Phospholipid-binding Plasma Proteins Research Articles

Interaction of the antiphospholipid syndrome autoantigen beta-2 glycoprotein I with DNA and neutrophil extracellular traps

Beta-2 glycoprotein I (β2GPI) is a phospholipid-binding plasma protein and prominent autoantigen in antiphospholipid syndrome (APS). Here, we tested the hypothesis that β2GPI might bind to not only phospholipids, but also cell-free DNA and neutrophil extracellular traps (NETs). We report that β2GPI interacts with cell-free DNA from different species, as well as NETs, in a dose-dependent manner, retarding their migration in an agarose-gel electrophoretic mobility shift assay. We confirm the direct binding interaction of β2GPI with DNA and NETs by ELISA. We also demonstrate that β2GPI colocalizes with NET strands by immunofluorescence microscopy. Finally, we provide evidence that β2GPI-DNA complexes can be detected in the plasma of APS patients, where they positively correlate with an established biomarker of NET remnants. Taken together, our findings indicate that β2GPI interacts with DNA and NETs, and suggest that this interaction may play a role as a perpetuator and/or instigator of autoimmunity in APS.

Gastrointestinal symptoms as the first manifestation of antiphospholipid syndrome

BackgroundAntiphospholipid syndrome (APS) is an acquired pre-thrombotic autoimmune condition, which produces autoantibodies called antiphospholipid antibodies (APL) against phospholipid-binding plasma proteins. The diagnosis of APS requires at least one of Sapporo standard clinical manifestations and one laboratory criteria (persistently medium/high titer anticardiolipin antibodies, and/or medium/high titer anti-β2-glycoprotein I antibodies, and/or a positive lupus anticoagulant test). Gastrointestinal lesions are rarely reported in APS patients. APS cases with recurrent abdominal pain as the first clinical manifestation are even rarer.Case presentationThis report describes an APS case with recurrent abdominal pain as the first clinical manifestation of antiphospholipid syndrome. The patient has a history of two miscarriages. Computed tomography of the abdomen confirmed mesenteric thrombosis and intestinal obstruction while laboratory tests for serum antiphospholipid and anti-β2-glycoprotein I antibodies were positive. This led to the diagnosis of APS.ConclusionsThis paper provides useful information on gastrointestinal manifestations and APS, also including a brief literature review about possible gastrointestinal symptoms of APS.

A Simplified Understanding of the Black Swan: Anti-phospholipid Antibody Syndrome

Anti-phospholipid Antibody Syndrome or Hugh's syndrome is a heterogeneous disorder, first fully described in 1980s. The syndrome is caused by the presence of specific antibodies against phospholipid binding plasma proteins in the serum of the patient, with or without underlying autoimmune diseases, that causes prolongation of tests of coagulation. High index of clinical suspicion is required for diagnosis of Anti-phospholipid Antibody Syndrome. Stroke or myocardial infarction in young, unprovoked recurrent deep vein thrombosis and recurrent pregnancy loss are typical scenarios where Anti-Phospholipid Antibody Syndrome should be suspected. Presence of non-criteria manifestations like livedo reticularis, skin ulcers, nephropathy, valvular heart disease and thrombocytopenia adds to diagnostic clue for presence of Anti-Phospholipid Antibody Syndrome. Treatment of Anti-Phospholipid Antibody Syndrome has preventive and therapeutic aspects that usually focus on thrombotic and obstetric manifestations of the disease. Therapeutic anti-coagulation with heparin followed by warfarin is required for patients presenting with acute thrombosis. Those with venous thrombosis are given moderate intensity warfarin International Normalized Ratio, 2-3), whereas those with arterial thrombosis or recurrent venous thrombosis even on warfarin are treated with high intensity warfarin (International Normalized Ratio, 3-4). Similarly, anticoagulation with heparin is advised in patients with obstetric Anti-Phospholipid Antibody Syndrome throughout pregnancy and up to six weeks postpartum. Treatment recommendations are still not clear for asymptomatic Anti-Phospholipid Antibody Syndrome positive patients and in those with non-criteria manifestations of the disease. Steroids, intravenous immunoglobulin and immunosuppressant are reported to be effective in severe cases of catastrophic antiphospholid syndrome characterized by rapid small vessel thrombotic involvement of multiple organ systems. Studies are evaluating the efficacy of direct thrombin inhibitors in the management of refractory cases. Keywords: anticoagulants; anti-phospholipid syndrome; obstetric APS; thrombotic APS.

Antiphospholipid syndrome – an update

Antiphospholipid syndrome (APS) is an autoantibody-mediated acquired thrombophilia. It is characterized by the presence of antiphospholipid antibodies (APL) that are directed against phospholipid-binding plasma proteins, such as beta-2-glycoprotein I (b2GPI). Its main manifestations are recurrent vascular thromboses (so-called "thrombotic APS") and pregnancy complications ("obstetric APS"). According to the current consensus criteria, a persistently positive functional lupus anticoagulant (LA) assay and/or the presence of anti-b2GPI and/or anti-cardiolipin antibodies, together with clinical symptoms, is mandatory for the diagnosis of APS. Other clinical features, such as thrombocytopenia, Coombs-positive haemolytic anaemia, heart valve disease, renal microangiopathy and neurologic disorders are also common in APL-positive patients. APS can be associated with other autoimmune disorders, such as systemic lupus erythematosus. In rare cases, catastrophic APS (CAPS) occurs, with the development of excessive thrombosis at multiple sites, usually affecting small vessels and leading to multi-organ dysfunction and organ failure. Treatment usually comprises antithrombotic therapy using antiplatelet and anticoagulant agents. However, there is no consensus concerning the intensity or duration of therapy. Despite apparently adequate anticoagulation, the risk of recurrent thrombosis remains high. For patients with CAPS, a combined therapeutic approach that includes anticoagulation, glucocorticoids, plasma exchange and/or intravenous immunoglobulin seems to be the best treatment option. Keywords: Antiphospholipid syndrome, lupus anticoagulants, anti-cardiolipin, anti-beta-2-glycoprotein I, vascular thrombosis, pregnancy complication.

Autoantibody profiling in APS.

The international consensus for the classification of antiphospholipid syndrome (APS) requires clinical and laboratory criteria to be considered at an equal level for diagnosing APS. Thus, detection of antiphospholipid antibodies (aPL) being a hallmark of APS has been the object of intensive investigation over the past 40 years. However, appropriate detection of aPL still remains a laboratory challenge due to their heterogeneity comprising autoantibodies reactive to different phospholipid-binding plasma proteins, such as beta-2 glycoprotein I (β2GPI) and prothrombin. The relevance of aPL interacting with phospholipids other than cardiolipin (CL, diphosphatidylglycerol), such as phosphatidylserine (PS), remains elusive with regard to the diagnosis of APS. Recently, the concept of aPL profiling has been introduced to assess the risk of thrombotic complications in patients with APS. New assay techniques, apart from enzyme-linked immunosorbent assays (ELISAs) recommended by the international consensus for the classification of APS, have been proposed for multiplexing of aPL testing. Line immunoassays (LIAs) employing a novel hydrophobic solid phase for the simultaneous detection of different aPL seem to be an intriguing alternative. We evaluated a novel multiplex LIA employing a hydrophobic membrane coated with different phospholipid (PL)-binding proteins or PLs. The performance characteristics of this new multiplexing assay technique demonstrated its usefulness for aPL profiling.

Atypical hemolytic-uremic syndrome associated with antiphospholipid antibodies and antiphospholipid syndrome; a novel presentation

Atypical hemolytic-uremic syndrome (HUS) is defined by the presence of microangiopathic hemolytic anemia, acute renal failure, and thrombocytopenia without a diarrheal prodrome. It is responsible for only ten percent of cases in children. The role of genetic deficiencies of complement regulation, Von Willebrand factor cleaving protease (ADAMTS 13), and intracellular defects of vitamin B12 metabolism has been known in pathogenesis of disease. Antiphospholipid antibodies (aPLs) are autoantibodies against negatively charged phospholipids or phospholipid binding plasma proteins. It is clear that their presence is associated with thrombosis, pregnancy morbidity, hematologic, skin, neurological conditions, and microangiopathy.

Clinical significance of phospholipid-cofactor antibodies in patients with systemic lupus erythematosus-associated antiphospholipid syndrome

To establish whether antibodies directed against phospholipid-binding plasma proteins such as β2-glycoprotein I (β2GPI), prothrombin (PT), and Annexin V (AnxV) constitute a risk factor for thrombosis in patients with systemic lupus erythematosus (SLE)-associated antiphospholipid syndrome (SLE/APS). A group of SLE patients (with and without APS) and patients with primary APS (PAPS) were included in this study. Fifteen patients with deep vein thrombosis but without antiphospholipid (aPL) antibodies, and another 15 age-matched and sex-matched apparently healthy individuals served as a control group. All patients were investigated for lupus anticoagulants and detection of anticardiolipin (aCL) immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies. Antibodies against β2GPI (IgG and IgM), PT (IgG and IgM), and AnxV (IgG) were also measured using the respective enzyme-linked immunosorbent assays. The study included 58 SLE patients (18 SLE/APS patients and 40 patients without APS) as well as 40 patients with PAPS, mean age 43 years (range: 18–74 years). IgG and/or IgM aCL antibodies were detected in all patients with PAPS (100%), whereas the prevalence rates of aPL-cofactor antibodies were as follows: 75% anti-β2GPI, 70% anti-PT, and 25% anti-AnxV antibodies. In SLE patients without APS, aCL antibodies were detected in 17.5%, anti-β2GPI antibodies in 20%, anti-AnxV antibodies in 20%, and anti-PT antibodies in 10% of patients. None of the antibodies measured were detected in deep vein thrombosis cases or healthy controls. Measurement of antiphospholipid-cofactor antibodies in addition to the more widely used aCL and anti-β2GPI antibodies could be a useful prognostic marker for the risk of thrombosis in SLE/APS patients.

Seronegative antiphospholipid syndrome

APS is an autoimmune disease that leads to arterial and/or venous thrombosis, recurrent pregnancy loss and persistently positive aPLs. Patients with clinical manifestations highly suggestive of APS but persistently negative conventional aPLs are classified as having seronegative APS. Ongoing research has revealed the existence of non-criteria antibodies proposed to be relevant to APS and that can be potentially included in the disease's classification criteria. We present a literature review on the most promising antibodies of this heterogeneous aPL family, which includes antibodies to a zwitterionic phospholipid, namely phosphatidylethanolamine, phospholipid-binding plasma proteins, phospholipid-protein complexes and anionic phospholipids other than cardiolipin. Although these molecules can increase the diagnostic yield of APS, their clinical relevance is still debatable and needs to be confirmed by interlaboratory efforts toward standardizing diagnostic tools, in addition to experimental data and larger longitudinal studies.

Diffuse Alveolar Hemorrhage and Libman-Sacks Endocarditis as a Manifestation of Possible Primary Antiphospholipid Syndrome

Antiphospholipid syndrome (APS) is an autoimmune disease characterized by the presence of circulating autoantibodies against phospholipid-binding plasma proteins, leading to an increased risk of thrombosis and pregnancy loss. The most common manifestation of lung disease in APS is pulmonary embolism, which may often be the presenting symptom. We present a 30-year-old man with probable primary APS (with no history of thromboses) presenting with diffuse alveolar hemorrhage, an uncommon presentation. He was also found to have severe mitral valve regurgitation and during valve replacement surgery had cardiac vegetations compatible with a presentation of Libman-Sacks endocarditis. There are only 21 other reported cases of diffuse alveolar hemorrhage occurring as a result of APS. This is the first case of Libman-Sacks endocarditis in the setting of probable APS and alveolar hemorrhage.Diffuse alveolar hemorrhage should be considered as a nonthrombotic manifestation of APS, even in the absence of known thromboses, and may be the presenting symptom.

Pathophysiology of Thrombosis and Potential Targeted Therapies in Antiphospholipid Syndrome

The antiphospholipid syndrome (APS) is an autoimmune disease in which recurrent vascular thrombosis, pregnancy morbidity or a combination of these events is associated with the persistent presence of circulating antiphospholipid antibodies (aPL). Evidence shows that the dominant antigenic targets for aPL in APS are phospholipid-binding plasma proteins such as b2glycoprotein I and prothrombin. The pathogenic role of aPL in thrombosis is widely accepted but the mechanisms by which these antibodies mediate disease are only partially understood. aPL may affect the normal procoagulant and anticoagulant reactions occurring on cell surface, and also may interact with certain cells, altering the expression and secretion of procoagulant substances. The intracellular signal transduction triggered by aPL has been a focus of intensive research and the p38 mitogen activated protein kinase (MAPK) pathway has been revealed as a major player in the aPL-mediated cell activation. In addition, some candidates as cell-receptor for phospholipid-binding plasma proteins have been identified. The recognition of the intracellular signaling triggered by aPL is a step forward in the design of new modalities of targeted therapies for thrombosis in APS including specific inhibitors of MAPK pathway or antagonists of the putative receptors. Furthermore, novel findings regarding the role of aPL in T-cells responses mark new advances in the understanding of the immunological reactions in APS and open new insights into possible therapeutic approaches to APS. In this article, we review the pathophysiological mechanisms of thrombosis and the specific new targeted therapies for the treatment in APS.

Anti-Prothrombin Antibodies are Associated with Adverse Pregnancy Outcome

Women with antiphospholipid antibodies (aPL) such as lupus anticoagulant, anticardiolipin antibodies, and anti-β(2) glycoprotein-1 antibodies are at high risk of late pregnancy complications, such as severe pre-eclampsia, placental insufficiency, and fetal loss. It has been observed that aPL consists of a heterogeneous group of antibodies targeting several phospholipid-binding plasma proteins, including also anti-prothrombin (anti-PT), anti-protein S (anti-PS), and anti-protein C (anti-PC) antibodies. Their potential role in late pregnancy complications is not known. The aim of this work was to investigate the association between those autoantibodies and histories for adverse pregnancy outcome. Anti-PT, anti-PS, and anti-PC antibodies were evaluated in 163 patients with previous severe pre-eclampsia, fetal death, and/or placental abruption and in as many women with previous uneventful pregnancies, negative for aPL. The prevalence of anti-PT antibodies was higher in cases than in controls (OR, 95% CI: 10.92, 4.52-26.38). The highest prevalence was observed in subjects with fetal death. Anti-PT antibodies appear to be associated with adverse pregnancy outcome, irrespectively of aPL.

A novel mechanism of thrombosis in antiphospholipid antibody syndrome

Antiphospholipid antibody syndrome (APS) is an autoimmune thrombophilia mediated by autoantibodies directed against phospholipid-binding plasma proteins, mainly β2 Glycoprotein I (β2GPI)-a plasma apolipoprotein and prothrombin (PT). A subgroup of these antibodies termed “Lupus Anticoagulant” (LA) elongate in vitro the clotting times, this elongation not corrected by adding normal plasma in the detection system. The exact mechanism by which these autoantibodies induce thrombosis is not well understood. Resistance to natural anticoagulants such as protein C, impaired fibrinolysis, activation of endothelial cells to a pro-coagulant phenotype and activation of platelets, are among the mechanisms partially supported by experimental evidence. Artificially dimerized β2GPI binds tightly to platelet membrane activating them. We search for mechanisms of natural dimerization of β2GPI by proteins of the platelet membranes and found that platelet factor 4 (PF4) assembled in homotetramers binds two molecules of β2GPI and this complex is recognized by anti-β2GPI antibodies, the whole complexes being thrombogenic in terms of activating platelets as confirmed by p38MAP kinase phosphorylation and thromboxane B2 production. Of note PF4/heparin complexes are also immunogenic triggering the production of anti-PF4/heparin antibodies which activate also platelets (the so-called “heparin-induced thrombocytopenia and thrombosis syndrome”, HITT). The anti-β2GPI antibodies activate platelets by their F(ab)2, while the anti-PF4/heparin by their Fc fragments. Thus PF4 is a common denominator in the pathogenesis of APS and HITT which share also clinical characteristics such as thrombocytopenia and thrombosis.

Association between periodontitis and anti‐cardiolipin antibodies in Buerger disease

Anti-cardiolipin (CL) antibodies can be induced in Buerger disease (BD), an inflammatory occlusive disorder affecting peripheral blood vessels, in response to bacteria bearing homology to the TLRVYK peptide of a phospholipid-binding plasma protein beta-2-glycoprotein I. TLRVYK homologies are present in Porphyromonas gingivalis (TLRIYT) and Treponema denticola (TLALYK). This study investigated the association between periodontal infection and anti-CL antibodies in BD patients. Periodontal conditions were examined in 19 BD patients and 25 systemically healthy control subjects. All subjects were heavy smokers. Serum anti-CL, anti-TLRVYK, anti-TLRIYT, and anti-TLALYK antibodies were assessed using the enzyme-linked immunosorbent assay. BD patients had a significantly higher prevalence of periodontitis, more severe periodontal destruction and increased titres of serum anti-CL, anti-TLRVYK, anti-TLRIYT, and anti-TLALYK antibodies compared with healthy subjects. The levels of anti-CL antibodies positively correlated with those of the three anti-peptide antibodies. Anti-CL antibody titres were significantly associated with the percentage of sites with clinical attachment level >or=4 mm in BD patients. Elevated anti-CL antibody levels were associated with periodontal destruction in BD patients. Periodontopathic bacteria may serve as exogenous antigens that stimulate the anti-CL antibody production through molecular mimicry between the bacterial peptides and a host plasma protein.

New approaches for managing antiphospholipid syndrome

Antiphospholipid antibodies (aPL) are a family of autoantibodies directed against phospholipid-binding plasma proteins, most commonly beta(2)-glycoprotein-I. Primary thrombosis prevention in persistently aPL-positive individuals requires a risk-stratified approach; elimination of reversible thrombosis risk factors and aggressive prophylaxis during high-risk periods are crucial. The effectiveness of aspirin in persistently aPL-positive patients without vascular involvement is not supported by data from prospective, controlled studies. For the secondary prevention of thrombosis in persistently aPL-positive individuals, the current recommendation is life-long warfarin; however, determining the intensity and duration of warfarin treatment, as well as the role of alternative anticoagulants, requires further research. The effectiveness of high-intensity anticoagulation in patients with antiphospholipid syndrome (APS) and vascular involvement is not supported by data from prospective, controlled studies. Patients with catastrophic APS usually receive a combination of anticoagulants, corticosteroids, intravenous immunoglobulin and plasma exchange; however, despite this aggressive approach, the mortality rate remains high. Potential new approaches for the management of persistently aPL-positive patients include hydroxychloroquine, statins, rituximab, complement inhibition, and other targeted therapies that have been effective in experimental APS models.

Venous Thrombosis in the Antiphospholipid Syndrome

The antiphospholipid syndrome is a relatively common acquired cause of venous thrombosis. Up to 20% of cases of deep vein thrombosis, with and without pulmonary embolism, may be associated with antiphospholipid antibodies. These antibodies are typically detected in lupus anticoagulant assays and tests for anticardiolipin antibodies. Most antiphospholipid antibodies are directed against several phospholipid-binding plasma proteins. The most common antigens are beta2-glycoprotein I and prothrombin. Immunoassays using these purified antigens are now available. In addition to being markers for thrombotic risk, antiphospholipid antibodies have been shown to directly contribute to hypercoagulability in animal models and in various in vitro studies. Prevention of recurrent venous thrombosis in patients with the antiphospholipid syndrome requires long-term anticoagulation. The optimal intensity of warfarin therapy is an ongoing issue, but most clinicians currently favor a target INR in the 2.0 to 3.0 range. In certain patients, antiphospholipid antibodies may interfere with determination of the INR, requiring other approaches to monitor and adjust the warfarin dose. Low-dose aspirin is typically recommended for primary prevention of thrombosis in asymptomatic patients with moderate to high levels of antiphospholipid antibodies, although strong supporting data are lacking.

Annexin A5 binding to giant phospholipid vesicles is differentially affected by anti- 2-glycoprotein I and anti-annexin A5 antibodies

Anti-phospholipid antibodies have been recognized to play a role in vascular thrombosis and pregnancy morbidity. They were first thought to be directed to phospholipids, but it is now known that the majority of pathogenic antibodies recognizes epitopes on phospholipid-binding plasma proteins such as beta2-glycoprotein I (beta2GPI) or possibly also annexin A5 (ANXA5). The mechanism of their prothrombotic action is still not completely understood. The aim of the present study was to observe the effect of antibodies against ANXA5 (aANXA5) and antibodies against beta2GPI (abeta2GPI) on the binding of ANXA5 to the negatively charged phospholipid membrane. Giant phospholipid vesicles (GPVs) were used as a simple model of the membrane surface. GPVs composed of phosphatidylserine and phosphatidylcholine were produced in an aqueous medium. A single GPV was transferred to the solution containing ANXA5 conjugated with Alexa Fluor 488 (FANXA5) and (i) aANXA5 or abeta2GPI and (ii) different concentrations of abeta2GPI together with beta2GPI. The emission of the fluorescent light from the GPV surface, as the result of FANXA5 binding, was measured. Beta2GPI together with abeta2GPI reduced the binding of FANXA5 to GPVs. On the contrary, aANXA5 enhanced the binding of ANXA5 to the GPV surface. Our results point to the competition between FANXA5 and complexes of beta2GPI-abeta2GPI for the same binding sites and therefore support the hypothesis of the disruption of the ANXA5 protective shield on procoagulant phospholipid surface. The influence of increased cell surface ANXA5 concentration in the presence of aANXA5 on coagulation needs to be further studied.

Antiphospholipid-Mediated Thrombosis: Interplay Between Anticardiolipin Antibodies and Vascular Cells

The antiphospholipid syndrome (APS) is characterized by thrombosis or pregnancy morbidity in the presence of antiphospholipid autoantibodies (aPL). aPL are a heterogeneous family of autoantibodies with diverse cross-reactivities whose origin and role have not been fully elucidated. Many of the autoantibodies associated with APS are directed against phospholipid-binding plasma proteins, such as beta2-GPI and prothrombin, or phospholipid-protein complexes. The mechanisms by which aPL cause thrombosis are not completely understood. There is no unique mechanism able to explain all symptoms associated with the presence of aPL. Different theories have been proposed, including the effect of aPL on endothelial cells, monocytes, and platelets. aPL are able to recognize, injure, or activate cultured vascular endothelial cells. Cultured endothelial cells incubated with aPL express increased levels of cell adhesion molecules and tissue factor (TF), an effect mediated by beta2-GPI, and may promote inflammation and thrombosis. Overexpression of TF has been also shown in monocytes in vitro and ex vivo. TF is the major initiator of coagulation in vivo; thus, its dysregulation may be one of the most important contributors to thrombosis. Effects of aPL upon platelets are not completely elucidated. aPL bind anionic phospholipid but they are normally in the inner side of cell membranes. When platelets are activated by different agonists, anionic phospholipids are exposed. There is some evidence showing that activated platelets are present in aPL-positive patients. Increased levels of beta-thrombomodulin, and microvesicle formation seem to support this hypothesis. Activated platelets may contribute to thrombosis by persistent exposure of a procoagulant surface.

Antigen Specificity and Clinical Relevance of Antiphospholipid Syndrome- Related Autoantibodies

The presence of antiphospholipid antibodies (aPL) combined with venous or arterial thrombosis, and/or obstetric complications defines the antiphospholipid syndrome (APS). Lupus anticoagulants (LA) and anticardiolipin antibodies (aCL) were the first described aPL. It has been shown that aPL, despite their name, are not directed against anionic phospholipids, as had previously thought, but are a part of a large family of autoantibodies against phospholipid-binding plasma proteins. The most important and relevant antigenic targets involved in APS are β2 glycoprotein I (β2GPI) and prothrombin. However, an increasing number of other phospholipid-binding proteins with crucial functions in the regulation of blood coagulation and fibrinolysis are also targeted by APS-related autoantibodies. For clinical purposes, it is important to find which aPL markers has the best prognostic value. While it has been clearly demonstrated that LA and, to a lesser extent, aCL represent a risk for thrombosis, the role of antibodies directed against β2GPI, prothrombin as well as other targets is still uncertain and controversial. Data from a prospective study showed the presence of antibodies to prothrombin and/or β2GPI is a predictor of first or recurrent thromboembolic events in patients with LA and/or aCL. This review intends to highlight the antigen specificity and clinical relevance of APSrelated aPL. Keywords: antiphospholipid antibodies, antiphospholipid syndrome, glycoprotein I, pregnancy loss, prothrombin, thrombosis

Prevalence and Clinical Correlation of Anti-Phospholipid–Binding Protein Antibodies in Anticardiolipin-Negative Patients With Systemic Lupus Erythematosus and Women With Unexplained Recurrent Miscarriages

Anti-phospholipid antibodies (aPL) are a heterogeneous group of autoantibodies, the presence of which is associated with thrombotic events and miscarriage. To establish whether antibodies directed against phospholipid-binding plasma proteins such as beta(2)-glycoprotein I (beta(2)GPI), prothrombin (PT), and annexin V (Anx V) constitute a risk factor for thromboembolism in patients with systemic lupus erythematosus (SLE) and for miscarriage in women with recurrent pregnancy loss (RPL), independently of the presence of the classic anticardiolipin (aCL) antibodies, and whether their determination together with that of aCL would help to increase the diagnostic sensitivity of aPL tests. The prevalence of various antibodies directed toward phospholipids (CL and other anionic phospholipids [APL]) and phospholipid-binding proteins (beta(2)GPI, PT, and Anx V) was determined by immunoenzymatic methods in 311 serum samples. Twenty-five patients with aCL-positive primary anti-phospholipid syndrome (pAPS); 89 patients with SLE, 23 of whom had thrombotic complications (SLE/APS) and 66 of whom had no thrombosis; and 77 women with unexplained recurrent pregnancy loss comprised our study group. One hundred twenty healthy subjects matched for age and sex were studied as the control group. Immunoglobulin (Ig) G and/or IgM aAPL, anti-beta(2)GPI, anti-PT, and IgG anti-Anx V antibodies were detected in 25 (100%), 20 (80%), 15 (60%), and 6 (24%), respectively, of the 25 aCL-positive pAPS patients; IgG and/or IgM aCL, aAPL, anti-beta(2)GPI, anti-PT, and IgG anti-Anx V antibodies were detected in 33 (37%), 42 (47%), 31 (35%), 40 (45%), and 12 (13%) of the 89 SLE patients, respectively. Of the 56 SLE patients who proved to be aCL negative, anti-beta(2)GPI was present in 3 patients (5%), anti-PT in 13 (23%) patients, and anti-Anx V in 5 (9%) patients. In the subset of 23 SLE/APS patients, IgG anti-PT prevalence was higher than that of the other autoantibodies (87% vs 70% aCL, 66% aAPL, 57% anti-beta(2)GPI, and 4% anti-Anx V), and in 26% of cases, IgG anti-PT was the only antibody present. Anti-PT had a slightly lower specificity than aCL (46% vs 49%); however, the occurrence of both antibodies brought the specificity to 92.4%. The highest risk for thrombosis in SLE patients was associated with the presence of IgG anti-PT antibody (odds ratio [OR] 15.3, P < .001, vs 6.5 aCL, 3.5 aAPL, 3.4 anti-beta(2)GPI, 0.2 anti-Anx V). Fifty-one of the 77 women with recurrent pregnancy loss were negative for all antibodies investigated; the prevalence of IgG and/or IgM aCL, aAPL, anti-beta(2)GPI, anti-PT, and IgG anti-Anx V antibodies was 6% (5), 12% (9), 6% (5), 16% (12), and 17% (13), respectively. Of the 67 aCL-negative women, none had anti-beta(2)GPI antibodies, 7 (11%) were anti-PT positive, and 13 (19%) were anti-Anx V positive. In the subgroup of 26 recurrent pregnancy loss patients who had at least one antibody, anti-Anx V was present in 50% of cases (in 42% as the sole antibody) and was the only antibody significantly associated with miscarriage (P = .02). The results of this study indicate that it is useful to measure anti-PT antibodies in addition to the more widely used aCL and anti-beta(2)GPI antibodies in the prognostic evaluation of SLE patients for the risk of thrombosis, and the results also confirm that anti-Anx V antibodies may play an important role in recurrent pregnancy loss.

Atherogenic role of protein-modified oxidized low-density lipoproteins and related autoantibodies

Elevated serum levels of antiphospholipid antibodies, venous or arterial thrombosis, and/or pregnancy morbidity (miscarriages, recurrent fetal loss) are the major serologic and clinical features of the antiphospholipid syndrome (APS). Antiphospholipid antibodies belong to a heterogeneous family of autoantibodies that play a crucial role in the development of thrombosis in patients with APS. These antibodies may react with negatively charged phospholipids, phospholipid/protein complexes, and certain plasma proteins attached to suitable surfaces (i.e., activated cell membranes, oxygenated polystyrene). Most antiphospholipid antibodies derived from patients with APS require the presence of certain plasma proteins for optimal phospholipid binding activity. β2-Glycoprotein I (β2GPI), a phospholipid-binding plasma protein, is now recognized as the most clinically relevant antigenic target for antiphospholipid antibodies. Antibodies to β2GPI are more specific for thrombosis (and APS) than anticardiolipin (aCL) antibodies. Recent prospective studies have shown that β2GPI-dependent aCL and anti-β2GPI antibodies were significant predictors of arterial thrombosis (myocardial infarction and stroke). In addition to natural anticoagulant properties, β2GPI may also bind to oxidized low-density lipoprotein (oxLDL) likely to quench its proinflammatory and atherogenic effects. Circulating oxLDL/β2GPI complexes are immunogenic, and anti-oxLDL/β2GPI antibodies accelerate their macrophage uptake and the development of autoimmune-mediated atherothrombosis. Autoantibodies against oxLDL/β2GPI complexes strongly correlated with arterial thrombosis in patients with systemic lupus erythematosus (SLE) and APS. These findings suggest that β2GPI and anti-β2GPI antibodies play a central pathogenic role in thrombosis, and particularly in autoimmune-mediated atherosclerosis.