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Background: Hereditary angioedema with normal C1-inhibitor (HAE-nC1-INH) is a rare genetic disease. The symptoms can resemble other forms of hereditary angioedema (HAE), but the specific laboratory values are inconspicuous. The knowledge about treatment strategies in HAE-nC1-INH remains insufficient; most of the drugs are only licensed and approved for other types of HAE. Methods: An analysis of all patients with HAE-nC1-INH was carried out in a certified angioedema treatment center in southern Germany. Only patients with a confirmed HAE-nC1-INH mutation were included. The impact of disease was monitored with validated questionnaires. Results: Eighteen patients were included: two families with a factor XII mutation and seven families with a plasminogen mutation. All individuals received icatibant for on-demand therapy—efficient treatment response was reported. Three patients were severely affected, and prophylaxis was initiated with lanadelumab. According to the questionnaires, the clinical course and symptoms improved significantly under this prophylactic regime. Conclusion: This is one of the first descriptions of the clinical outcomes as a response to prophylactic treatment with lanadelumab in HAE-nC1-INH patients with a known mutation. The therapeutic management of HAE-1 and HAE-2 should also be the basis of HAE-nC1-INH, including prophylaxis. [ABSTRACT FROM AUTHOR]

Group A Streptococcus (GAS) is a human-specific bacterial pathogen that can exploit the plasminogen-plasmin fibrinolysis system to dismantle blood clots and facilitate its spread and survival within the human host. In this study, we use affinity-enrichment mass spectrometry to decipher the host-pathogen protein-protein interaction between plasminogen and streptolysin O, a key cytolytic toxin produced by GAS. This interaction accelerates the conversion of plasminogen to plasmin by both the host tissue-type plasminogen activator and streptokinase, a bacterial plasminogen activator secreted by GAS. Integrative structural mass spectrometry analysis shows that the interaction induces local conformational shifts in plasminogen. These changes lead to the formation of a stabilised intermediate plasminogen-streptolysin O complex that becomes significantly more susceptible to proteolytic processing by plasminogen activators. Our findings reveal a conserved and moonlighting pathomechanistic function for streptolysin O that extends beyond its well-characterised cytolytic activity. Group A Streptococcus exploits the human fibrinolytic system to promote infection. Here, the authors reveal that the bacterial toxin streptolysin O binds to plasminogen and accelerates its conversion to plasmin, thereby dismantling blood clots and facilitating the spread within the human host. [ABSTRACT FROM AUTHOR]

Pulmonary fibrosis, a debilitating lung disorder characterised by excessive fibrous tissue accumulation in lung parenchyma, compromises respiratory function leading to a life‐threatening respiratory failure. While its origins are multifaceted and poorly understood, the urokinase system, including urokinase‐type plasminogen activator (uPA) and its receptor (uPAR), plays a significant role in regulating fibrotic response, extracellular matrix remodelling, and tissue repair. Mesenchymal stem/stromal cells (MSCs) hold promise in regenerative medicine for treating pulmonary fibrosis. Our study aimed to investigate the potential of MSCs to inhibit pulmonary fibrosis as well as the contribution of uPAR expression to this effect. We found that intravenous MSC administration significantly reduced lung fibrosis in the bleomycin‐induced pulmonary fibrosis model in mice as revealed by MRI and histological evaluations. Notably, administering the MSCs isolated from adipose tissue of uPAR knockout mice (Plaur‐/‐ MSCs) attenuated lung fibrosis to a lesser extent as compared to WT MSCs. Collagen deposition, a hallmark of fibrosis, was markedly reduced in lungs treated with WT MSCs versus Plaur‐/‐ MSCs. Along with that, endogenous uPA levels were affected differently; after Plaur‐/‐ MSCs were administered, the uPA content was specifically decreased within the blood vessels. Our findings support the potential of MSC treatment in attenuating pulmonary fibrosis. We provide evidence that the observed anti‐fibrotic effect depends on uPAR expression in MSCs, suggesting that uPAR might counteract the uPA accumulation in lungs. [ABSTRACT FROM AUTHOR]

As an essential thrombolytic agent, the tissue plasminogen activator receives increasing attention due to its longer half-life, lower immunogenicity, and easier administration, which are superior to other thrombolytic agents. In this study, the isolated and purified plasminogen activator from the sandworm (Perinereis aibuhitensis) was expressed in E. coli (Escherichia coli) to investigate its potential for simplifying the development process. The sandworm plasminogen activator was previously successfully cloned and expressed in E. coli with low yield and activity in the culture supernatant. This low yield and activity prompted us to optimize its DNA sequence. Furthermore, to raise the efficiency in the separation of the target protein, the protein's solubility was enhanced by fusing it with maltose-binding protein (MBP) tags. Eventually, the fibrinolytic activity was successfully restored after digestion with tobacco etch virus (TEV) protease. This study provides an innovative method of efficiently expressing and purifying plasminogen activators from sandworm in E. coli and broadens its applications in therapeutic treatment of cardiovascular diseases, including thrombosis, stroke, and coronary atherosclerotic heart disease. [ABSTRACT FROM AUTHOR]

Incident chronic kidney disease (CKD) varies in populations with hypertension of similar severity. Proteinuria promotes CKD progression in part due to activation of plasminogen to plasmin in the podocytes, resulting in oxidative stress‐mediated injury. Additional mechanisms include deficiency of renal alpha‐klotho, that inhibits Wnt/beta‐catenin, an up regulator of intra‐renal renin angiotensin system (RAS) genes. Alpha‐klotho deficiency therefore results in upregulation of the intra‐renal RAS via Wnt/beta‐catenin. In hypertensive, Dahl salt sensitive (DS) and spontaneously hypertensive rats (SHR), we investigated renal and vascular injury, miR‐155, AT1R, alpha‐klotho, and TNF‐α. Hypertensive high salt DS (DS‐HS), but not SHR developed proteinuria, plasminuria, and glomerulosclerosis. Compared to DS low salt (DS‐LS), in hypertensive DS‐HS alpha‐klotho decreased 5‐fold in serum and 2.6‐fold in kidney, whereas serum mir‐155 decreased 3.3‐fold and AT1R increased 52% in kidney and 77% in aorta. AT1R, alpha‐klotho, and miR‐155 remained unchanged in prehypertensive and hypertensive SHR. TNF‐α increased by 3‐fold in serum and urine of DS‐HS rats. These studies unveiled in salt sensitive DS‐HS, but not in SHR, a genetically conditioned dysfunction of the intermolecular network integrated by alpha‐klotho, RAS, miR‐155, and TNF‐α that is at the helm of their end‐organ susceptibility while plasminuria may participate as a second hit. [ABSTRACT FROM AUTHOR]

Background and Aim: The development of acute pancreatitis (AP) is strongly linked to blood clotting and fibrinolysis issues. Modern clinical practices now utilize advanced blood markers like thrombin–antithrombin III complex (TAT), plasmin‐α2‐plasmin inhibitor complex, thrombomodulin (TM), and tissue plasminogen activator–inhibitor complex (t‐PAIC) to assess thrombosis risk. Our study used a highly sensitive chemiluminescence technique to measure these markers in AP patients, aiming to determine their early predictive value for AP severity. Methods: There were 173 patients with AP, all of whom developed symptoms within 72 h; 102 individuals had onset symptoms within 48 h. The biomarkers were measured upon admission before determining the severity of AP. Results: The levels of TAT, plasmin‐α2‐plasmin inhibitor complex, TM, and t‐PAIC were significantly higher in the severe acute pancreatitis (SAP) group compared with the mild acute pancreatitis and moderate severe acute pancreatitis groups. For the patients within 72 h of onset, TAT, TM, and t‐PAIC predicted the occurrence of SAP. For the patients within 48 h of onset, TAT and t‐PAIC predicted the occurrence of SAP. The area under the curve (AUC) of prediction models is similar to Bedside Index for Severity in Acute Pancreatitis (BISAP) but significantly higher than C‐reactive protein (P < 0.05). Notably, t‐PAIC had a larger AUC than TAT, BISAP, and C‐reactive protein. Conclusion: In the initial 48 h, plasma TAT and t‐PAIC levels may predict the development of SAP. Within 72 h, plasma levels of TAT, TM, and t‐PAIC may predict the development of SAP, and the TAT + TM + t‐PAIC prediction model achieved a maximum AUC of 0.915, comparable to BISAP. [ABSTRACT FROM AUTHOR]

Background/Objectives: The negative energy balance activaties of spontaneous lipolysis. This may promotes inflammation within the adipose tissue. The aim of the study was to explain the development of inflammation during increased lactogenesis. It was hypothesized that lipolysis contributes synthesis of interleukin-6 and plasminogen. Methods: The study was in production conditions carried out using Holstein-Friesian cows. The period studied covered time of early lactation. Results: Up to the peak of lactation, milk yield strongly influenced the rate of loss of body condition. This had an impact on with the intensity of the release of the fatty acids. In both cases this relationships strengthened to the peak of production. Oobserved tendencies towards a decrease in the concentration of glucose and an increase in that of leptin. Loss of the body condition and the release of NEFA were were influencing to affect the blood concentrations of interleukin-6 and plasminogen. We have shown that IL-6 has a relatively strong correlation with the NEFA. They correlate with IL-6 independently of EB influence. This may suggest independent associations between these variables, which could potentially be applied in practice. Conclusions: The NEFA release in the long term can increase the inflammatory response within adipose tissue and can intensify the release of interleukin-6 and plasminogen. It is likely that in the initial stage of lactogenesis, the inflammatory process developing within adipose tissue is physiologically justified. Our results can provide background to this little-described area of research. [ABSTRACT FROM AUTHOR]

[Objective] To determine the feasibility of preparing plasminogen (Pg) with Fraction Ⅲ precipitation (hereinafter referred to as FⅢ-P) from low-temperature ethanol process by full chromatography (hereinafter referred to as FⅢ-P process). [Methods] The FⅢ-P was diluted with dissolution buffer at different dilution times and stirring time. The potency and antigen concentration of Pg in dissolution sample were detected and the dissolution and clarification conditions were determined. Pre-treatment of loading sample and pre-experiment of affinity chromatography were carried out on the FⅢ-P dissolution sample to judge whether the loading sample had an impact on the chromatography by observing the performance of the affinity chromatography column and to evaluate whether the affinity chromatography could achieve the purpose of purifying Pg by detecting the plasma protein antigen concentration and Pg potency of the samples in the process. Two batches of FⅢ-P process were studied step by step, and the specific activity, steps and total recovery, and the output of Pg per ton of plasma were calculated. The feasibility of preparing Pg by FⅢ-P process was evaluated by comparing with the data of full chromatography process using plasma as raw material (hereinafter referred to as plasma process). [Results] The FⅢ-P was dissolved with 10 times of dissolution buffer, stirred for 1 hour, centrifuged at room temperature of 10 000×g for 15 minutes. The supernatant was first filtered with a screen, then clarified with an 8/0.8 μm filter, and finally filtered with a 0.45/0.2 μm filter and loaded. Pre-test showed that from clarification and filtration to Pg affinity chromatography, the step recovery of activity and antigen was 39.51% and 108.64%, respectively, the antigen concentration of Pg increased by 31.16 times and the activity increased by 11.39 times after affinity chromatography, which reaching the effect of affinity chromatography purification of Pg. The results of 2 batches of step-by-step scale-up FⅢ-P process showed that the total recoveries of antigen and activity from plasma to SP chromatography of FⅢ-P process were (45.76±1.10)% and (24.15±0.59)%, respectively, which had a total loss of about 1/3 of antigen and about 2/3 of activity compared to the plasma process. The Pg specific activity of SP chromatography eluent was (4.68±0.25) U/mg, which was about half of that of plasma process, but meeted the internal standard of > 4 U/mg. The output of Pg antigen per ton of plasma in the FⅢ-P process was 68.73% of that in the plasma process, and the output of Pg activity per ton of plasma in the plasma process was 29.82% of that in the plasma process, which basically achieved the purpose of waste utilization of FⅢ-P. [Conclusion] The technical route of preparing Pg from FⅢ-P by full chromatography is feasible.

The study of reciprocal interactions between the plasminogen/plasmin system and the platelet componentof hemostasis is necessary both for understanding the biochemical mechanisms regulating the processes of thrombosis and thrombolysis and for elucidating the role of platelets in angiogenesis. Aim. The study aimed to investigate the peculiarities of plasminogen processing by cytosolic and plasma membrane-associated proteases of platelets. Methods. Gel-permeation filtration was used for the isolation of platelets from the donor’s blood plasma. Plasminogen was purified from Cohn’s fraction III2,3 of human blood plasma by affinity chromatography on lysine-Sepharose. The viability of washed platelets and their response to an agonist were assessed by optical aggregometry. The processing of plasminogen on platelets was induced by stimulating the cells with thrombin (1 NIH/ml) after pre-incubation with 0.25 μM Pg for 30, 60, or 120 min. Plasminogen and its fragments were detected by immunoblot with the use of previously obtained polyclonal antibodies to plasminogen kringles (K1-3 and K5). Results. It was established that exogenous plasminogen is adsorbed onto the plasma membrane of platelets, converted into the Lys-form, and further fragmented into angiostatins and mini-plasminogen. This indicates the involvement of various platelet proteases in plasminogen cleavage. It was shown that platelets are capable of internalizing exogenous plasminogen in its Glu-form, while formed angiostatins are not internalized by the cells. It has been determined that internalized Glu-plasminogen (0.25 μM) may change its conformation to a Lys-like form within  120 minutes of incubation with platelets, as immunochemically detected with the use of antibodies against K5 plasminogen fragment. Conclusion. The obtained results provide new insights into the mechanisms by which platelets may regulate the functioning of the plasminogen/plasmin system. This regulation occurs through their ability to generate plasminogen fragments (angiostatins) and having the potential for internalization and further secretion of the formed angiostatins by both native and activated platelets.

Venous thrombosis and pulmonary embolism (venous thromboembolism) are important causes of morbidity and mortality worldwide. In patients with venous thromboembolism, thrombi obstruct blood vessels and resist physiological dissolution (fibrinolysis), which can be life threatening and cause chronic complications. Plasminogen activator therapy, which was developed >50 years ago, is effective in dissolving thrombi but has unacceptable bleeding risks. Safe dissolution of thrombi in patients with venous thromboembolism has been elusive despite multiple innovations in plasminogen activator design and catheter-based therapy. Evidence now suggests that fibrinolysis is rigidly controlled by endogenous fibrinolysis inhibitors, including α2-antiplasmin, plasminogen activator inhibitor-1, and thrombin-activable fibrinolysis inhibitor. Elevated levels of these fibrinolysis inhibitors are associated with an increased risk of venous thromboembolism in humans. New therapeutic paradigms suggest that accelerated and effective fibrinolysis may be achieved safely by therapeutically targeting these fibrinolytic inhibitors in venous thromboembolism. In this article, we discuss the role of fibrinolytic components in venous thromboembolism and the current status of research and development targeting fibrinolysis inhibitors. [ABSTRACT FROM AUTHOR]

The study of reciprocal interactions between the plasminogen/plasmin system and the platelet component of hemostasis is necessary both for understanding the biochemical mechanisms regulating the processes of thrombosis and thrombolysis and for elucidating the role of platelets in angiogenesis. Aim. The study aimed to investigate the peculiarities of plasminogen processing by cytosolic and plasma membrane-associated proteases of platelets. Methods. Gel-permeation filtration was used for the isolation of platelets from the donor’s blood plasma. Plasminogen was purified from Cohn’s fraction III2,3 of human blood plasma by affinity chromatography on lysine-Sepharose. The viability of washed platelets and their response to an agonist were assessed by optical aggregometry. The processing of plasminogen on platelets was induced by stimulating the cells with thrombin (1 NIH/ml) after pre-incubation with 0.25 μM Pg for 30, 60, or 120 min. Plasminogen and its fragments were detected by immunoblot with the use of previously obtained polyclonal antibodies to plasminogen kringles (K1-3 and K5). Results. It was established that exogenous plasminogen is adsorbed onto the plasma membrane of platelets, converted into the Lys-form, and further fragmented into angiostatins and mini-plasminogen. This indicates the involvement of various platelet proteases in plasminogen cleavage. It was shown that platelets are capable of internalizing exogenous plasminogen in its Glu-form, while formed angiostatins are not internalized by the cells. It has been determined that internalized Glu-plasminogen (0.25 μM) may change its conformation to a Lys-like form within ≥ 120 minutes of incubation with platelets, as immunochemically detected with the use of antibodies against K5 plasminogen fragment. Conclusion. The obtained results provide new insights into the mechanisms by which platelets may regulate the functioning of the plasminogen/plasmin system. This regulation occurs through their ability to generate plasminogen fragments (angiostatins) and having the potential for internalization and further secretion of the formed angiostatins by both native and activated platelets. [ABSTRACT FROM AUTHOR]

Fibrinolysis refers to plasmin-mediated clot breakdown, which can be exaggerated in certain disease states, leading to severe bleeding. Antifibrinolytic drugs, including tranexamic acid and epsilon-aminocaproic acid, are a useful addition to the toolbox for the treatment of bleeding in small animals. These drugs are lysine analogs, which prevent binding of plasminogen to fibrin-binding sites on blood clots, limiting the conversion of plasminogen to plasmin and therefore preventing clot degradation. Common situations to consider antifibrinolytic drugs include in traumatic and surgical hemorrhage (e.g., spontaneous hemoperitoneum), in perioperative bleeding in certain breeds of dog (e.g., greyhounds), or as an adjunctive hemostatic agent in coagulopathies (e.g., hemophilia A). These drugs are well tolerated in most dogs, although patient selection is important to avoid development of inadvertent issues (e.g., obstructive blood clot in cases of urinary tract bleeding). Less information about the use of antifibrinolytic drugs in cats is available, although these drugs might play a role similar to their use in dogs. [ABSTRACT FROM AUTHOR]

Dislocation of anti‐adhesion materials, non‐specific tissue adhesion, and the induction of secondary fibrinolysis disorders are the main challenges faced by postoperative anti‐adhesion materials. Herein, a self‐leveling transient unilateral adhesive hydrogel is custom‐designed to conquer these challenges with a theoretically calculated and dual‐step tailored gellan gum (GG) as the sole agent. First, the maximum gelation temperature of GG is lowered from 42–25 °C through controlled perturbation of intra‐ and inter‐molecular hydrogen bonds, which is achieved by employing the methacrylic anhydride as a "hydrogen bond's perturbator" to form methacrylate GG (MeGG). Second, the "self‐leveling" injectability and wound shape adaptably are endowed by the formation of borate‐diol complexed MeGG (BMeGG). Finally, the transient unilateral tissue‐adhesive hydrogel (BMeGG‐H) barrier is prepared through photo‐controlled cross‐linking of reactive alkenyl groups. This degradable hydrogel demonstrates favorable rheological properties, light‐controlled unilateral adhesion properties, biocompatibility, anti‐fibrin adhesion, and anti‐cell adhesion properties in vitro. Comprehensive regulation of the fibrinolysis balance toward non‐adhesion is conformed in a rat model after intra‐abdominal surgery via anti‐autoinflammatory response, intestinal wall integrity repair, and Tissue plasminogen activator (t‐PA) and plasminogen activator inhibitor‐1 (PAI‐1) balance adjustment. Notably, the 14th day anti‐adhesion effective rate is 100%, indicating its significant potential in clinical applications for postoperative anti‐adhesion. [ABSTRACT FROM AUTHOR]

The association between high-density lipoprotein (HDL) cholesterol and breast cancer (BC) remains controversial due to the high complexity of the HDL particle and its functionality. The HDL proteome was determined in newly diagnosed BC classified according to the molecular type [luminal A or B (LA or LB), HER2, and triple-negative (TN)] and clinical stage of the disease. Women (n = 141) aged between 18 and 80 years with BC, treatment-naïve, and healthy women [n = 103; control group (CT)], matched by age and body mass index, were included. Data-independent acquisition (DIA) proteomics was performed in isolated HDL (D = 1.063–1.21 g/mL). Results: Paraoxonase1, carnosine dipeptidase1, immunoglobulin mMu heavy chain constant region (IGHM), apoA-4, and transthyretin were reduced, and serum amyloid A2 and tetranectin were higher in BC compared to CT. In TNBC, apoA-1, apoA-2, apoC-2, and apoC-4 were reduced compared to LA, LB, and HER2, and apoA-4 compared to LA and HER2. ComplementC3, lambda immunoglobulin2/3, serpin3, IGHM, complement9, alpha2 lysine rich-glycoprotein1, and complement4B were higher in TNBC in comparison to all other types; complement factor B and vitamin D-binding protein were in contrast to LA and HER2, and plasminogen compared to LA and LB. In grouped stages III + IV, tetranectin and alpha2-macroglobulin were reduced, and haptoglobin-related protein; lecithin cholesterol acyltransferase, serum amyloid A1, and IGHM were increased compared to stages I + II. Conclusions: A differential proteomic profile of HDL in BC based on tumor molecular classification and the clinical stage of the disease may contribute to a better understanding of the association of HDL with BC pathophysiology, treatment, and outcomes. [ABSTRACT FROM AUTHOR]

Pleural effusion is the most common manifestation of pleural disease, and chest ultrasound is crucial for diagnostic workup and post-treatment monitoring. Ultrasound helps distinguish the various types of pleural effusion and enables the detection of typical manifestations of empyema, which presents as a complicated, septated effusion. This may benefit from drainage and the use of intrapleural enzyme therapy or may require more invasive approaches, such as medical or surgical thoracoscopy. The mechanism of action of intrapleural enzymatic therapy (IPET) is the activation of plasminogen to plasmin, which breaks down fibrin clots that form septa or the loculation of effusions and promotes their removal. In addition, IPET has anti-inflammatory properties and can modulate the immune response in the pleural space, resulting in reduced pleural inflammation and improved fluid reabsorption. In this article, we briefly review the literature on the efficacy of IPET and describe a case series in which most practical applications of IPET are demonstrated, i.e., as a curative treatment but also as an alternative, propaedeutic, or subsequent treatment to surgery. [ABSTRACT FROM AUTHOR]

Objectives Materials and Methods Results Conclusions This study investigated the role of fibrin on neutrophil extracellular traps (NETs) formation from neutrophils and to elucidate the involvement of mitochondria in NETs formation during periodontitis.Plasminogen‐deficient (Plg−/−) mice were employed to evaluate the effects of fibrin deposition on inflammation, bone resorption, and neutrophil infiltration in periodontal tissues. In addition, in vitro tests evaluated fibrin's impact on neutrophil‐driven inflammation. Mitochondrial reactive oxygen species (mtROS) levels within neutrophils were quantified utilizing flow cytometry and immunofluorescence in vitro. Furthermore, the anti‐inflammatory properties of the mtROS scavenger, Mito‐TEMPO, were confirmed to regulate the NET formation in vitro and in vivo.Plasminogen deficiency resulted in increased fibrin deposition, neutrophil infiltration, inflammatory factors concentration, and alveolar bone resorption in periodontal tissues. After neutrophils were treated by fibrin in vitro, the expression of inflammatory factors, the formation of mtROS, and NETs enriched in mitochondrial DNA (mtDNA) were upregulated, which were reversed by Mito‐TEMPO in vitro. Moreover, Mito‐TEMPO alleviated inflammation in Plg−/− mice.This study showed that fibrin deposition in gingiva induced the NET formation in Plg−/− mice, in which the DNA in NETs was from mitochondria depending on increasing mtROS. [ABSTRACT FROM AUTHOR]

The pathway for axon regeneration in Caenorhabditis elegans is activated by SVH-1, a growth factor belonging to the HGF/plasminogen family. SVH-1 is a dual-function factor that acts as an HGF-like growth factor to promote axon regeneration and as a protease to regulate early development. It is important to understand how SVH-1 is converted from a protease to a growth factor for axon regeneration. In this study, we demonstrate that cytidine deaminase (CDD) SVH-17/CDD-2 plays a role in the functional conversion of SVH-1. We find that the codon exchange of His-755 to Tyr in the Asp–His–Ser catalytic triad of SVH-1 can suppress the cdd-2 defect in axon regeneration. Furthermore, the stem hairpin structure around the His-755 site in svh-1 mRNA is required for the activation of axon regeneration by SVH-1. These results suggest that CDD-2 promotes axon regeneration by transforming the function of SVH-1 from a protease to a growth factor through modification of svh-1 mRNA. Author summary: The axon regeneration pathway in C. elegans is activated by SVH-1, a growth factor that belongs to the HGF/plasminogen family. SVH-1 is a dual-functional factor that promotes axon regeneration as an HGF-like growth factor and regulates development as a plasminogen-like protease. It is crucial to understand the mechanism by which SVH-1 transforms from a protease to a growth factor during axon regeneration. In this study, we demonstrate that CDD-2, a cytidine deaminase, is critical for converting SVH-1 function from a protease to a growth factor by modifying svh-1 mRNA. [ABSTRACT FROM AUTHOR]

Objectives Materials and Methods Results Conclusion Periodontitis is a common oral disease that is aggravated by occlusal trauma. Fibrin is a protein that participates in blood clotting and is involved in several human diseases. The deposition of fibrin in periodontal tissues can induce periodontitis, while mechanical forces may regulate the degradation of fibrin. Our study investigated how occlusal trauma aggravating periodontitis through regulating the plasminogen/plasmin system and fibrin deposition.This study included 84 C57BL/6 mice in which periodontitis was induced with or without occlusal trauma. Micro‐computed tomography was used to assess bone resorption. Fibrin, fibrinogen, plasminogen, plasmin, tissue plasminogen activator (t‐PA), and urokinase plasminogen activator (u‐PA) levels were measured using Frazer–Lendrum staining, quantitative reverse transcription polymerase chain reaction, enzyme‐linked immunosorbent assay, western blotting, immunofluorescence staining, and immunohistochemistry staining.Occlusal trauma aggravated inflammation and bone resorption. The periodontitis group showed significant fibrin deposition. Occlusal trauma increased fibrin deposition and neutrophil aggregation. The periodontitis with occlusal trauma group had decreased fibrinogen, t‐PA, and u‐PA expression and plasmin and fibrin degradation product levels, as well as increased plasminogen levels.Occlusal trauma promotes excessive fibrin deposition by suppressing the plasminogen/plasmin system, thus exacerbating periodontitis. [ABSTRACT FROM AUTHOR]

• Tissue-type plasminogen activator (tPA) is a serine proteinase that catalyzes the conversion of plasminogen into plasmin. • The neurovascular unit is assembled by endothelial cells, pericytes, the perivascular basement membrane, astrocytes, neurons and microglia. • TPA is abundantly the cellular components of the NVU where it regulates its function under physiologic and pathologic conditions. The neurovascular unit (NVU) is assembled by endothelial cells (ECs) and pericytes, and encased by a basement membrane (BM) surveilled by microglia and surrounded by perivascular astrocytes (PVA), which in turn are in contact with synapses. Cerebral ischemia induces the rapid release of the serine proteinase tissue-type plasminogen activator (tPA) from endothelial cells, perivascular astrocytes, microglia and neurons. Owning to its ability to catalyze the conversion of plasminogen into plasmin, in the intravascular space tPA functions as a fibrinolytic enzyme. In contrast, the release of astrocytic, microglial and neuronal tPA have a plethora of effects that not always require the generation of plasmin. In the ischemic brain tPA increases the permeability of the NVU, induces microglial activation, participates in the recycling of glutamate, and has various effects on neuronal survival. These effects are mediated by different receptors, notably subunits of the N-methyl-D-aspartate receptor (NMDAR) and the low-density lipoprotein receptor-related protein-1 (LRP-1). Here we review data on the role of tPA in the NVU under non-ischemic and ischemic conditions, and analyze how this knowledge may lead to the development of potential strategies for the treatment of acute ischemic stroke patients. [ABSTRACT FROM AUTHOR]

Background: There is a need for a simpler and accessible intervention to heal tympanic membrane perforations than myringoplasty that is todaýs golden standard. Experimental studies have identified plasminogen as a promising agent for medical treatment of chronic tympanic membrane perforation. Aims/Objectives: This was a phase 1, prospective, randomized, placebo-controlled study with the main objective to evaluate the safety of injecting plasminogen in the vicinity of the tympanic membrane in subjects with chronic tympanic membrane perforation. Material and Methods: Adults diagnosed with a dry chronic tympanic membrane perforation were recruited for an injection schedule with Human plasminogen 10. Adverse events, audiometry, VAS fluctuations and size of perforation, were monitored throughout the length of the study. Results: It was possible to perform the injections according to schedule in all subjects. None of the subjects experienced any severe adverse events. Most common adverse event was ear pain. No signs of ototoxicity were reported. Conclusions and Significance: This pilot study suggests that plasminogen injections close to the tympanic membrane as treatment for chronic tympanic membrane injections are safe and feasible, encouraging further dose-escalating studies. [ABSTRACT FROM AUTHOR]

Background: Thromboembolism, which leads to pulmonary embolism and ischemic stroke, remains one of the main causes of death. Ultrasound‐assisted thrombolysis (UAT) is an effective thrombolytic method. However, further studies are required to elucidate the mechanism of ultrasound on arterial and venous thrombi. Methods: We employed the blood‐on‐a‐chip technology to simulate thrombus formation in coronary stenosis and deep vein valves. Subsequently, UAT was conducted on the chip to assess the impact of ultrasound on thrombolysis under varying flow conditions. Real‐time fluorescence was used to assess thrombolysis and drug penetration. Finally, scanning electron microscopy and immunofluorescence were used to determine the effect of ultrasound on fibrinolysis. Results: The study revealed that UAT enhanced the thrombolytic rate by 40% in the coronary stenosis chip and by 10% in the deep venous valves chip. This enhancement is attributed to the disruption of crosslinked fibrin fibers by ultrasound, leading to increased urokinase diffusion within the thrombus and accumulation of plasminogen on the fibrinogen α chain. Moreover, the acceleration of the dissolution rate of thrombi in the venous valve chip by ultrasound was not as significant as that in the coronary stenosis chip. Conclusion: These findings highlight the differential impact of ultrasound on thrombolysis under various flow conditions and emphasize the valuable role of the blood‐on‐a‐chip technology in exploring thrombolysis mechanisms. [ABSTRACT FROM AUTHOR]

Deposition of extracellular matrix (ECM) in the trabecular meshwork (TM) increases aqueous humour outflow resistance leading to elevation of intraocular pressure (IOP) in primary open-angle glaucoma, which remains the only modifiable risk factor. Resveratrol has been shown to counteract the steroid-induced increase in IOP and increase the TM expression of ECM proteolytic enzymes; however, its effects on the deposition of ECM components by TM and its associated pathways, such as TGF-ß-SMAD signalling remain uncertain. This study, therefore, explored the effects of trans-resveratrol on the expression of ECM components, SMAD signalling molecules, plasminogen activator inhibitor-1 and tissue plasminogen activator in dexamethasone-treated human TM cells (HTMCs). We also studied the nature of molecular interaction of trans -resveratrol with SMAD4 domains using ensemble docking. Treatment of HTMCs with 12.5 µM trans-resveratrol downregulated the dexamethasone-induced increase in collagen, fibronectin and a-smooth muscle actin at gene and protein levels through downregulation of TGF-ß1, SMAD4, and upregulation of SMAD7. Downregulation of TGF-ß1 signalling by trans-resveratrol could be attributed to its effect on the transcriptional activity due to high affinity for the MH2 domain of SMAD4. These effects may contribute to resveratrol's IOP-lowering properties by reducing ECM deposition and enhancing aqueous humour outflow in the TM. [ABSTRACT FROM AUTHOR]

Introduction: Plasminogen deficiency is an ultra rare disease whose patients may develop ligneous lesions if untreated. Prophylactic replacement therapy with plasma derived plasminogen, Ryplazim, is efficient in treating lesions and could benefit from pharmacokinetic (PK) tailoring. Aim: The objectives of this study are to develop, evaluate and integrate into the WAPPS‐Hemo platform a Population PK model supporting prophylactic replacement therapy for Plasminogen deficient patients. Methods: Population PK modelling and evaluations followed the same protocol performed for factor VIII and IX concentrates. Limited sampling analysis used dosing and sampling scenarios in accordance with recommended treatment for Ryplazim. Results: The population PK model, derived from 16 participants included in previous clinical studies, was a 2‐compartment model whose variability was best described by fat‐free mass. Evaluations showed that the model described well the data and Bayesian forecasting in limited sampling environment led to acceptable precision for PK parameters relevant to plasminogen treatment. Conclusion: The model was integrated into the WAPPS‐Hemo webservice to help individualize prophylactic treatment in plasminogen deficient patients. Prospective PK data to be collected through the WAPPS‐Hemo database will be used to better understand plasminogen PK and improve patient care. [ABSTRACT FROM AUTHOR]

The worldwide burden of skeletal diseases such as osteoporosis, degenerative joint disease and impaired fracture healing is steadily increasing. Tranexamic acid (TXA), a plasminogen inhibitor and anti-fibrinolytic agent, is used to reduce bleeding with high effectiveness and safety in major surgical procedures. With its widespread clinical application, the effects of TXA beyond anti-fibrinolysis have been noticed and prompted renewed interest in its use. Some clinical trials have characterized the effects of TXA on reducing postoperative infection rates and regulating immune responses in patients undergoing surgery. Also, several animal studies suggest potential therapeutic effects of TXA on skeletal diseases such as osteoporosis and fracture healing. Although a direct effect of TXA on the differentiation and function of bone cells in vitro was shown, few mechanisms of action have been reported. Here, we summarize recent findings of the effects of TXA on skeletal diseases and discuss the underlying plasminogen-dependent and -independent mechanisms related to bone metabolism and the immune response. We furthermore discuss potential novel indications for TXA application as a treatment strategy for skeletal diseases. Tranexamic acid serves as a potential candidate in the treatment of skeletal diseases through plasminogen-dependent and -independent processes in relation to bone metabolism, immune system, and inflammatory responses. [Display omitted] [ABSTRACT FROM AUTHOR]

Objective: This study aimed to identify the binding sites for plasminogen (Pg) and its kringle-containing fragments within the αC-region of fibrin(ogen). This investigation is crucial while the conversion of fibrinogen into fibrin induces conformational changes that expose binding sites for Pg and tissue-type Pg activator (tPA), facilitating effective zymogen activation on the fibrin surface. Methods: Two C-terminal fragments of the Aα chain ‒ 45 kDa (225Val-610Val) and 40 kDa (225Val-580Lys), were obtained through plasmin hydrolysis of human fibrinogen and subsequently characterized using MALDI TOF mass spectrometry. The interactions of Glu-Pg and Lys-Pg, as well as Pg kringle fragments (K1-3, K4, and K5), with the obtained αC truncated polypeptides were analyzed using ELISA and Western blot techniques with the use of specific antibodies. Results: It was demonstrated that Pg and its fragments K1-3, K4, and K5 interact exclusively with the 45-kDa fragment (225Val-610Val) of the αC region of fibrinogen with high affinity in a concentration-dependent manner (Kd values for Glu-Pg = 7.10 × 10−9 M, Lys-Pg = 6.01 × 10−9 M, K1-3 = 1.08 × 10−7 M, K4 = 5.06 × 10−7 M, and K5 = 2.50 × 10−7 M). This fragment, unlike the 40-kDa polypeptide (225Val-580Lys), contains the α581Ser-610Val sequence. Conclusions: It was shown that the sequence 581Ser-610Val of fibrinogen Aα-chain, which becomes exposed during the conversion of fibrinogen to fibrin, is essential for the formation of complexes between Pg and αC regions of fibrin(ogen), thereby contributing to the initiation and regulation of fibrinolysis.

This study aimed to evaluate the clot-lysing and membrane stabilizing capacities of ascorbic acid (AA) using in vitro and in silico methods. For this, we used in vitro clot lysis and hemolyzing tests to check the anti-atherothrombosis and membrane-stabilizing properties of AA, respectively. Additionally, molecular docking studies were performed to investigate AA’s interactions with cyclooxygenase-1 (COX-1) and plasminogen enzymes. Findings suggest that AA exhibited a concentration-dependent effect, with 43.95 ± 1.27 % clot lysis and 64.46 ± 0.01 % membrane stabilization at 100 µg/mL. The IC50 values for clot lysis and membrane stabilization were 215.19 ± 1.09 and 57.21 ± 2.11 µg/mL, respectively. In silico analysis showed strong binding affinities of AA with COX-1 (−6.2 kcal/mol) and plasminogen (−5.8 kcal/mol), supporting its observed clot lysis and membrane protection activities. Taken together, AA showed moderate clot-lysing and robust membrane-stabilizing effects, which may be due to its strong antioxidant and anti-inflammatory properties. AA might be a good therapeutic agent for atherothrombosis and membrane damage, highlighting the need for further investigation into its underlying molecular mechanisms and potential clinical applications. AA shows promising clot-lysing and membrane-stabilizing effects, highlighting its therapeutic potential for atherothrombosis and membrane damage.

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.

Abstract ADAMTS13, a disintegrin and metalloprotease with a thrombospondin type 1 motif, member 13, regulates the length of Von Willebrand factor (VWF) multimers and their platelet-binding activity. ADAMTS13 is constitutively secreted as an active protease and is not inhibited by circulating protease inhibitors. Therefore, the mechanisms that regulate ADAMTS13 protease activity are unknown. We performed an unbiased proteomics screen to identify ligands of ADAMTS13 by optimizing the application of BioID to plasma. Plasma BioID identified 5 plasma proteins significantly labeled by the ADAMTS13-birA* fusion, including VWF and plasminogen. Glu-plasminogen, Lys-plasminogen, mini-plasminogen, and apo(a) bound ADAMTS13 with high affinity, whereas micro-plasminogen did not. None of the plasminogen variants or apo(a) bound to a C-terminal truncation variant of ADAMTS13 (MDTCS). The binding of plasminogen to ADAMTS13 was attenuated by tranexamic acid or ε-aminocaproic acid, and tranexamic acid protected ADAMTS13 from plasmin degradation. These data demonstrate that plasminogen is an important ligand of ADAMTS13 in plasma by binding to the C-terminus of ADAMTS13. Plasmin proteolytically degrades ADAMTS13 in a lysine-dependent manner, which may contribute to its regulation. Adapting BioID to identify protein-interaction networks in plasma provides a powerful new tool to study protease regulation in the cardiovascular system.

Abstract Parkinson's disease (PD) is the second most frequently diagnosed neurodegenerative disease, and it is characterized by the intracellular and extracellular accumulation of α-synuclein (α-syn) and Tau, which are major components of cytosolic protein inclusions called Lewy bodies, in the brain. Currently, there is a lack of effective methods that preventing PD progression. It has been suggested that the plasminogen activation system, which is a major extracellular proteolysis system, is involved in PD pathogenesis. We investigated the functional roles of plasminogen in vitro in an okadaic acid-induced Tau hyperphosphorylation NSC34 cell model, ex vivo using brains from normal controls and methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice, and in vivo in a widely used MPTP-induced PD mouse model and an α-syn overexpression mouse model. The in vitro, ex vivo and in vivo results showed that the administered plasminogen crossed the blood‒brain barrier (BBB), entered cells, and migrated to the nucleus, increased plasmin activity intracellularly, bound to α-syn through lysine binding sites, significantly promoted α-syn, Tau and TDP-43 clearance intracellularly and even intranuclearly in the brain, decreased dopaminergic neurodegeneration and increased the tyrosine hydroxylase levels in the substantia nigra and striatum, and improved motor function in PD mouse models. These findings indicate that plasminogen plays a wide range of pivotal protective roles in PD and therefore may be a promising drug candidate for PD treatment.

Intravenous thrombolysis with recombinant tissue plasminogen activator (rtPA) is the primary treatment for ischemic stroke. However, rtPA treatment can substantially increase blood‐brain barrier (BBB) permeability and susceptibility to hemorrhagic transformation. Herein, the mechanism underlying the side effects of rtPA treatment is investigated and demonstrated that ferroptosis plays an important role. The ferroptosis inhibitor, liproxstatin‐1 (Lip) is proposed to alleviate the side effects. A well‐designed macrocyclic carrier, glucose‐modified azocalix[4]arene (GluAC4A), is prepared to deliver Lip to the ischemic site. GluAC4A bound tightly to Lip and markedly improved its solubility. Glucose, modified at the upper rim of GluAC4A, imparts BBB targeting to the drug delivery system owing to the presence of glucose transporter 1 on the BBB surface. The responsiveness of GluAC4A to hypoxia due to the presence of azo groups enabled the targeted release of Lip at the ischemic site. GluAC4A successfully improved drug accumulation in the brain, and Lip@GluAC4A significantly reduced ferroptosis, BBB leakage, and neurological deficits induced by rtPA in vivo. These findings deepen the understanding of the side effects of rtPA treatment and provide a novel strategy for their effective mitigation, which is of great significance for the treatment and prognosis of patients with ischemic stroke. [ABSTRACT FROM AUTHOR]

Intravenous thrombolysis with a recombinant tissue plasminogen activator (rt-PA) is the first-line treatment of acute ischemic stroke. However, successful recanalization is relatively low and the underlying processes are not completely understood. The goal was to provide insights into clinically important factors potentially limiting rt-PA efficacy such as clot size, rt-PA concentration, clot age and also rt-PA in combination with heparin anticoagulant. We established a static in vitro thrombolytic model based on red blood cell (RBC) dominant clots prepared using spontaneous clotting from the blood of healthy donors. Thrombolysis was determined by clot mass loss and by RBC release. The rt-PA became increasingly less efficient for clots larger than 50 μl at a clinically relevant concentration of 1.3 mg/l. A tenfold decrease or increase in concentration induced only a 2-fold decrease or increase in clot degradation. Clot age did not affect rt-PA-induced thrombolysis but 2-hours-old clots were degraded more readily due to higher activity of spontaneous thrombolysis, as compared to 5-hours-old clots. Finally, heparin (50 and 100 IU/ml) did not influence the rt-PA-induced thrombolysis. Our study provided in vitro evidence for a clot size threshold: clots larger than 50 μl are hard to degrade by rt-PA. Increasing rt-PA concentration provided limited thrombolytic efficacy improvement, whereas heparin addition had no effect. However, the higher susceptibility of younger clots to thrombolysis may prompt a shortened time from the onset of stroke to rt-PA treatment. [ABSTRACT FROM AUTHOR]