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The Journal of Biological Chemistry Oct 2023Most serine proteases are synthesized as inactive zymogens that are activated by cleavage by another protease in a tightly regulated mechanism. The urokinase-type...
Most serine proteases are synthesized as inactive zymogens that are activated by cleavage by another protease in a tightly regulated mechanism. The urokinase-type plasminogen activator (uPA) and plasmin cleave and activate each other, constituting a positive feedback loop. How this mutual activation cycle begins has remained a mystery. We used hydrogen deuterium exchange mass spectrometry to characterize the dynamic differences between the inactive single-chain uPA (scuPA) and its active form two-chain uPA (tcuPA). The results show that the C-terminal β-barrel and the area around the new N terminus have significantly reduced dynamics in tcuPA as compared with scuPA. We also show that the zymogen scuPA is inactive but can, upon storage, become active in the absence of external proteases. In addition to plasmin, the tcuPA can activate scuPA by cleavage at K158, a process called autoactivation. Unexpectedly, tcuPA can cleave at position 158 even when this site is mutated. TcuPA can also cleave scuPA after K135 or K136 in the disordered linker, which generates the soluble protease domain of uPA. Plasmin cleaves scuPA exclusively after K158 and at a faster rate than tcuPA. We propose a mechanism by which the uPA receptor dimerization could promote autoactivation of scuPA on cell surfaces. These results resolve long-standing controversies in the literature surrounding the mechanism of uPA activation.
PubMed: 37607618
DOI: 10.1016/j.jbc.2023.105179 -
PLoS Pathogens Jul 2023Leptospirosis, a zoonosis with worldwide distribution, is caused by pathogenic spirochetes belonging to the genus Leptospira. Bacterial outer membrane proteins (OMPs),...
Leptospirosis, a zoonosis with worldwide distribution, is caused by pathogenic spirochetes belonging to the genus Leptospira. Bacterial outer membrane proteins (OMPs), particularly those with surface-exposed regions, play crucial roles in pathogen dissemination and virulence mechanisms. Here we characterized the leptospiral Membrane Protein L36 (MPL36), a rare lipoprotein A (RlpA) homolog with a C-terminal Sporulation related (SPOR) domain, as an important virulence factor in pathogenic Leptospira. Our results confirmed that MPL36 is surface exposed and expressed during infection. Using recombinant MPL36 (rMPL36) we also confirmed previous findings of its high plasminogen (PLG)-binding ability determined by lysine residues of the C-terminal region of the protein, with ability to convert bound-PLG to active plasmin. Using Koch's molecular postulates, we determined that a mutant of mpl36 has a reduced PLG-binding ability, leading to a decreased capacity to adhere and translocate MDCK cell monolayers. Using recombinant protein and mutant strains, we determined that the MPL36-bound plasmin (PLA) can degrade fibrinogen. Finally, our mpl36 mutant had a significant attenuated phenotype in the hamster model for acute leptospirosis. Our data indicates that MPL36 is the major PLG binding protein in pathogenic Leptospira, and crucial to the pathogen's ability to attach and interact with host tissues during infection. The MPL36 characterization contributes to the expanding field of bacterial pathogens that explore PLG for their virulence, advancing the goal to close the knowledge gap regarding leptospiral pathogenesis while offering a novel potential candidate to improve diagnostic and prevention of this important zoonotic neglected disease.
Topics: Cricetinae; Animals; Leptospira; Plasminogen; Fibrinolysin; Leptospira interrogans; Protein Binding; Leptospirosis; Bacterial Outer Membrane Proteins; Recombinant Proteins
PubMed: 37486929
DOI: 10.1371/journal.ppat.1011313 -
Blood Advances Nov 2023Thrombosis and bleeding are significant contributors to morbidity and mortality in patients with hematological cancer, and the impact of altered fibrinolysis on bleeding...
Thrombosis and bleeding are significant contributors to morbidity and mortality in patients with hematological cancer, and the impact of altered fibrinolysis on bleeding and thrombosis risk is poorly understood. In this prospective cohort study, we investigated the dynamics of fibrinolysis in patients with hematological cancer. Fibrinolysis was investigated before treatment and 3 months after treatment initiation. A dynamic clot formation and lysis assay was performed beyond the measurement of plasminogen activator inhibitor 1, tissue- and urokinase-type plasminogen activators (tPA and uPA), plasmin-antiplasmin complexes (PAP), α-2-antiplasmin activity, and plasminogen activity. Clot initiation, clot propagation, and clot strength were assessed using rotational thromboelastometry. A total of 79 patients were enrolled. Patients with lymphoma displayed impaired fibrinolysis with prolonged 50% clot lysis time compared with healthy controls (P = .048). They also displayed decreased clot strength at follow-up compared with at diagnosis (P = .001). A patient with amyloid light-chain amyloidosis having overt bleeding at diagnosis displayed hyperfibrinolysis, indicated by a reduced 50% clot lysis time, α-2-antiplasmin activity, and plasminogen activity, and elevated tPA and uPA. A patient with acute promyelocytic leukemia also displayed marked hyperfibrinolysis with very high PAP, indicating extreme plasmin generation, and clot formation was not measurable, probably because of the extremely fast fibrinolysis. Fibrinolysis returned to normal after treatment in both patients. In conclusion, patients with lymphoma showed signs of impaired fibrinolysis and increased clot strength, whereas hyperfibrinolysis was seen in patients with acute promyelocytic leukemia and light-chain amyloidosis. Thus, investigating fibrinolysis in patients with hematological cancer could have diagnostic value.
Topics: Humans; Fibrinolysis; Fibrin Clot Lysis Time; alpha-2-Antiplasmin; Fibrinolysin; Leukemia, Promyelocytic, Acute; Prospective Studies; Lymphoma; Thrombosis; Hematologic Neoplasms; Antifibrinolytic Agents; Urokinase-Type Plasminogen Activator; Amyloidosis; Plasminogen
PubMed: 37756519
DOI: 10.1182/bloodadvances.2023011379 -
Shock (Augusta, Ga.) Sep 2023Excessive blood loss in the prehospital setting poses a significant challenge and is one of the leading causes of death in the United States. In response, emergency... (Review)
Review
Excessive blood loss in the prehospital setting poses a significant challenge and is one of the leading causes of death in the United States. In response, emergency medical services (EMS) have increasingly adopted the use of tranexamic acid (TXA) and calcium chloride (CaCl 2 ) as therapeutic interventions for hemorrhagic traumas. Tranexamic acid functions by inhibiting plasmin formation and restoring hemostatic balance, while calcium plays a pivotal role in the coagulation cascade, facilitating the conversion of factor X to factor Xa and prothrombin to thrombin. Despite the growing utilization of TXA and CaCl 2 in both prehospital and hospital environments, a lack of literature exists regarding the comparative effectiveness of these agents in reducing hemorrhage and improving patient outcomes. Notably, Morgan County Indiana EMS recently integrated the administration of TXA with CaCl 2 into their treatment protocols, offering a valuable opportunity to gather insight and formulate updated guidelines based on patient-centered outcomes. This narrative review aims to comprehensively evaluate the existing evidence concerning the administration of TXA and CaCl 2 in the prehospital management of hemorrhages, while also incorporating and analyzing data derived from the co-administration of these medications within the practices of Morgan County EMS. This represents the inaugural description of the concurrent use of both TXA and CaCl 2 to manage hemorrhages in the scientific literature.
Topics: Humans; Tranexamic Acid; Calcium Chloride; Antifibrinolytic Agents; Hemorrhage; Emergency Medical Services
PubMed: 37477447
DOI: 10.1097/SHK.0000000000002177 -
International Journal of Molecular... Dec 2023Epithelial sodium channel (ENaC) are integral to maintaining salt and water homeostasis in various biological tissues, including the kidney, lung, and colon. They enable... (Review)
Review
Epithelial sodium channel (ENaC) are integral to maintaining salt and water homeostasis in various biological tissues, including the kidney, lung, and colon. They enable the selective reabsorption of sodium ions, which is a process critical for controlling blood pressure, electrolyte balance, and overall fluid volume. ENaC activity is finely controlled through proteolytic activation, a process wherein specific enzymes, or proteases, cleave ENaC subunits, resulting in channel activation and increased sodium reabsorption. This regulatory mechanism plays a pivotal role in adapting sodium transport to different physiological conditions. In this review article, we provide an in-depth exploration of the role of proteolytic activation in regulating ENaC activity. We elucidate the involvement of various proteases, including furin-like convertases, cysteine, and serine proteases, and detail the precise cleavage sites and regulatory mechanisms underlying ENaC activation by these proteases. We also discuss the physiological implications of proteolytic ENaC activation, focusing on its involvement in blood pressure regulation, pulmonary function, and intestinal sodium absorption. Understanding the mechanisms and consequences of ENaC proteolytic activation provides valuable insights into the pathophysiology of various diseases, including hypertension, pulmonary disorders, and various gastrointestinal conditions. Moreover, we discuss the potential therapeutic avenues that emerge from understanding these mechanisms, offering new possibilities for managing diseases associated with ENaC dysfunction. In summary, this review provides a comprehensive discussion of the intricate interplay between proteases and ENaC, emphasizing the significance of proteolytic activation in maintaining sodium and fluid balance in both health and disease.
Topics: Epithelial Sodium Channels; Proteolysis; Serine Endopeptidases; Serine Proteases; Sodium
PubMed: 38139392
DOI: 10.3390/ijms242417563 -
Thrombosis Journal Sep 2023Upon cellular injury, damage-associated molecular patterns (DAMPs) are released into the extracellular space and evoke proinflammatory and prothrombotic responses in...
BACKGROUND
Upon cellular injury, damage-associated molecular patterns (DAMPs) are released into the extracellular space and evoke proinflammatory and prothrombotic responses in animal models of sterile inflammation. However, in clinical settings, the dynamics of DAMP levels after trauma and links between DAMPs and trauma-associated coagulopathy remain largely undetermined.
METHODS
Thirty-one patients with severe trauma, who were transferred to Kagoshima City Hospital between June 2018 and December 2019, were consecutively enrolled in this study. Blood samples were taken at the time of delivery, and 6 and 12 h after the injury, and once daily thereafter. The time-dependent changes of coagulation/fibrinolysis markers, including thrombin-antithrombin complex, α2-plasmin inhibitor (α2-PI), plasmin-α2-PI complex, and plasminogen activator inhibitor-1 (PAI-1), and DAMPs, including high mobility group box 1 and histone H3, were analyzed. The relationship between coagulation/fibrinolysis markers, DAMPs, Injury Severity Score, in-hospital death, and amount of blood transfusion were analyzed.
RESULTS
The activation of coagulation/fibrinolysis pathways was evident at the time of delivery. In contrast, PAI-1 levels remained low at the time of delivery, and then were elevated at 6-12 h after traumatic injury. Histone H3 and high mobility group box 1 levels were elevated at admission, and gradually subsided over time. PAI-1 levels at 6 h were associated with serum histone H3 levels at admission. Increased histone H3 levels and plasmin-α2-PI complex levels were associated with in-hospital mortality. α2-PI levels at admission showed the strongest negative correlation with the amount of blood transfusion.
CONCLUSION
The elevation of histone H3 levels and fibrinolysis perturbation are associated with fatal outcomes in patients with traumatic injury. Patients with low α2-PI levels at admission tend to require blood transfusion.
PubMed: 37674235
DOI: 10.1186/s12959-023-00536-w -
Neuroscience Mar 2024The neurovascular unit (NVU) is assembled by endothelial cells (ECs) and pericytes, and encased by a basement membrane (BM) surveilled by microglia and surrounded by... (Review)
Review
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.
Topics: Humans; Tissue Plasminogen Activator; Fibrinolysin; Ischemic Stroke; Endothelial Cells; Brain Ischemia; Brain; Fibrinolytic Agents
PubMed: 37574107
DOI: 10.1016/j.neuroscience.2023.08.011 -
Acta Neurologica Belgica Apr 2024Alzheimer's disease (AD) is the most common type of dementia associated with amyloid beta (Aβ) deposition. Dysfunction of the neuronal clearance pathway promotes the... (Review)
Review
Alzheimer's disease (AD) is the most common type of dementia associated with amyloid beta (Aβ) deposition. Dysfunction of the neuronal clearance pathway promotes the accumulation of Aβ. The plasminogen-activating system (PAS) is controlled by various enzymes like tissue plasminogen activators (tPA). Neuronal tPA enhances the conversion of plasminogen to plasmin, which cleaves Aβ; this function is controlled by many inhibitors of PAS, including a plasminogen-activating inhibitor (PAI-1) and neuroserpin. Therefore, the objective of the present narrative review was to explore the potential role of tPA/neuroserpin in the pathogenesis of AD. PAI-1 activity is increased in AD, which is involved in accumulating Aβ. Progressive increase of Aβ level during AD neuropathology is correlated with the over-production of PAI-1 with subsequent reduction of plasmin and tPA activities. Reducing plasmin and tPA activities promote Aβ by reducing Aβ clearance. Neuroserpin plays a critical role in the pathogenesis of AD as it regulates the expression and accumulation of Aβ. Higher expression of neuroserpin inhibits the neuroprotective tPA and the generation of plasmin with subsequent reduction in the clearance of Aβ. These observations raise conflicting evidence on whether neuroserpin is neuroprotective or involved in AD progression. Thus, neuroserpin over-expression with subsequent reduction of tPA may propagate AD neuropathology.
Topics: Humans; Alzheimer Disease; Tissue Plasminogen Activator; Amyloid beta-Peptides; Neuroserpin; Fibrinolysin; Plasminogen Activator Inhibitor 1; Plasminogen
PubMed: 37917293
DOI: 10.1007/s13760-023-02403-x -
Kardiologia Polska 2024Patients with transient ST-segment elevation myocardial infarction or spontaneous reperfusion, which occurs in approximately 20% of patients with ST-segment elevation... (Review)
Review
Patients with transient ST-segment elevation myocardial infarction or spontaneous reperfusion, which occurs in approximately 20% of patients with ST-segment elevation myocardial infarction (STEMI), have smaller infarcts and more favorable clinical outcomes than patients without spontaneous reperfusion. Understanding the mechanisms underlying spontaneous reperfusion is therefore important since this may identify possible novel therapeutic targets to improve outcomes in patients with STEMI. In this review, we discuss some of the possible determinants of spontaneous reperfusion including pro-thrombotic profile, endogenous fibrinolytic status, lipoprotein(a) (Lp[a]), inflammatory markers, and neutrophil extracellular traps (NETs). Effective (rapid) endogenous fibrinolysis, as assessed in whole blood in vitro, using a point-of-care technique assessment of global thrombotic status, has been strongly linked to spontaneous reperfusion. Lp(a), which has a high degree of homology to plasminogen, may impair fibrinolysis through competitive inhibition of tissue plasminogen activator-mediated plasminogen activation as well as tissue plasminogen activator-mediated clot lysis and contribute to pathogenic clot properties by decreasing fibrin clot permeation. NETs appear to negatively modulate clot lysis by increasing thrombin fiber diameter and inhibiting plasmin-driven lysis of plasma clots. There are limited data that oral anticoagulation may modulate endogenous fibrinolysis but antiplatelet agents currently appear to have no impact. Phase III trials involving subcutaneous P2Y12 or glycoprotein IIb/IIIa inhibitors, oral factor XIa inhibitors, interleukin-6 inhibitors, and apolipoprotein(a) antisense oligonucleotides in patients with cardiovascular disease are ongoing. Future studies will be needed to determine the impact of these novel antithrombotic, anti-inflammatory, and lipid-lowering therapies on endogenous fibrinolysis and spontaneous reperfusion.
Topics: Humans; ST Elevation Myocardial Infarction; Fibrinolysis; Extracellular Traps; Myocardial Reperfusion; Lipoprotein(a)
PubMed: 38493469
DOI: 10.33963/v.phj.99737 -
CEN Case Reports Nov 2023Infection-related glomerulonephritis (IRGN) is one of the most common causes of acute kidney injury (AKI). Positive glomerular staining of the nephritis-associated...
Infection-related glomerulonephritis (IRGN) is one of the most common causes of acute kidney injury (AKI). Positive glomerular staining of the nephritis-associated plasmin receptor (NAPlr) has been reported as a useful biomarker of IRGN. Although the infection can provoke acute tubulointerstitial nephritis (AIN), there are few reports of positive staining for NAPlr with AIN. We report a case of methicillin-sensitive Staphylococcus aureus (MSSA) infection-related nephritis complicated with AIN, which showed positive staining for tubulointerstitial NAPlr. The patient developed AKI and nephrotic syndrome during an intraperitoneal MSSA infection. A diagnosis of IRGN complicated by infection-related acute tubulointerstitial nephritis (IRAIN) was made based on glomerular endocapillary proliferation with tubulointerstitial infiltrating cells and tubular atrophy. Tubulointerstitial infiltrating cells were positive for NAPlr staining and plasmin activity. Treatment of the infection by antibiotics and drainage did not improve the AKI, but steroid administration improved that. NAPlr evaluation is a helpful tool for identifying causes of AIN during infection.
Topics: Humans; Glomerulonephritis; Glomerulonephritis, IGA; Nephritis; Nephritis, Interstitial; Acute Kidney Injury; Staphylococcal Infections; Immunoglobulin A
PubMed: 36920749
DOI: 10.1007/s13730-023-00782-x