capacity, where it exhibited 50.46 ± 3.81 % clot lysis at a concentration of 160 μg/mL. In the latter case, the IC50 value was I158.22 ± 2.21 µg/mL. CTL exhibit potent membrane-stabilizing and moderate clot-lysis activity. We suppose that CTL may exert these effects, possibly through its capacity to inhibit inflammatory mediators. Further in vivo and in silico studies are required to elucidate CTL’s exact molecular mechanisms behind these biological effects.

Complement activation is associated with right ventricular dysfunction and the severity of pulmonary embolism: links with prothrombotic state.

Little is known about the role of complement activation in acute pulmonary embolism (PE). We investigated whether complement activation is associated with the severity of acute PE, along with the associated prothrombotic state, systemic inflammation and neutrophil extracellular traps (NETs) formation. We studied 109 normotensive, non-cancer PE patients (aged 58.1±15.0 years). On admission prior to initiation of anticoagulation, plasma soluble complement components, i.e., C3a and sC5b-9, were measured with enzyme-linked immunosorbent assay (ELISA), along with thrombin generation, fibrinolysis proteins (plasminogen, antiplasmin, plasminogen activator inhibitor-1), factor VIII (FVIII) activity, and fibrin clot properties, including clot permeability (Ks, a measure of clot density) and clot lysis time (CLT). Moreover, we determined inflammatory markers and citrullinated histone H3, a specific marker of NETs formation. Patients in the lower tertile of C3a (≤1.45 ng/mL, n=37) had lower simplified Pulmonary Embolism Severity Index (sPESI) values and were less likely to have right ventricular (RV) dysfunction compared to the remaining subjects. The former subgroup also had 13% lower FVIII activity, but not fibrinogen, interleukin-6, fibrinolysis proteins, and thrombin generation. Plasma C3a levels correlated inversely with Ks and positively with CLT indicating formation of denser and poorly lysable clots in subjects with elevated C3a. Despite a positive association between C3a and sC5b-9, the latter parameter was solely associated with higher FVIII, but not with other variables. We showed that in acute PE enhanced complement activation characterizes patients with poorer short-term prognosis who display prothrombotic fibrin clot properties and elevated FVIII, which supports the involvement of complement proteins in acute thromboembolism.

&lt;i&gt;In vitro&lt;/i&gt; influence on fibrinolysis parameters of aqueous extracts of mushrooms growing on woody plants

Introduction. Various compounds of mushroom extracts are of interest as a source of new active pharmaceutical substances with antithrombotic activity.Aim. The aim of the work is to determine the potential antithrombotic activity in vitro of aqueous extracts of xylotrophic mushrooms: Lentinula edodes, Pholiota squarrosa, Flammulina velutipes (Curtis) Signer, Kuehneromyces mutabilis, Armillaria cepistipes, Armillaria borealis, Hypholoma capnoides, Lentinellus cochleatus, Léccinum aurantíacum, Pleurotus ostreatus.Materials and methods. The study of fibrinolytic activity of water extracts of mushrooms in vitro was carried out. The effect of test objects on plasminogen activation, on the activity of enzyms of the fibrinolysis system α2-antiplasmin, on the time of euglobulin clot lysis and on the concentration of fibrinogen was evaluated. For the analysis we used reagent kits of SPD «RENAM» and LLC Company «Тechnology-Standard» (Russia).Results and discussion. Pronounced antithrombotic activity is demonstrated by an aqueous extract of L. edodes and fractions of aqueous extract. Extracts of L. edodes affect all studied parameters of fibrinolysis to varying degrees. Caps and stems of P. squarrosa, K. mutabilis, and F. velutipes aqueous extracts and fractions of aqueous extracts also demonstrate antithrombotic (fibrinolytic) activity in vitro, having a statistically significant effect on various parameters of fibrinolysis. A less pronounced effect on fibrinolysis parameters in comparison with the above samples is observed when using an aqueous extract of L. cochleatus marked. A. cepistipes, A. borealis, H. capnoides, L. aurantiacum, P. ostreatus fruiting bodies aqueous extracts do not affect the studied parameters of fibrinolysis and the concentration of fibrinogen.Conclusion. The data obtained make it possible to recommend extracts from L. edodes, F. velutipes, P. squarrosa and K. mutabilis for further study, for example, in order to develop functional food products for cardiovascular diseases or to create antithrombotic drugs.

Units of fibrinolytic system in mice with urokinase gene knockout in presence of growing B16/F10 melanoma

Units of fibrinolytic system in mice with urokinase gene knockout in presence of growing B16/F10 melanoma

Daytime plasma cortisol and cortisol response to dexamethasone suppression are associated with a prothrombotic state in hypertension.

A prothrombotic state was demonstrated in patients with Cushing's syndrome and is involved in the development and progression of cardiovascular and renal damage in hypertensive patients. This study was designed to examine the relationships between cortisol secretion and the hemostatic and fibrinolytic systems in hypertension. In 149 middle-aged, nondiabetic, essential hypertensive patients free of cardiovascular and renal complications, we measured hemostatic markers that express the spontaneous activation of the coagulation and fibrinolytic systems and assessed daily cortisol levels (8 AM, 3 PM, 12 AM; area under the curve, AUC-cortisol) together with the cortisol response to dexamethasone overnight suppression (DST-cortisol). Plasma levels of D-dimer (D-dim), prothrombin fragment 1+2 (F1+2), and von Willebrand factor (vWF) were progressively and significantly higher across tertiles of AUC-cortisol and DST-cortisol, whereas no differences were observed in fibrinogen, tissue plasminogen activator, plasminogen activator inhibitor-1, antithrombin III, protein C, and protein S. D-dim, F1+2, and vWF were significantly and directly correlated with age and both AUC-cortisol and DST-cortisol. Multivariate regression analysis showed that both AUC-cortisol and DST-cortisol were related to plasma D-dim, F1+2, and vWF independently of age, body mass index, blood pressure, and renal function. Greater daily cortisol profile and cortisol response to overnight suppression are independently associated with a prothrombotic state in hypertensive patients and might contribute to the development of organ damage and higher risk of cardiovascular complications.

Comparative studies between humans and golden Syrian hamsters via thromboelastography.

Thromboelastography (TEG) is a widely utilized clinical testing method for real-time monitoring of platelet function and the thrombosis process. Lipid metabolism disorders are crucial risk factors for thrombosis. The lipid metabolism characteristics of hamsters resemble those of humans more closely than mice and rats, and their relatively large blood volume makes them suitable for studying the mechanisms of thrombosis related to plasma lipid mechanisms. Whole blood samples from golden Syrian hamsters and healthy humans were obtained following standard clinical procedures. TEG was employed to evaluate coagulation factor function, fibrinogen (Fib) function, platelet function, and the fibrinolytic system. The whole blood from hamster or healthy human was isolated following the clinical procedure, and TEG was employed to evaluate the coagulation factor function, Fib function, platelet function, and fibrinolytic system. Coagulation analysis used ACLTOP750 automatic coagulation analysis pipeline. Blood routine testing used XN-2000 automatic blood analyzer. TEG parameters revealed that hamsters exhibited stronger coagulation factor function than humans (reaction time [R], p = 0.0117), with stronger Fib function (alpha angle, p < 0.0001; K-time [K], p < 0.0001). Platelet function did not differ significantly (maximum amplitude [MA], p = 0.077). Hamsters displayed higher coagulation status than humans (coagulation index [CI], p = 0.0023), and the rate of blood clot dissolution in hamsters differed from that in humans (percentage lysis 30 min after MA, p = 0.02). Coagulation analysis parameters indicated that prothrombin time (PT) and activated partial thromboplastin time (APTT) were faster in hamsters than in humans (PT, p = 0.0014; APTT, p = 0.03), whereas the Fib content was significantly lower in hamsters than in humans (p < 0.0001). No significant difference was observed in thrombin time (p = 0.1949). In summary, TEG could be used to evaluate thrombosis and bleeding parameters in whole blood samples from hamsters. The platelet function of hamsters closely resembled that of humans, whereas their coagulation function was significantly stronger.

Injectable polyamide-amine dendrimer-crosslinked meloxicam-containing poly-γ-glutamic acid hydrogel for prevention of postoperative tissue adhesion through inhibiting inflammatory responses and balancing the fibrinolytic system

Establishing a physical barrier between the peritoneum and the cecum is an effective method to reduce the risk of postoperative abdominal adhesions. Meloxicam (MX), a nonsteroidal anti-inflammatory drug has also been applied to prevent postoperative adhesions. However, its poor water solubility has led to low bioavailability. Herein, we developed an injectable hydrogel as a barrier and drug carrier for simultaneous postoperative adhesion prevention and treatment. A third-generation polyamide-amine dendrimer (G3) was exploited to dynamically combine with MX to increase the solubility and the bioavailability. The formed G3@MX was further used to crosslink with poly-γ-glutamic acid (γ-PGA) to prepare a hydrogel (GP@MX hydrogel) through the amide bonding. In vitro and in vivo experiments evidenced that the hydrogel had good biosafety and biodegradability. More importantly, the prepared hydrogel could control the release of MX, and the released MX is able to inhibit inflammatory responses and balance the fibrinolytic system in the injury tissues in vivo. The tunable rheological and mechanical properties (compressive moduli: from ∼ 57.31 kPa to ∼ 98.68 kPa;) and high anti-oxidant capacity (total free radical scavenging rate of ∼ 94.56 %), in conjunction with their syringeability and biocompatibility, indicate possible opportunities for the development of advanced hydrogels for postoperative tissue adhesions management.

Interactions and Trends of Interleukins, PAI-1, CRP, and TNF-α in Inflammatory Responses during the Perioperative Period of Joint Arthroplasty: Implications for Pain Management-A Narrative Review.

Inflammation during the perioperative period of joint arthroplasty is a critical aspect of patient outcomes, influencing both the pathophysiology of pain and the healing process. This narrative review comprehensively evaluates the roles of specific cytokines and inflammatory biomarkers in this context and their implications for pain management. Inflammatory responses are initiated and propagated by cytokines, which are pivotal in the development of both acute and chronic postoperative pain. Pro-inflammatory cytokines play essential roles in up-regulating the inflammatory response, which, if not adequately controlled, leads to sustained pain and impaired tissue healing. Anti-inflammatory cytokines work to dampen inflammatory responses and promote resolution. Our discussion extends to the genetic and molecular influences on cytokine production, which influence pain perception and recovery rates post-surgery. Furthermore, the role of PAI-1 in modulating inflammation through its impact on the fibrinolytic system highlights its potential as a therapeutic target. The perioperative modulation of these cytokines through various analgesic and anesthetic techniques, including the fascia iliac compartment block, demonstrates a significant reduction in pain and inflammatory markers, thus underscoring the importance of targeted therapeutic strategies. Our analysis suggests that a nuanced understanding of the interplay between pro-inflammatory and anti-inflammatory cytokines is required. Future research should focus on individualized pain management strategies.

Thrombin generation, fibrin clot permeation and lysis in patients with severe mitral regurgitation undergoing transcatheter edge-to-edge mitral valve repair: Mitral Fibrin Study.

Intricate management of heart failure (HF), especially in the context of reduced ejection fraction, is further complicated by an elevated risk of thromboembolic events. Studies published so far offer inconclusive insight into the interplay between mitral regurgitation (MR) and the coagulation system. This study aimed to investigate the impact of transcatheter edge‑to‑edge repair (TEER) on specific coagulation parameters in HF patients. A cohort of 31 HF patients with severe MR treated with TEER underwent a systematic evaluation at 3 visits (V1, V2, and V3). Coagulation parameters, including fibrinogen concentration, thrombin generation, fibrin clot permeability, and clot lysis time (CLT) were assessed (n = 27 at V2; n = 25 at V3). TEER induced changes in fibrinogen levels (P = 0.01; V3 vs V2) and improved fibrin clot properties over 50‑day follow‑up (P = 0.01; V3 vs V2). No significant differences were observed between time points in the analyzed blood clot parameters. Correlation analysis showed that baseline CLT was associated with ΔN‑terminal pro-B‑type natriuretic peptide (NT‑proBNP) level (P = 0.049; r = 0.4). Multivariable analysis identified baseline CLT as an independent predictor of early post‑TEER NT‑proBNP change (R2 = 0.55; P = 0.02). We found decreased level of fibrinogen and increased permeation coefficient over a median 50 (interquartile range, 32.5-75.5)-day post‑TEER follow‑up, as compared with early postprocedural assessments. Other blood coagulation parameters remained unchanged from baseline at both follow‑up time points after TEER. Finally, CLT was an independent predictor of early NT‑proBNP increase, emphasizing its role as an indicator of hemodynamic response to TEER.

Homocysteine thiolactone and other sulfur-containing amino acid metabolites are associated with fibrin clot properties and the risk of ischemic stroke

Homocysteine (Hcy) and Hcy-thiolactone (HTL) affect fibrin clot properties and are linked to cardiovascular disease. Factors that influence fibrin clot properties and stroke are not fully understood. To study sulfur-containing amino acid metabolites, fibrin clot lysis time (CLT) and maximum absorbance (Absmax) in relation to stroke, we analyzed plasma and urine from 191 stroke patients (45.0% women, age 68 ± 12 years) and 291 healthy individuals (59.7% women, age 50 ± 17 years). Plasma and urinary levels of sulfur-containing amino acid metabolites and fibrin clot properties were significantly different in stroke patients compared to healthy individuals. Fibrin CLT correlated with fibrin Absmax in healthy males (R2 = 0.439, P = 0.000), females (R2 = 0.245, P = 0.000), female stroke patients (R2 = 0.187, P = 0.000), but not in male stroke patients (R2 = 0.008, P = ns). Fibrin CLT correlated with age in healthy females but not males while fibrin Absmax correlated with age in both sexes; these correlations were absent in stroke patients. In multiple regression analysis in stroke patients, plasma (p)CysGly, pMet, and MTHFR A1298C polymorphism were associated with fibrin Absmax, while urinary (u)HTL, uCysGly, and pCysGly were significantly associated with fibrin CLT. In healthy individuals, uHTL and uGSH were significantly associated with fibrin Absmax, while pGSH, and CBS T833C 844ins68 polymorphism were associated with fibrin CLT. In logistic regression, uHTL, uHcy, pCysGly, pGSH, MTHFR C677T polymorphism, and Absmax were independently associated with stroke. Our findings suggest that HTL and other sulfur-containing amino acid metabolites influence fibrin clot properties and the risk of stroke.

Association of Metallic and Nonmetallic Elements with Fibrin Clot Properties and Ischemic Stroke.

Objectives-Metallic elements and fibrin clot properties have been linked to stroke. We examined metallic and nonmetallic elements, fibrin clot lysis time (CLT), and maximum absorbance (Absmax) in relation to ischemic stroke. Design-A case-control study of ischemic stroke patients vs. healthy individuals. Subjects and Methods-Plasma and serum were collected from 260 ischemic stroke patients (45.0% women; age, 68 ± 12 years) and 291 healthy controls (59.7% women; age, 50 ± 17 years). Fibrin CLT and Absmax were measured using a validated turbidimetric assay. Serum elements were quantified by inductively coupled plasma mass spectrometry (ICP-MS) and optical emission spectrometry (ICP-OES). Data were analyzed by bivariate correlations and multiple or logistic regression. Results-In female stroke patients, copper, lithium, and aluminum were significantly lower compared with controls; in male stroke patients, potassium was lower, and beryllium was elevated. In female and male stroke patients, iron, zinc, nickel, calcium, magnesium, sodium, and silicon were significantly lower, while strontium was elevated. Positive correlations between fibrin clot properties and metals, observed in healthy controls, were lost in ischemic stroke patients. In multivariate regression analysis, fibrin CLT and/or Absmax was associated with zinc, calcium, potassium, beryllium, and silicon in stroke patients and with sodium, potassium, beryllium, and aluminum in controls. In logistic regression analysis, stroke was independently associated with lithium, nickel, beryllium, strontium, boron, and silicon and with sodium, potassium, calcium, and aluminum but not with fibrin CLT/Absmax. Conclusions-Various elements were associated with fibrin clot properties and the risk of ischemic stroke. Lithium, sodium, calcium, and aluminum abrogated the association of fibrin clot properties with ischemic stroke.

The Role of Fibrinolytic Activity of Blood Plasma in the Development of Obstetric Complications in Overweight Pregnant Women with Metabolic Dysfunction-Associated Steatohepatitis: A Prospective Cohort Study

Background: Obesity and hyperlipidaemia during pregnancy increase the risk of metabolic dysfunction-associated steatotic liver disease. Our aim was to evaluate changes in the fibrinolytic system in overweight pregnant women due to metabolic dysfunction-associated steatohepatitis, compared with a control group and its impact on the development of gestational complications. Methods: Prospective cohort study included 69 overweight pregnant women with metabolic dysfunction-associated steatohepatitis and 30 healthy pregnant women (control group). All pregnants with metabolic dysfunction-associated steatohepatitis and obesity were divided into 3 subgroups: IA – 23 overweight pregnant women, IB – 25 pregnants with obesity grade 1, IC – 24 pregnant women with obesity grade 2. To evaluate the fibrinolysis system, we studied total, enzymatic and non-enzymatic fibrinolytic activity of plasma. Results: The total fibrinolytic activity of IA, IB and IC groups were, respectively, 15%, 19% and 23% lower than that of controls and the enzymatic fibrinolytic activity in overweight pregnant women with metabolic dysfunction-associated steatohepatitis of IA, IB and IC groups were, respectively 28%, 43% and 54% lower than in controls. Marked suppression of the total fibrinolytic activity and enzymatic fibrinolytic activity of the blood plasma was established in overweight pregnant women with metabolic dysfunction-associated steatohepatitis. These changes can serve as a prerequisite for the occurrence of microthrombosis with the subsequent development of placental dysfunction, fetal growth restriction and fetal distress. Conclusions: There are disorders in the system of the coagulation function with a tendency to peripheral microthrombosis, disseminated intravascular coagulation syndrome and macrothrombosis, which can be early prognostic criteria in the development of obstetric and perinatal complications in overweight pregnant women with metabolic dysfunction-associated steatohepatitis. Clinical Trial Registration: This work is a fragment of the complex research work of the Department of Obstetrics and Gynecology of I. Horbachevsky Ternopil National Medical University of the Ministry of Health of Ukraine “A comprehensive approach to symptom control, recurrent and long-term prognosis in conditions of comorbid pathology in internal medicine clinic and family doctor’s practice” (state registration number 0223U000022, 0118U000361 https://nrat.ukrintei.ua/en/searchdoc/).

ANTIOXIDANT, ANTI-INFLAMMATORY, ANTIDIABETIC, ANTHELMINTHIC, AND THROMBOLYTIC ACTIVITY OF THE METHANOL EXTRACT OF CITRULLUS LANATUS PEEL

Background: The present study aimed for in vitro evaluation of Citrullus lanatus Peel (CLP) methanol extract for antioxidative, anti-inflammatory, antidiabetic, anthelmintic, and thrombolytic activity. Method: CLP was extracted with methanol and tested for in vitro antioxidative (DPPH and H2O2 scavenging method), thrombolytic, anthelmintic, antidiabetic, and anti-inflammatory properties (human red blood cell membrane stabilization method, α-amylase inhibitory effect by using Iodine-Starch method. Results: The percentage yield was 7% and phytochemical profile showed the presence of steroid, tannins, saponins, flavonoids, glycoside and reducing sugar. In DPPH scavenging assay, 88.7% was at 1000 g/ml, whereas 34% to 48.9% (IC50 27.65 µg/ml) in H2O2 and from 61.8% to 95.2% (IC50 value: 07 µg/ml) in ASCOR. In in vitro anti-inflammatory activity, the maximum protection percentage was observed 58% and 44%. Consecutively, in anti-diabetic activity, IC50 values of 1137.64 μg/ml whereas reference (Acarbose) exhibited IC50 at 671.71 μg/ml. On the other hand, the inhibitory effect of Acarbose showed 57% and 98% respectively at 2000 μg/ml. In earthworms, it considered as very potential for the helminthic infection and helminthiasis. A significant clot lysis activity of 44% was noticed when the clots were treated with 100 µl of CLP extract (10 mg/ml) in in-vitro thrombolytic activity assay. Conclusions: The findings of this study revealed that the peel possessed the antioxidant, anti-inflammatory, antidiabetic, anthelminthic and thrombolytic activities. These findings will be motivated to do further research on molecular level to finds into more depth which is considered as the limitation of this study. Peer Review History: Received 3 February 2024; Revised 12 March 2024; Accepted 20 April; Available online 15 May 2024 Academic Editor: Dr. A.A. Mgbahurike, University of Port Harcourt, Nigeria, amaka_mgbahurike@yahoo.com Average Peer review marks at initial stage: 6.0/10 Average Peer review marks at publication stage: 7.5/10 Reviewers: Dr. Gehan Fawzy Abdel Raoof Kandeel, Pharmacognosy Department, National Research Centre, Dokki, 12622, Giza, Egypt, gehankandeel9@yahoo.com Dr. Francis Adou Yapo, Felix Houphouet Boigny, University of Abidjan, Ivory Coast, fyapo@yahoo.fr