Plasmapheresis to remove amyloid fibrin(ogen) particles for treating the post-COVID-19 condition.

BACKGROUND: The post-COVID-19 condition (PCC) consists of a wide array of symptoms including fatigue and impaired daily living. People seek a wide variety of approaches to help them recover. A new belief, arising from a few laboratory studies, is that 'microclots' cause the symptoms of PCC. This belief has been extended outside these studies, suggesting that to recover people need plasmapheresis (an expensive process where blood is filtered outside the body). We appraised the laboratory studies, and it was clear that the term 'microclots' is incorrect to describe the phenomenon being described. The particles are amyloid and include fibrin(ogen); amyloid is not a part of a thrombus which is a mix of fibrin mesh and platelets. Initial acute COVID-19 infection is associated with clotting abnormalities; this review concerns amyloid fibrin(ogen) particles in PCC only. We have reported here our appraisal of laboratory studies investigating the presence of amyloid fibrin(ogen) particles in PCC, and of evidence that plasmapheresis may be an effective therapy to remove amyloid fibrin(ogen) particles for treating PCC. OBJECTIVES: Laboratory studies review To summarize and appraise the research reports on amyloid fibrin(ogen) particles related to PCC. Randomized controlled trials review To assess the evidence of the safety and efficacy of plasmapheresis to remove amyloid fibrin(ogen) particles in individuals with PCC from randomized controlled trials. SEARCH METHODS: Laboratory studies review We searched for all relevant laboratory studies up to 27 October 2022 using a comprehensive search strategy which included the search terms 'COVID', 'amyloid', 'fibrin', 'fibrinogen'. Randomized controlled trials review We searched the following databases on 21 October 2022: Cochrane COVID-19 Study Register; MEDLINE (Ovid); Embase (Ovid); and BIOSIS Previews (Web of Science). We also searched the WHO International Clinical Trials Registry Platform and ClinicalTrials.gov for trials in progress. SELECTION CRITERIA: Laboratory studies review Laboratory studies that investigate the presence of amyloid fibrin(ogen) particles in plasma samples from patients with PCC were eligible. This included studies with or without controls. Randomized controlled trials review Studies were eligible if they were of randomized controlled design and investigated the effectiveness or safety of plasmapheresis for removing amyloid fibrin(ogen) particles for treating PCC. DATA COLLECTION AND ANALYSIS: Two review authors applied study inclusion criteria to identify eligible studies and extracted data. Laboratory studies review We assessed the risk of bias of included studies using pre-developed methods for laboratory studies. We planned to perform synthesis without meta-analysis (SWiM) as described in our protocol. Randomized controlled trials review We planned that if we identified any eligible studies, we would assess risk of bias and report results with 95% confidence intervals. The primary outcome was recovery, measured using the Post-COVID-19 Functional Status Scale (absence of symptoms related to the illness, ability to do usual daily activities, and a return to a previous state of health and mind). MAIN RESULTS: Laboratory studies review We identified five laboratory studies. Amyloid fibrin(ogen) particles were identified in participants across all studies, including those with PCC, healthy individuals, and those with diabetes. The results of three studies were based on visual images of amyloid fibrin(ogen) particles, which did not quantify the amount or size of the particles identified. Formal risk of bias assessment showed concerns in how the studies were conducted and reported. This means the results were insufficient to support the belief that amyloid fibrin(ogen) particles are associated with PCC, or to determine whether there is a difference in the amount or size of amyloid fibrin(ogen) particles in the plasma of people with PCC compared to healthy controls. Randomized controlled trials review We identified no trials meeting our inclusion criteria. AUTHORS' CONCLUSIONS: In the absence of reliable research showing that amyloid fibrin(ogen) particles contribute to the pathophysiology of PCC, there is no rationale for plasmapheresis to remove amyloid fibrin(ogen) particles in PCC. Plasmapheresis for this indication should not be used outside the context of a well-conducted randomized controlled trial.

Immobilized fibrinogen activates human platelets through glycoprotein VI.

Glycoprotein VI, a major platelet activation receptor for collagen and fibrin, is considered a particularly promising, safe antithrombotic target. In this study, we show that human glycoprotein VI signals upon platelet adhesion to fibrinogen. Full spreading of human platelets on fibrinogen was abolished in platelets from glycoprotein VI- deficient patients suggesting that fibrinogen activates platelets through glycoprotein VI. While mouse platelets failed to spread on fibrinogen, human-glycoprotein VI-transgenic mouse platelets showed full spreading and increased Ca2+ signaling through the tyrosine kinase Syk. Direct binding of fibrinogen to human glycoprotein VI was shown by surface plasmon resonance and by increased adhesion to fibrinogen of human glycoprotein VI-transfected RBL-2H3 cells relative to mock-transfected cells. Blockade of human glycoprotein VI with the Fab of the monoclonal antibody 9O12 impaired platelet aggregation on preformed platelet aggregates in flowing blood independent of collagen and fibrin exposure. These results demonstrate that human glycoprotein VI binds to immobilized fibrinogen and show that this contributes to platelet spreading and platelet aggregation under flow.

Ischaemic Stroke, Thromboembolism and Clot Structure.

Ischaemic stroke is a major cause of morbidity and mortality worldwide. Blood clotting and thromboembolism play a central role in the pathogenesis of ischaemic stroke. An increasing number of recent studies indicate changes in blood clot structure and composition in patients with ischaemic stroke. In this review, we aim to summarise and discuss clot structure, function and composition in ischaemic stroke, including its relationships with clinical diagnosis and treatment options such as thrombolysis and thrombectomy. Studies are summarised in which clot structure and composition is analysed both in vitro from patients' plasma samples and ex vivo in thrombi obtained through interventional catheter-mediated thrombectomy. Mechanisms that drive clot composition and architecture such as neutrophil extracellular traps and clot contraction are also discussed. We find that, while in vitro clot structure in plasma samples from ischaemic stroke patients are consistently altered, showing denser clots that are more resistant to fibrinolysis, current data on the composition and architecture of ex vivo clots obtained by thrombectomy are more variable. With the potential of advances in technologies underpinning both the imaging and retrieving of clots, we expect that future studies in this area will generate new data that is of interest for the diagnosis, optimal treatment strategies and clinical management of patients with ischaemic stroke.

Glucose variability is associated with an adverse vascular profile but only in the presence of insulin resistance in individuals with type 1 diabetes: An observational study.

AIMS/HYPOTHESIS: We hypothesised that the detrimental effect of high glucose variability (GV) in people with type 1 diabetes is mainly evident in those with concomitant insulin resistance. METHODS: We conducted secondary analyses on continuous glucose monitoring (CGM) using baseline observational data from three randomised controlled trials and assessed the relationship with established vascular markers. We used standard CGM summary statistics and principal component analysis to generate individual glucose variability signatures for each participant. Cluster analysis was then employed to establish three GV clusters (low, intermediate, or high GV, respectively). The relationship with thrombotic biomarkers was then investigated according to insulin resistance, assessed as estimated glucose disposal rate (eGDR). RESULTS: Of 107 patients, 45%, 37%, and 18% of patients were assigned into low, intermediate, and high GV clusters, respectively. Thrombosis biomarkers (including fibrinogen, plasminogen activator inhibitor-1, tissue factor activity, and tumour necrosis factor-alpha) increased in a stepwise fashion across all three GV clusters; this increase in thrombosis markers was evident in the presence of low but not high eGDR and at a threshold of eGDR <5.1 mg/kg/min. CONCLUSION: Higher GV is associated with increased thrombotic biomarkers in type 1 diabetes but only in those with concomitant insulin resistance.

Fibrinogen αC-subregions critically contribute blood clot fibre growth, mechanical stability, and resistance to fibrinolysis.

Fibrinogen is essential for blood coagulation. The C-terminus of the fibrinogen α-chain (αC-region) is composed of an αC-domain and αC-connector. Two recombinant fibrinogen variants (α390 and α220) were produced to investigate the role of subregions in modulating clot stability and resistance to lysis. The α390 variant, truncated before the αC-domain, produced clots with a denser structure and thinner fibres. In contrast, the α220 variant, truncated at the start of the αC-connector, produced clots that were porous with short, stunted fibres and visible fibre ends. These clots were mechanically weak and susceptible to lysis. Our data demonstrate differential effects for the αC-subregions in fibrin polymerisation, clot mechanical strength, and fibrinolytic susceptibility. Furthermore, we demonstrate that the αC-subregions are key for promoting longitudinal fibre growth. Together, these findings highlight critical functions of the αC-subregions in relation to clot structure and stability, with future implications for development of novel therapeutics for thrombosis.

Fibrin clot structure remains unaffected in young, healthy individuals after transient exposure to diesel exhaust.

Exposure to urban particulate matter has been associated with an increased risk of cardiovascular disease and thrombosis. We studied the effects of transient exposure to diesel particles on fibrin clot structure of 16 healthy individuals (age 21-44). The subjects were randomly exposed to diesel exhaust and filtered air on two separate occasions. Blood samples were collected before exposure, and 2 and 6 hours after exposure. There were no significant changes on clot permeability, maximum turbidity, lag time, fibre diameter, fibre density and fibrinogen level between samples taken after diesel exhaust exposure and samples taken after filtered air exposure. These data show that there are no prothrombotic changes in fibrin clot structure in young, healthy individuals exposed to diesel exhaust.

Impact of γ'γ' fibrinogen interaction with red blood cells on fibrin clots.

AIM: γ' fibrinogen has been associated with thrombosis. Here the interactions between γ'γ' or γAγA fibrinogen and red blood cells (RBCs), and their role on fibrin clot properties were studied. MATERIALS & METHODS: Atomic Force microscopy (AFM)-based force spectroscopy, rheological, electron and confocal microscopy, and computational approaches were conducted for both fibrinogen variants. RESULTS & CONCLUSION: AFM shows that the recombinant human (rh)γ'γ' fibrinogen increases the binding force and the frequency of the binding to RBCs compared with rhγAγA, promoting cell aggregation. Structural changes in rhγ'γ' fibrin clots, displaying a nonuniform fibrin network were shown by microscopy approaches. The presence of RBCs decreases the fibrinolysis rate and increases viscosity of rhγ'γ' fibrin clots. The full length of the γ' chain structure, revealed by computational analysis, occupies a much wider surface and is more flexible, allowing an increase of the binding between γ' fibers, and eventually with RBCs.

The 95RGD97 sequence on the Aα chain of fibrinogen is essential for binding to its erythrocyte receptor.

BACKGROUND: Erythrocyte aggregation, a cardiovascular risk factor, is increased by high plasma fibrinogen levels. Here, the effect of different fibrinogen mutations on binding to its human erythrocyte receptor was assessed in order to identify the interaction sites. METHODS: Three fibrinogen variants were tested, specifically mutated in their putative integrin recognition sites on the Aα chain (mutants D97E, D574E and D97E/D574E) and compared with wild-type fibrinogen. RESULTS: Atomic force microscopy-based force spectroscopy measurements showed a significant decrease both on the fibrinogen-erythrocyte binding force and on its frequency for fibrinogen with the D97E mutation, indicating that the corresponding arginine-glycine-aspartate sequence (residues 95-97) is involved in this interaction, and supporting that the fibrinogen receptor on erythrocytes has a ß3 subunit. Changes in the fibrin clot network structure obtained with the D97E mutant were observed by scanning electron microscopy. CONCLUSION: These findings may lead to innovative perspectives on the development of new therapeutic approaches to overcome the risks of fibrinogen-driven erythrocyte hyperaggregation.

A fibrin biofilm covers blood clots and protects from microbial invasion.

Hemostasis requires conversion of fibrinogen to fibrin fibers that generate a characteristic network, interact with blood cells, and initiate tissue repair. The fibrin network is porous and highly permeable, but the spatial arrangement of the external clot face is unknown. Here we show that fibrin transitioned to the blood-air interface through Langmuir film formation, producing a protective film confining clots in human and mouse models. We demonstrated that only fibrin is required for formation of the film, and that it occurred in vitro and in vivo. The fibrin film connected to the underlying clot network through tethering fibers. It was digested by plasmin, and formation of the film was prevented with surfactants. Functionally, the film retained blood cells and protected against penetration by bacterial pathogens in a murine model of dermal infection. Our data show a remarkable aspect of blood clotting in which fibrin forms a protective film covering the external surface of the clot, defending the organism against microbial invasion.