Covid-19, induced activation of hemostasis, and immune reactions: Can an auto-immune reaction contribute to the delayed severe complications observed in some patients?

Covid-19 is characterized by weak symptoms in most affected patients whilst severe clinical complications, with frequent fatal issues, occur in others. Disease severity is associated with age and comorbidities. Understanding of viral infectious mechanisms, and antibody immune response, can help to better control disease progression.

Laboratory assessment of Activated Protein C Resistance/Factor V-Leiden and performance characteristics of a new quantitative assay.

Activated Protein C Resistance is mainly associated to a factor V mutation (RQ506), which induces a deficient inactivation of activated factor V by activated protein C, and is associated to an increased risk of venous and arterial thrombosis in affected individuals, caused by the prolonged activated factor V survival. Its prevalence is mainly in Caucasians (about 5%), and this mutation is absent in Africans and Asians. Presence of Factor V-Leiden is usually evidenced with clotting methods, using a two-step APTT assay performed without or with APC: prolongation of blood coagulation time is decreased if this factor is present.

Anti-phospholipid syndrome: Current opinion on mechanisms involved, laboratory characterization and diagnostic aspects.

Anti-phospholipid syndrome is a complex and severe clinical situation, associated with symptoms such as recurrent thrombosis, arterial or venous, at any site, pregnancy loss, and other related syndromes. These clinical burdens, are highly variable from patient to patient, and are associated with biological abnormalities, such as the presence of the Lupus Anticoagulant or phospholipid dependent antibodies, confirmed on two occasions at least 12 weeks apart. From the diagnosis standpoint, both, functional (clotting) or immunological assays, are difficult to standardize and to optimize, due to the absence of reference material, or a characteristic clinical group, and international reference preparations.

Update on functional and genetic laboratory assays for the detection of platelet microvesicles.

Functional and genetic assays for measuring platelet microvesicles (PMVs) are presented and discussed. Functional assays concern two groups of methods: a) homogeneous assays using the cofactor activity of phospholipids (PPLs) contained in PMVs and present in assayed plasmas, and a coagulation or a thrombin generation assay (TGA) as "end points"; b) capture-based assays, in which PMVs bind to an immobilized ligand, such as Annexin V in the presence of calcium, or monoclonal antibodies (MoAbs) specific for membrane proteins. Genetic assays aim to detect micro-RNA (miRNA) present in PMVs: miRNA must be extracted from plasma, and the expression pattern can be determined by various methods such as quantitative real-time PCR, microarray or sequencing.

Increased thrombin generation in persons with echogenic carotid plaques.

Echolucent carotid plaques are associated with higher risk for future ischemic cerebrovascular events (CVE) than echogenic plaques independent of the degree of stenosis. Elevated markers of thrombin generation are associated with atherosclerotic plaques and are increased in the acute and chronic phases of CVE. The present study was conducted to investigate the influence of plaque morphology on thrombin generation in persons with carotid stenosis.

Endothelial dysfunction and systemic inflammation in persons with echolucent carotid plaques.

Echolucent carotid plaques are associated with high risk for future ischemic cerebrovascular events independent of the degree of stenosis. Elevated levels of markers of systemic inflammation and endothelial dysfunction are predictors for future myocardial infarction and stroke. The present study was undertaken to investigate the relations between plaque morphology, endothelial dysfunction assessed by tissue-plasminogen activator antigen (t-PA ag) and vonWillebrand factor (vWF), and systemic inflammation in persons with carotid stenosis.

Affinity purification of heparin-dependent antibodies to platelet factor 4 developed in heparin-induced thrombocytopenia: biological characteristics and effects on platelet activation.

Antibodies to heparin platelet factor 4 (H-PF4) complexes were purified from the plasma of three patients with heparin-induced thrombocytopenia (HIT) using affinity chromatography. From each plasma, the largest amount of antibodies was eluted with 2 M NaCl at pH 7.5 (peak 1) and the remainder was obtained using 0.

Generation and pathogenicity of anti-platelet factor 4 antibodies: diagnostic implications.

Recent studies have elucidated the target antigens for heparin-dependent antibodies. The major one is generated in the presence of platelet factor 4 (PF4) and heparin at a well-defined concentration that allows formation of heparin-PF4 complexes and induces an alteration of the PF4 molecule. It then exposes neoepitopes, which bind the heparin-dependent antibodies.

Elevated plasma fibrinogen and increased fibrin turnover among healthy women who both smoke and use low-dose oral contraceptives--a preliminary report.

Among users of low-dose oral contraceptives (OC), cardiovascular diseases occur mainly in smokers. The mechanisms by which OC and smoking increase the risk for arterial thrombotic risk have not been adequately explained. Epidemiological evidence suggests that changes in blood coagulation and fibrinolysis may play an important role as determinants of thrombotic events.

Antibodies to platelet factor 4-heparin after cardiopulmonary bypass in patients anticoagulated with unfractionated heparin or a low-molecular-weight heparin : clinical implications for heparin-induced thrombocytopenia.

Background: Cardiopulmonary bypass (CPB) induces platelet activation with release of platelet factor 4 (PF4), and patients are exposed to high doses of heparin (H). We investigated whether this contributes to the development of antibodies to H-PF4 and heparin-induced thrombocytopenia (HIT).

Methods And Results: CPB was performed with unfractionated heparin (UFH) in 328 patients.

Prevalence of antiphospholipid-related antibodies in unselected patients with history of venous thrombosis.

Antiphospholipid antibodies (aPL) are heterogeneous and are now accepted to be mainly phospholipid-protein-dependent antibodies. Although these antibodies are classically associated with thrombosis, their clinical relevance remains to be established. The subgroups of antibodies characterized by their proteic targets were reported to be more appropriate thrombotic markers.

High incidence of anti-heparin/platelet factor 4 antibodies after cardiopulmonary bypass surgery.

Fifty-one patients undergoing cardiopulmonary bypass (CPB) were studied on day 0 and day 8 for heparin-induced thrombocytopenia (HIT). The platelet aggregation test (PAT) and tests for anti-heparin-platelet factor 4 (anti-H.PF4), anti-IL8 and anti-neutrophil activating peptide 2 (anti-NAP2) antibodies (Ab) were performed by ELISA.

New insights into the negative regulation of hematopoiesis by chemokine platelet factor 4 and related peptides.

Platelet factor 4 (PF4) has been recognized as an inhibitor of myeloid progenitors. However, the mechanism of action of this chemokine remains poorly understood. The present study was designed to determine its structure/function relationship.

Longitudinal study on activated factors XII and VII levels during normal pregnancy.

Levels of activated factor XII (FXIIa) and VII (FVIIa) were determined in 100 women with uneventful pregnancies. Samples were divided into five study intervals: three during pregnancy, one at delivery and one 3 d postpartum. The median (range) for FXIIa levels were 3.

Differences in specificity of heparin-dependent antibodies developed in heparin-induced thrombocytopenia and consequences on cross-reactivity with danaparoid sodium.

Heparin-induced thrombocytopenia (HIT) is frequently associated with antibodies (Abs) to heparin-PF4 complexes (H-PF4). In order to investigate whether there are variations in specificity of Abs, we studied 63 samples from patients with suspected HIT. Two groups of samples were separated after comparing their reactivity against H-PF4 or recombinant PF4 (r-PF4) using ELISA.

Annexin V, a new marker of platelet storage lesion: correlation with dMPV.

Released annexin V, an intracellular platelets glycoprotein, was used to determine the cellular injury which occurred during storage of platelet concentrates. Twenty-eight units of leuco-reduced apheresis platelet concentrates, obtained without leucocyte filtration, were analysed. Released annexin V showed a significant correlation with EDTA-induced shape changes of platelet (r = 0.

Absence of cross-reactivity of SR90107A/ORG31540 pentasaccharide with antibodies to heparin-PF4 complexes developed in heparin-induced thrombocytopenia.

New carbohydrate-based anticoagulants devoid of the side effects of unfractionated heparin are currently under development and show a major potential for patients with heparin-induced thrombocytopenia (HIT) who still require efficient antithrombotic therapy. As HIT is usually associated with antibodies to heparin-platelet factor 4 (H-PF4) complexes, cross-reactivity of the heparin pentasaccharide SR90107A/ORG31540 was tested in the presence of PF4 with the plasma from 49 patients with HIT. No cross-reactivity was observed whatever the pentasaccharide concentrations.

Different target specificities of phospholipid-dependent antibodies.

Phospholipid dependent antibodies are usually measured with assays for antiphospholipid/anticardiolipin antibodies (aPLA) or for lupus anticoagulant (LA) activity. Most of them are targeted to complexes of beta 2-glycoprotein I (beta 2-GPI) and anionic phospholipids (PLP) or to prothrombin for some LA. New understandings allow a better standardisation and optimisation of assays' reactivity.

Population correlates of coagulation factor VII. Importance of age, sex, and menopausal status as determinants of activated factor VII.

Factor VII coagulant activity (FVIIc) has been found to be related to cardiovascular risk factors and may be an independent predictor of coronary heart disease (CHD). Whether these associations are due to changes in FVII activation rather than FVII concentration remain unclear. Therefore, we investigated the relationships between activated factor VII (FVIIa) and CHD risk factors in healthy subjects (336 men and 348 women) aged 25 to 64 years.

Presence of autoantibodies to interleukin-8 or neutrophil-activating peptide-2 in patients with heparin-associated thrombocytopenia.

Eighty-seven patients with heparin-associated thrombocytopenia (HAT) showed either a positive heparin platelet aggregometry test result and/or the presence of antibodies to heparin-platelet factor 4 (H-PF4) complexes by enzyme-linked immunosorbent assay (ELISA). Fifteen of these patients lacked antibodies to H-PF4, and plasma from these patients was analyzed for the presence of antibodies to PF4-related chemokines, Neutrophil-activating peptide-2 (NAP-2) and interleukin-8 (IL-8). Of these 15 patients, 6 showed antibodies to IL-8 and 3 to the platelet basic protein (PBP)-derived protein, NAP-2.

Generation of antibodies to heparin-PF4 complexes without thrombocytopenia in patients treated with unfractionated or low-molecular-weight heparin.

The incidence of antibodies to heparin-PF4 complexes (H-PF4) has been evaluated in patients who were under heparin therapy for more than 7 days: 109 patients treated with unfractionated heparin (UH) and 100 patients with low-molecular-weight heparin (LMWH). The presence of antibodies was identified in 17% of the former group and 8% of the latter. In both the UH and the LMWH groups, IgM antibodies were found in all but four patients who showed IgA antibodies.

Pathogenicity of IgA and/or IgM antibodies to heparin-PF4 complexes in patients with heparin-induced thrombocytopenia.

Antibodies to heparin-PF4 (H-PF4) complexes have been tested and isotyped in 38 patients who developed severe heparin-induced thrombocytopenia (type II HIT). All patients had a platelet count <120x10(9)/1 or a reduction of > 30% of the initial value, occurring at least 5d after the onset of heparin. Thrombocytopenia, which rapidly reversed following the withdrawal of heparin, was associated with thrombosis in nine patients.

A new, semi-quantitative and individual ELISA for rapid measurement of plasma D-dimer in patients suspected of pulmonary embolism.

The performance of a new membrane ELISA for semi-quantitative determination of plasma D-dimer has been evaluated. Its cut-off is about 500 ng/ml FEU and this single test is completed within 10 min. D-dimer was measured in 301 patients suspected of pulmonary embolism by conventional microplate and membrane ELISA.

A new sensitive membrane based ELISA technique for instantaneous D.Dimer evaluation in emergency.

D.Dimer is currently used as a diagnotic help in thromboembolic events. The first application widely validated concerns the exclusion diagnosis of deep vein thrombosis and pulmonary embolism.