Tag: fibrin

Fibrin microthrombi in bladder urothelium after SARS-CoV-2 infection: Case report

Abstract:

A 45-year-old male with diabetes, hypertension and hyperlipidemia was referred to urology due to persistent symptoms of urinary frequency, urgency, nocturia, erectile dysfunction, and constant pain localized to the bladder, pelvis, and perineal area, 3–4 months after SARS-CoV-2 infection. A bladder biopsy showed urothelial mucosa and submucosa with hemorrhage and fibrin microthrombi in blood vessels. Hydrodistention of the bladder and pelvic floor physical therapy resolved symptoms, though bladder and pain symptoms returned upon reinfection with SARS-CoV-2. Urinalysis revealed elevated urinary interleukin-8, which may indicate localized bladder inflammation.

Source: Hoang Roberts L, Zwaans BMM, Jabbar K, Bartolone SN, Padmanabhan P, Peters KM. Fibrin microthrombi in bladder urothelium after SARS-CoV-2 infection: Case report. Urol Case Rep. 2023 Sep 25;51:102575. doi: 10.1016/j.eucr.2023.102575. PMID: 37829494; PMCID: PMC10565678. https://www.sciencedirect.com/science/article/pii/S2214442023002619 (Full text)

Are fibrinaloid microclots a cause of autoimmunity in Long Covid and other post-infection diseases?

Abstract:

It is now well established that the blood-clotting protein fibrinogen can polymerise into an anomalous form of fibrin that is amyloid in character; the resultant clots and microclots entrap many other molecules, stain with fluorogenic amyloid stains, are rather resistant to fibrinolysis, can block up microcapillaries, are implicated in a variety of diseases including Long COVID, and have been referred to as fibrinaloids. A necessary corollary of this anomalous polymerisation is the generation of novel epitopes in proteins that would normally be seen as ‘self’, and otherwise immunologically silent.

The precise conformation of the resulting fibrinaloid clots (that, as with prions and classical amyloid proteins, can adopt multiple, stable conformations) must depend on the existing small molecules and metal ions that the fibrinogen may (and is some cases is known to) have bound before polymerisation. Any such novel epitopes, however, are likely to lead to the generation of autoantibodies.

A convergent phenomenology, including distinct conformations and seeding of the anomalous form for initiation and propagation, is emerging to link knowledge in prions, prionoids, amyloids and now fibrinaloids. We here summarise the evidence for the above reasoning, which has substantial implications for our understanding of the genesis of autoimmunity (and the possible prevention thereof) based on the primary process of fibrinaloid formation.

Source: Kell DB, Pretorius E. Are fibrinaloid microclots a cause of autoimmunity in Long Covid and other post-infection diseases? Biochem J. 2023 Aug 16;480(15):1217-1240. doi: 10.1042/BCJ20230241. PMID: 37584410. https://portlandpress.com/biochemj/article/480/15/1217/233389/Are-fibrinaloid-microclots-a-cause-of-autoimmunity (Full text)

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

Abstract:

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.

Source: Fox T, Hunt BJ, Ariens RA, Towers GJ, Lever R, Garner P, Kuehn R. Plasmapheresis to remove amyloid fibrin(ogen) particles for treating the post-COVID-19 condition. Cochrane Database Syst Rev. 2023 Jul 26;7(7):CD015775. doi: 10.1002/14651858.CD015775. PMID: 37491597; PMCID: PMC10368521. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10368521/ (Full text)

Rapid flow cytometric analysis of fibrin amyloid microclots in Long COVID

Abstract:

Long COVID has become a significant global health and economic burden, yet there are currently no established diagnostic tools to identify which patients might benefit from specific treatments. One of the major pathophysiological factors contributing to Long COVID is the presence of hypercoagulability; this results in insoluble amyloid microclots that are resistant to fibrinolysis.

Our previous research using fluorescence microscopy has demonstrated a significant amyloid microclot load in Long COVID patients. However, this approach lacked statistical robustness, objectivity, and rapid throughput. In the current study, we have used imaging flow cytometry for the first time to show significantly increased concentration and size of these microclots.

We identified notable variations in size and fluorescence between microclots in Long COVID and those of controls even using a 20x objective. By combining cell imaging and the high-event-rate nature of a conventional flow cytometer, imaging flow cytometry can eliminate erroneous results and increase accuracy in gating and analysis beyond what pure quantitative measurements from conventional flow cytometry can provide.

Although imaging flow cytometry was used in our study, our results suggest that the signals indicating the presence of microclots should be easily detectable using a conventional flow cytometer. Flow cytometry is a more widely available technique which has been used in pathology laboratories for decades, rendering it a potentially more suitable and accessible method for detecting microclots in individuals suffering from both Long COVID and other conditions with similar pathology, such as myalgic encephalomyelitis.

Source: Turner, Simone and Laubscher, Gert Jacobus and Khan, M. Asad and Kell, Douglas and Pretorius, Etheresia, Rapid Flow Cytometric Analysis of Fibrin Amyloid Microclots in Long COVID. Available at SSRN: https://ssrn.com/abstract=4405265 or http://dx.doi.org/10.2139/ssrn.4405265 https://assets.researchsquare.com/files/rs-2731434/v1/0b4877b0-99fa-499c-9d65-3b6e43865d86.pdf?c=1680099696 (Full text)

Treatment of Long COVID symptoms with triple anticoagulant therapy

Abstract:

Background: Fibrin(ogen) amyloid microclots and platelet hyperactivation are key pathological findings in patients with acute COVID-19 infection and also in those with Long COVID/Post-Acute Sequelae of COVID-19 (PASC). These pathologies may represent a suitable target for pharmacological treatment of Long COVID.

Methods: Here we report on the symptoms displayed by a cohort of 91 South African Long COVID patients at baseline and after a clinician-initiated anticoagulant regime was completed. For laboratory analysis, patients provided a blood sample before and after treatment. Fibrinaloid microclot presence was studied by adding thioflavin T to platelet poor plasma (PPP), whilst platelet hyperactivation was studied using two platelet markers- PAC1 and CD62P (P-selectin). The anticoagulant regime included dual antiplatelet therapy (DAPT- Clopidogrel 75mg + Aspirin 75mg) once a day, and a direct oral anticoagulant (DOAC- Apixaban) 5mg twice a day. A proton pump inhibitor (PPI) pantoprazole 40 mg/day was also prescribed for gastric protection. Each of the treated cases reported their main Long COVID symptoms, and whether their symptoms resolved following treatment or not.

Results: In our cohort a most participants did not report any comorbidities before acute COVID-19 infection. Hypertension and dyslipidaemia were the commonest underlying illnesses, whilst the most commonly reported Long COVID symptoms included fatigue, cognitive dysfunction, shortness of breath, and joint and muscle pains. Following completion of treatment, each of the different symptoms resolved in the majority of patients. This was also reflected in the laboratory analysis, where a decrease in the severity of fibrin amyloid microclotting and the degree of platelet pathology was noted. No serious adverse bleeding events were reported.

Conclusions: Fibrin amyloid microclots, platelet hyperactivation/ aggregation, and  widespread endothelialitis inhibit the transport of oxygen at a capillary/cellular level. This provides a ready explanation for the symptoms of Long COVID. By normalizing the failed clotting physiology and reversal of the endothelialitis, triple anticoagulant therapy represents a promising treatment option that appears to be highly efficacious, and warrants controlled clinical studies. We caution that such a regime must only be followed under expert medical supervision in view of the risk of  bleeding.

Source: Gert J Laubscher, M Asad Khan, Chantelle Venter, Etheresia Pretorius et al. Treatment of Long COVID symptoms with triple anticoagulant therapy, 21 March 2023, PREPRINT (Version 1) available at Research Square https://doi.org/10.21203/rs.3.rs-2697680/v1 (Full text)

Increased levels of inflammatory molecules in blood of Long COVID patients point to thrombotic endotheliitis

Abstract:

The prevailing hypotheses for the persistent symptoms of Long COVID have been narrowed down to immune dysregulation and autoantibodies, widespread organ damage, viral persistence, and fibrinaloid microclots (entrapping numerous inflammatory molecules) together with platelet hyperactivation. Here we demonstrate significantly increased concentrations of Von Willebrand Factor, platelet factor 4, serum amyloid A, alpha-2-antiplasmin E-selectin, and platelet endothelial cell adhesion molecule-1, in the soluble part of the blood.

It was noteworthy that the mean level of alpha-2-antiplasmin exceeded the upper limit of the laboratory reference range in Long COVID patients, and the other 5 were significantly elevated in Long COVID patients as compared to the controls. This is alarming if we take into consideration that a significant amount of the total burden of these inflammatory molecules has previously been shown to be entrapped inside fibrinolysis-resistant microclots (thus decreasing the apparent level of the soluble molecules). We also determined that by individually adding E-selectin and PECAM-1 to healthy blood, these molecules may indeed be involved in protein-protein interactions with plasma proteins (contributing to microclot formation) and platelet hyperactivation. This investigation was performed as a laboratory model investigation and the final exposure concentration of these molecules was chosen to mimic concentrations found in Long COVID.

We conclude that presence of microclotting, together with relatively high levels of six inflammatory molecules known to be key drivers of endothelial and clotting pathology, points to thrombotic endotheliitis as a key pathological process in Long COVID. This has implications for the choice of appropriate therapeutic options in Long COVID.

Source: Simone Turner, Caitlin Naidoo, Thomas Usher, Arneaux Kruger, Chantelle Venter, Gert J Laubscher, M Asad Khan, Douglas B Kell, Etheresia Pretorius. Increased levels of inflammatory molecules in blood of Long COVID patients point to thrombotic endotheliitis. medRxiv 2022.10.13.22281055; doi: https://doi.org/10.1101/2022.10.13.22281055 (Full text available as PDF file)

A central role for amyloid fibrin microclots in long COVID/PASC: origins and therapeutic implications

Abstract:

Post-acute sequelae of COVID (PASC), usually referred to as ‘Long COVID’ (a phenotype of COVID-19), is a relatively frequent consequence of SARS-CoV-2 infection, in which symptoms such as breathlessness, fatigue, ‘brain fog’, tissue damage, inflammation, and coagulopathies (dysfunctions of the blood coagulation system) persist long after the initial infection. It bears similarities to other post-viral syndromes, and to myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS).

Many regulatory health bodies still do not recognize this syndrome as a separate disease entity, and refer to it under the broad terminology of ‘COVID’, although its demographics are quite different from those of acute COVID-19. A few years ago, we discovered that fibrinogen in blood can clot into an anomalous ‘amyloid’ form of fibrin that (like other β-rich amyloids and prions) is relatively resistant to proteolysis (fibrinolysis). The result, as is strongly manifested in platelet-poor plasma (PPP) of individuals with Long COVID, is extensive fibrin amyloid microclots that can persist, can entrap other proteins, and that may lead to the production of various autoantibodies. These microclots are more-or-less easily measured in PPP with the stain thioflavin T and a simple fluorescence microscope.

Although the symptoms of Long COVID are multifarious, we here argue that the ability of these fibrin amyloid microclots (fibrinaloids) to block up capillaries, and thus to limit the passage of red blood cells and hence O2 exchange, can actually underpin the majority of these symptoms. Consistent with this, in a preliminary report, it has been shown that suitable and closely monitored ‘triple’ anticoagulant therapy that leads to the removal of the microclots also removes the other symptoms. Fibrin amyloid microclots represent a novel and potentially important target for both the understanding and treatment of Long COVID and related disorders.

Source: Kell DB, Laubscher GJ, Pretorius E. A central role for amyloid fibrin microclots in long COVID/PASC: origins and therapeutic implications. Biochem J. 2022 Feb 17;479(4):537-559. doi: 10.1042/BCJ20220016. PMID: 35195253. https://portlandpress.com/biochemj/article/479/4/537/230829/A-central-role-for-amyloid-fibrin-microclots-in (Full text)