Thrombosis In Antiphospholipid Antibody Syndrome Research Articles

Platelets and Thrombotic Antiphospholipid Syndrome

Antiphospholipid antibody syndrome (APS) is an autoimmune disorder characterised by thrombosis and the presence of antiphospholipid antibodies (aPL): lupus anticoagulant and/or IgG/IgM anti-β2-glycoprotein I and anticardiolipin antibodies. APS carries significant morbidity for a relatively young patient population from recurrent thrombosis in any vascular bed (arterial, venous, or microvascular), often despite current standard of care, which is anticoagulation with vitamin K antagonists (VKA). Platelets have established roles in thrombosis at any site, and platelet hyperreactivity is clearly demonstrated in the pathophysiology of APS. Together with excess thrombin generation, platelet activation and aggregation are the common end result of all the pathophysiological pathways leading to thrombosis in APS. However, antiplatelet therapies play little role in APS, reserved as a possible option of low dose aspirin in addition to VKA in arterial or refractory thrombosis. This review outlines the current evidence and mechanisms for excessive platelet activation in APS, how it plays a central role in APS-related thrombosis, what evidence for antiplatelets is available in clinical outcomes studies, and potential future avenues to define how to target platelet hyperreactivity better with minimal impact on haemostasis.

The Lupus Anticoagulant Titer Is Associated with Elevated Anti-Phosphatidyl-Serine/Prothrombin Antibody Levels

Background Laboratory diagnosis of the antiphospholipid antibody syndrome (APS) requires evidence of persistently positive lupus anticoagulant (LAC) activity in clot-based assays and/or the presence of medium-high titer anticardiolipin (aCL) and/or anti-beta 2 glycoprotein I (aB2GPI) antibodies. Although aCL and aB2GPI methodologies provide quantitative information about the antibody titer, LAC testing is interpreted in a qualitative fashion (presence versus absence). Objectives In this study, we describe a method for measuring the LAC titer and evaluate the relationship between the LAC titer, other antiphospholipid antibodies and clinical outcomes. Methods Residual plasma from 51 LAC-positive samples (50 unique patients) were included in this study. To estimate LAC-titers, samples were tested undiluted and subjected to serial (dependent) twofold dilutions in pooled normal plasma (PrecisionBiologic) up to a final dilution of 1:128, as appropriate. The dilute Russell's viper venom screening time (DRVVT screen, LA Check, PrecisionBiologic) and activated partial thromboplastin screening time (APTT, SynthasIL, Werfen) were measured for each dilution. A four-parameter logistic model of the dilution factor versus the clotting time was used to interpolate the LAC-titer at the upper limit of normal for both assays (APTT of 37s and DRVVT screen ratio of 1.2). Measurements of aCL, aB2GPI and anti-phosphatidylserine/prothrombin antibodies (aPS/PT) were performed using ELISA methodologies (QUANTA Lite, Werfen). Available clinical data were extracted from the electronic medical record. Continuity corrected chi-square test was used as a categorical test of association and quantile regression was used to compare medians for interval/ratio data across subgroups (Stata v17). Results There was a strong positive linear relationship between the APTT and DRVVT titers (Pearson's correlation coefficient, r=0.92, Figure 1A). There was only a moderate relationship between the APTT titer and the APTT screen in the undiluted sample (r=0.50) and almost no relationship between the DRVVT titer and the DRVVT screen ratio or DRVVT screen/confirm ratio in the undiluted sample (r=0.06 and r=0.13, respectively). Data-driven partitioning identified two DRVVT titer groups: low-titer (<10) and high-titer (≥10). Empirical cutpoint estimation (Youden) identified an APTT titer threshold of 5 for predicting the DRVVT titer group. Among patients with available clinical history, 15 of 25 (60%) low-DRVVT-titer and 16 of 20 (80%) high-DRVVT-titer patients had thromboembolic/obstetric events (p=0.26). Autoimmune diagnoses were more prevalent in the low-DRVVT-titer group (19 of 25 patients, 13 with SLE) than the high-DRVVT-titer group (8 of 20 patients, 3 with SLE, p=0.03). Comparing the low and high-DRVVT-titer groups, median aCL IgM (p=0.27), aCL IgG (p=0.99), aB2GPI IgM (p=0.72) and aB2GPI IgG (p=1.00) titers were not different. High-DRVVT-titer samples had a significantly higher median aPS/PT IgM compared to low-DRVVT-titer samples (p <0.01, Figure 1B) but there was no difference observed for IgG aPS/PT (p=0.57, Figure 1B). Conclusions APTT and DRVVT screening times are not strongly correlated with the corresponding APTT or DRVVT LAC-titer and may not indicate LAC potency. High DRVVT LAC-titers were strongly associated with IgM aPS/PT levels, but not with aCL, aB2GPI or IgG aPS/PT levels. Further research is required to evaluate the association between the LAC-titer and recurrent thrombosis in APS. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

A targetable pathway in neutrophils mitigates both arterial and venous thrombosis.

Arterial and venous thrombosis constitutes a major source of morbidity and mortality worldwide. Long considered as distinct entities, accumulating evidence indicates that arterial and venous thrombosis can occur in the same populations, suggesting that common mechanisms are likely operative. Although hyperactivation of the immune system is a common forerunner to the genesis of thrombotic events in both vascular systems, the key molecular control points remain poorly understood. Consequently, antithrombotic therapies targeting the immune system for therapeutics gain are lacking. Here, we show that neutrophils are key effectors of both arterial and venous thrombosis and can be targeted through immunoregulatory nanoparticles. Using antiphospholipid antibody syndrome (APS) as a model for arterial and venous thrombosis, we identified the transcription factor Krüppel-like factor 2 (KLF2) as a key regulator of neutrophil activation. Upon activation through genetic loss of KLF2 or administration of antiphospholipid antibodies, neutrophils clustered P-selectin glycoprotein ligand 1 (PSGL-1) by cortical actin remodeling, thereby increasing adhesion potential at sites of thrombosis. Targeting clustered PSGL-1 using nanoparticles attenuated neutrophil-mediated thrombosis in APS and KLF2 knockout models, illustrating the importance and feasibility of targeting activated neutrophils to prevent pathological thrombosis. Together, our results demonstrate a role for activated neutrophils in both arterial and venous thrombosis and identify key molecular events that serve as potential targets for therapeutics against diverse causes of immunothrombosis.

Antiphospholipid antibodies and vitamin D deficiency in COVID-19 infection with and without venous or arterial thrombosis: A pilot case-control study.

Coronavirus disease-2019 (COVID-19) is associated with thromboembolism. Antiphospholipid antibody (APLa) formation is one of the mechanisms. Vitamin D deficiency has been associated with thrombosis in antiphospholipid antibody syndrome. Measure APLa and vitamin D in hospitalized COVID-19 patients with and without thrombosis to evaluate if thromboembolism is associated with concomitant APLa and vitamin D deficiency. Case-control study. Hospitalized COVID-19 patients with a thromboembolic event (ischemic stroke, myocardial infarction, deep venous thrombosis/pulmonary embolism, Cases n = 20). Controls (n = 20): Age, sex-matched without thromboembolic events. Patients with autoimmune disorders, antiphospholipid antibody syndrome, thrombophilia, anticoagulation therapy, prior thromboembolism, chronic kidney disease 3b, 4, end-stage renal disease, and malignancy were excluded. Given the limited current literature on the role of concomitant antiphospholipid antibodies and vitamin D deficiency in causing venous and/or arterial thrombosis in hospitalized COVID-19 patients, we enrolled 20 patients in each arm. Anti-cardiolipin IgG/IgM, beta-2 glycoprotein-1 IgG/IgM, lupus anticoagulant and vitamin D levels were measured in both groups. Cases were 5.7 times more likely to be vitamin D deficient (OR:5.7, 95% CI:1.3-25.6) and 7.4 times more likely to have any one APLa (OR:7.4, 95% CI: 1.6-49.5) while accounting for the effects of sex. Patients with both APLa and vitamin D deficiency had significantly more thrombosis compared to patients who were antibody positive without vitamin D deficiency (100% vs 47.4%; p = 0.01). Thrombosis in COVID-19 was associated with concomitant APLa and vitamin D deficiency. Future studies in COVID-19 should assess the role of vitamin D in reducing thrombosis.

A rare case of ulnar and popliteal artery thrombosis in antiphospholipid antibody syndrome

We report an extremely rare case of medium vessel thrombosis (ulnar and popliteal artery) in a 26-year-old female presenting with recurrent abortion and gangrenous right ring finger and pregangrenous toes of the left feet. Medium vessel thrombosis i.e. selective involvement of ulnar and popliteal vessels is an extremely rare phenomenon described in the literature of antiphospholipid antibody (APLA) syndrome. We conservatively managed the patient with antiplatelets and anticoagulants which substantially improved the ischemia of the finger and toes. Our case is a unique description of ulnar and popliteal artery thrombosis in APLA syndrome which has not been described in world literature so far.

Nanoparticle-Directed Targeting of Clustered PSGL-1 Mitigates Neutrophil-Derived Thrombosis in Antiphospholipid Antibody Syndrome

Multiorgan thromboses and subsequent organ failure are among the more devastating consequences of antiphospholipid antibody syndrome (APS). APS poses a relatively unique risk to patients by virtue of its propensity to cause thrombi of both the arterial and venous vascular systems. While recent studies have explored mechanisms of thrombosis in singular vascular beds, there remains a dearth of targetable effectors of arteriovenous thrombosis in APS. Here, we examine the potential of neutrophils as key players of both arterial and venous thrombosis in APS. From this and previous studies, we have also identified a critical transcriptional regulator in neutrophils whose loss mimics arterial and venous thrombosis susceptibility seen in APS. This factor, Krüppel-like factor 2 (KLF2), is down regulated in APS patients as well as in cells treated with antiphospholipid antibody (APLA) ex vivo. Loss of KLF2 in neutrophils contributes to classic neutrophil activation phenotypes including NETosis, increased tissue factor release, and heightened chemotaxis; remarkably, blocking each of these functions greatly abrogates the thrombotic phenotype. Utilizing the myeloid KLF2 knockout (K2KO) system as a model for aberrant neutrophil activation seen in APS, we have identified clustering of P-selectin glycoprotein ligand 1 (PSGL-1) secondary to actin rearrangement to be operative in neutrophil-mediated thrombosis in arterial and venous systems. While neutralizing antibodies against PSGL-1 have been explored as therapeutic options in APS, there remains a challenge in establishing local specificity to activated immune cells in order to increase efficacy. To circumvent this, we have designed a nanoparticle (NP) based approach that utilizes NPs decorated with anti-PSGL-1 immunoglobulins to better target clustered PSGL-1 on activated neutrophils. PSGL-1 NPs are more effective at inhibiting neutrophil rolling and adhesion in in vitro assays while, incubating activated neutrophils in PSGL-1 NPs prior to infusion attenuates their ability to contribute to both arterial and venous thrombosis. Further, injection of PSGL-1 NPs directly into K2KO mice reduces their extensive thrombotic phenotype. Importantly, this effect occurs at a substantially lower dose of PSGL-1 NPs than that which would be needed of PSGL-1 antibody alone. Finally, PSGL-1 NPs greatly reduce thrombotic burden in mice injected with APLA, diminishing clotting in both arteries and veins. Together, this work identifies key molecular and cellular players in APLA-induced arteriovenous thrombosis. Furthermore, it provides early pre-clinical evidence at specifically targeting clustered PSGL-1 on neutrophils as a viable option in combatting thrombosis in this particularly vulnerable patient population. Disclosures No relevant conflicts of interest to declare.

Simultaneous Occurrence of Deep Vein Thrombosis and Carotid Artery Thrombosis in Antiphospholipid Antibody Syndrome.

Simultaneous Occurrence of Deep Vein Thrombosis and Carotid Artery Thrombosis in Antiphospholipid Antibody Syndrome.

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.

Role of tissue factor in thrombosis in antiphospholipid antibody syndrome

Antiphospholipid syndrome (APS) is an acquired autoimmune disorder defined by the presence of an antiphospholipid antibody (aPL) and the occurrence of at least one associated clinical condition that includes venous thrombosis, arterial thrombosis or pregnancy morbidity. The aPL detected in APS have long been thought to have a direct prothrombotic effect in vivo. However, the pathophysiology underlying their coagulopathic effect has not been defined. Emerging data suggest a role for the procoagulant protein tissue factor (TF). In this review we provide an overview of TF, describe mouse models used in the evaluation of the role of TF in thrombosis, as well as summarize recent work on TF and APS.

Role of tissue factor in thrombosis in antiphospholipid antibody syndrome

Role of tissue factor in thrombosis in antiphospholipid antibody syndrome

Rituximab Effectively Reverses Papilledema Associated with Cerebral Venous Sinus Thrombosis in Antiphospholipid Antibody Syndrome

A case of bilateral papilledema secondary to cerebral venous sinus thrombosis treated with Rituximab, an anti-CD20 monoclonal antibody. A 23 year old obese female with a one week history of blurred vision, headaches and vomiting presented with bilateral papilledema. Her BCVA was 20/50 in right eye and 20/200 in the left eye with severe reduction of visual fields. Laboratory investigations revealed thrombocytopenia, prolonged prothrombin time (not reversed when mixed with normal plasma) and anticardiolipin antibodies. Besides, cerebral angiogram showed presence of cerebral venous thrombosis. The patient to have anti phospholipid antibody syndrome and treated with rituximab I.V. 375 mg/m 2 weekly x 4 doses, acetazolamide 500 mg BID, methyl prednisolone I. V. QID x 4. At 1-month, her headaches and vision improved to 20/30 in both eyes with partial resolution of papilledema and complete restoration of visual fields. Nine months later, patient had 20/25 vision in right eye and 20/30 in left eye with complete resolution of papilledema and cerebral sinus thrombosis. Rituximab was effective in reversing papilledema and cerebral sinus thrombosis, while preserving the vision in patient with antiphospholipid antibody syndrome. It is efficacious in treating papilledema in patients refractory to treatment with systemic steroids and immunoglobulin, with better clinical compliance and no side effects.

Evidence-based management of thrombosis in the antiphospholipid antibody syndrome.

Recommendations for the management of thrombosis in the antiphospholipid antibody syndrome have been based largely on retrospective case series. Several clinical trials have been recently published on the management of thrombosis in general. Two clinical trials have specifically addressed thrombosis in antiphospholipid antibody syndrome, and one is underway. These new clinical trials have challenged the previous dogma of a target International Normalized Ratio of 3 to 4 ("high-intensity" warfarin).

Identification of an Fc? receptor-independent mechanism by which intravenous immunoglobulin ameliorates antiphospholipid antibody-induced thrombogenic phenotype

Patients with the antiphospholipid antibody syndrome (APS) often experience recurrent arterial and venous thrombosis and pregnancy losses. Intravenous immunoglobulin (IVIG) therapy has prevented pregnancy loss in some women with APS and has reversed fetal resorption rates in murine models of pregnancy loss. Although the basis for these effects is unknown, effector mechanisms of pathogenic antibodies often involve receptors for IgG (Fc gamma receptors [Fc gammaR]). We examined the potential mechanisms of action of WIG in an in vivo murine model of antiphospholipid antibody (aPL)-induced thrombosis and endothelial cell activation. Mice infused with IgG containing human anticardiolipin antibodies (aCL) were treated with IVIG (36 microg i.v.), saline, or ovalbumin. Surgically induced thrombus formation and in vivo leukocyte adhesion to endothelial cells were measured. Circulating levels of aCL were measured by enzyme-linked immunosorbent assay. To determine whether Fc gammaR are required for the effects of IVIG, we treated mice deficient in stimulatory Fc gammaR. To examine the effects of IVIG on endogenously generated antibody, we treated mice immunized with beta2-glycoprotein I (beta2GPI). IVIG treatment inhibited aPL-induced endothelial cell activation and enhancement of thrombosis in mice passively infused with human aPL-containing IgG, and this was associated with a decrease in aPL levels. Similarly, IVIG lowered aPL levels and inhibited thrombogenesis in mice immunized with beta2GPI. The thrombophilic effects of aPL were evident in Fc gammaR-deficient mice. Treatment with IVIG inhibits the thrombogenic effects of aPL in vivo and reduces the levels of aCL in the circulation. Blockade of stimulatory Fc gammaR on inflammatory cells is not necessary for this effect. The mechanism of action of IVIG is more likely saturation of the IgG transport receptor, leading to accelerated catabolism of pathogenic aPL. These results have implications in the management of thrombosis in APS and may have applications for pregnant patients with a history of APS.

Isolation and purification of anticardiolipin antibody from plasma of a patient with antiphospholipid syndrome: induced generation of platelet thromboxane A 2 synthesis

Antiphospholipid antibodies, particularly anticardiolipin antibodies (aCL) are autoantibodies frequently detected in the serum of patients with systemic lupus erythematosus (SLE) and the primary antiphospholipid antibody syndrome (PAPS). These patients commonly suffer from thrombosis, recurrent fetal loss and thrombocytopenia. Since platelet aggregation is pivotal in the genesis of thrombosis, we tested the hypothesis that perturbation of platelet membrane by aCL β 2- glycoprotein ( aCL β 2 GP ) complex could trigger the biosynthesis of TXA 2, a proaggregatory metabolite of AA. The preincubation of 14C-arachidonic acid ( 14C-AA)-labeled platelet pellets ( 14C-PP) from normal individuals with aCL alone followed by incubation with thrombin, resulted in a moderate increase in platelet thromboxane B 2 ( 14C-TXB 2) biosynthesis when compared to controls (without aCL). Similar incubations with β 2GP-I alone resulted in negligible 14C-TXB 2 biosynthesis. In contrast, the preincubations of normal 14C-PP with aCL β 2 GP-I complex resulted in marked thrombin-induced TXB 2 biosynthesis, underscoring the requirement of β 2GP-I in aCL-induced platelet TXB 2 biosynthesis. Taken together, these results are consistent with the view that aCL β 2 GP-I platelet interactions do play a role, at least in part, in platelet hyperactivity and thrombosis in antiphospholipid antibody syndrome.