Pathophysiology of the antiphospholipid syndrome: roles of anticardiolipin antibodies in thrombosis and fibrinolysis

Antiphospholipid antibodies (aPL) (anticardiolipin antibodies and lupus anticoagulant) are associated with thrombosis and pregnancy morbidity, the antiphospholipid syndrome (APS). Despite the clear association between aPL and those manifestations, the precise underlying disease mechanisms remain unclear. APL may affect the normal procoagulant and anticoagulant reactions occurring on cell membranes, and also may interact with certain cells, altering the expression and secretion of procoagulant substances. In this article, we review the immunological characteristics of anticardiolipin antibodies and their potential effects on the coagulation and fibrinolytic systems implicated in the development of thrombotic complications in patients with APS.     

Adrenal involvement in the antiphospholipid syndrome

Although Addison disease and hypoadrenalism are rare in patients with systemic lupus erythematosus (SLE), early reports of the association suggested the presence of antiphospholipid antibodies (aPL) in these patients. Data from literature reveal that adrenal failure was present in between 10 and 26% of patients with catastrophic APS and that one-third of patients presented with adrenal involvement during the course of catastrophic APS. Adrenal involvement may be the first clinical manifestation of this syndrome, whereas a few patients may have a history of Addison's disease in the past. The pathological mechanisms involved in the production of adrenal insufficiency in APS are still not clearly understood, but the hypercoagulable state in these patients supports the concept that adrenal haemorrhagic infarction may possibly be related to adrenal vein thrombosis. In the present article,we review the pathogenic mechanisms and main clinical, laboratory and treatment features of patients suffering adrenal involvement with aPL to support the idea that APS leads to the development of adrenal insufficiency.     

Pathogenic role of antiphospholipid antibodies

The antiphospholipid antibody syndrome (APS) is characterized by recurrent arterial and venous thrombosis and/or pregnancy in association with antiphospholipid (aPL) antibodies. The pathogenic mechanisms in APS that lead to in vivo injury are incompletely understood. Recent evidence suggests that APL antibodies alter regulation of haemostasis and induce activation of complement. We will discuss the current knowledge on how aPL antibodies trigger increased inflammation and enhanced thrombotic tendency, and thereby lead to tissue damage.     

Antiphospholipid syndrome, “the best prophet of the future”

The antiphospholipid syndrome (APS) is an autoimmune disorder characterized by the occurrence of venous and arterial thromboses, often multiple, and obstetric-related adverse events in the presence of antiphospholipid antibodies (aPL). APS, first described in 1983, as thrombosis, abortion and cerebral disease, is nowadays recognised as a systemic disease with a wide constellation of clinical manifestations related to acute and chronic vascular lesions. The presence of aPL is the serological hallmark of APS representing a heterogeneous population of autoantibodies with many antigenic specificities directed to phospholipid-binding proteins, either alone or in combination with phospholipids. Many assays have been developed for aPL detection. Particularly, anticardiolipin antibodies, anti-β2-glycoprotein I antibodies and lupus anticoagulant are essential tools for APS diagnosis. The cumulative evidence indicates that aPL are pathogenic autoantibodies binding to target cells and promoting thrombosis and pregnancy complications through a wide range of pathological mechanisms not yet fully understood. Finally, the recognition of the important role of aPL to assess the individual risk of thrombosis or pregnancy complications has expanded the concept of aPL, and currently aPL profile is regarded as a major risk factor for clinical thrombotic events.     

Placentation, antiphospholipid syndrome and pregnancy outcome

The antiphospholipid antibodies (aPLs) are a diverse group of autoantibodies associated with a pattern of disease known as antiphospholipid syndrome (APS). Pregnancy complications secondary to placental insufficiency are key features of this disease. The mechanisms underlying the placental pathology remain unclear. In this article the process of placentation in healthy and pathological pregnancies is reviewed. The evidence for defective placentation in APS pregnancies and involvement of aPLs in this process is summarized. Finally hypotheses based on the interpretation of these studies are discussed.     

What have we learned about antiphospholipid syndrome from patients and antiphospholipid carrier cohorts?

Venous or arterial thrombosis or pregnancy morbidity in the presence of circulating antiphospholipid antibodies (aPL) define the antiphospholipid syndrome (APS). In terms of accepted APS criteria, aPL are detected by one coagulation test (lupus anticoagulant; LAC) and two immunoassays (anticardiolipin antibodies and anti-β2-glycoptrotein I antibodies). In patients with APS, a single positive test carries a much lower risk of thrombosis recurrence or new pregnancy loss than does multiple (or triple) positivity. The same holds true for aPL carriers, namely subjects with laboratory tests but without clinical criteria for APS. Thus, very different risk categories exist among patients with APS as well as in carriers of aPL. Triple positivity apparently identifies the pathogenic autoantibody (antidomain I-II of β2-glycoptrotein I); it is in this category of patients that trials on new therapeutic strategies should focus.     

Therapeutic strategies in antiphospholipid syndrome

The classical clinical picture of the antiphospholipid syndrome (APS) is characterized by venous or arterial thromboses, fetal losses and thrombocytopenia, in the presence of antiphospholipid antibodies (aPL), namely lupus anticoagulant, anticardiolipin antibodies or antibodies directed to various proteins, mainly β2 glycoprotein I, or all three. Apart from being "primary" (without any discernable underlying systemic autoimmune disease), or associated to another disease (usually to systemic lupus erythematosus), it may also occur rapidly over days or weeks when it has been termed "catastrophic" APS. Therapy should not primarily be directed at effectively reducing the aPL levels and the use of immunotherapy (including high dose steroid administration, immunosuppression or plasma exchange) is generally not indicated, unless in the catastrophic APS. Treatment of APS patients should be based on the use of antiaggregant and anticoagulant therapy.     

Current concepts on the pathogenesis of the antiphospholipid syndrome

The antiphospholipid syndrome (APS) is an important cause of acquired thrombophilia. It is characterized by the core clinical manifestations of thrombosis, either venous or arterial, and in women it can also be associated with recurrent fetal loss. The detection of persistently elevated levels of antiphospholipid antibodies (aPL Abs) is a requisite laboratory feature for the diagnosis to be made. The dominant antigenic targets in APS are beta 2-glycoprotein I (beta2-GPI) and prothrombin. There is an accumulating body of experimental evidence that suggests that specific subgroups of aPL Abs may directly contribute to disease pathogenesis. This review critically examines the experimental evidence underlying the various propositions made to explain how these antibodies may predispose to disease in humans. Furthermore, it also examines the evidence relating to the immunologic mechanisms that may contribute to the breakage of peripheral tolerance in this disorder. Delineating the strengths and limitations of the experimental evidence accumulated thus far will hopefully stimulate further experimentation toward achieving the ultimate goal of precisely defining the dominant pathogenic mechanisms operational in APS. This may pave the way for the development of improved therapies.     

Mechanisms of aPL-mediated thrombosis: Effects of aPL on endothelium and platelets

Antiphospholipid antibodies (aPL) are associated with thrombosis and pregnancy loss in patients with systemic lupus erythematosus and antiphospholipid syndrome. Strong evidence demonstrates that aPL are pathogenic in vivo from studies that utilized animal models of thrombosis, endothelial cell activation, and pregnancy loss. However, the mechanisms by which aPL mediate disease are only partially understood, and our knowledge is limited by the polyspecificity of the antibodies, the multiple potential end-organ targets, and the variability of the clinical context in which the disease may present. This review discusses and summarizes the most current data available on molecular interactions and pathogenic mechanisms in antiphospholipid syndrome.     

Activation of complement mediates antiphospholipid antibody-induced pregnancy loss

Although it is clear that the specific antigenic reactivity of antiphospholipid (aPL) antibodies is critical to their effect, the pathogenic mechanisms that result in injury in vivo are incompletely understood. We hyphothesized that aPL antibodies targeted to the placenta activate complement locally, generating split products that mediate placental injury and lead to foetal loss and growth retardation. To test this hypothesis, we used a murine model of APS in which pregnant mice are injected with human IgG containing aPL antibodies. Mice treated with inhibitors of complement activation and mice deficient in complement components were protected from aPL antibody-induced foetal damage. Although the cause of tissue injury in this disease is probably multifactoral, we have shown that complement activation is an absolute requirement for foetal loss and growth restriction and, therefore, thatthis pathway acts upstream of other important effector mechanisms. Identification of complement activation as a mechanism that is necessary for aPL-induced tissue damage and definition ofthe complement components necessary to trigger such injury is likely to lead to a better understanding of the pathogenesis of vascular and tissue injury in SLE and to new and improved treatments.     

Pathogenetic mechanisms of antiphospholipid antibody production in antiphospholipid syndrome

Antiphospholiipid syndrome (APS) is an autoimmune disease characterized by the pathological action of antiphospholipid antibodies (aPL), that leads to recurrent pregnancy loss and thrombosis. Despite limited evidence, it is clear that there are both inherited and acquired components of the ontogeny of these antibodies. Animal genetic studies and human familial and population studies highlight the influence of genetic factors in APS, particularly human leukocyte antigen associations. Similarly, both animal and human studies have reported the importance of acquired factors in APS development and infectious agents in particular have a great impact on aPL production. Bacterial and viral agents have been implicated in the induction of autoimmune responses by various mechanisms including molecular mimicry, cryptic autoantigens exposure and apoptosis. In this review we highlight the latest updates with regards to inherited and acquired factors leading to the manufacturing of pathogenic antibodies and APS.     

The antiphospholipid syndrome and infection

Infectious agents have been implicated in the induction of antiphospholipid (aPL) antibodies and the development of the antiphospholipid syndrome (APS). This review focuses on the types of aPL antibodies detected in infections and addresses whether these antibodies are of clinical importance in patients with infections. Hepatitis C virus (HCV) infection is given special attention because this virus has the propensity to induce various autoimmune phenomena. Several aspects are emphasized that should be considered carefully when interpreting results. Most of the published data agree that thrombophilia is not observed in patients with infections (including HCV) because aPL antibodies are mostly the natural or nonpathogenic type. Thus, we do not recommend routinely testing for HCV in patients with APS. However, not all infection-associated aPL antibodies are cofactor independent. For instance, infections are increasingly recognized as a major precipitating condition of the catastrophic variant of APS, perhaps via mechanisms of molecular mimicry. Therefore, it may be possible to prevent this devastating evolution if the infectious process is promptly recognized and exhaustively treated.     

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

OBJECTIVE: 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. METHODS: 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). RESULTS: 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. CONCLUSION: 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.     

Gene polymorphisms of tissue plasminogen activator and plasminogen activator inhibitor-1 in patients with antiphospholipid antibodies

OBJECTIVE: Impaired fibrinolytical outcomes may be one of the pathogenic factors for thrombotic events in patients with antiphospholipid antibodies (aPL). We investigated the consequences of the gene polymorphisms of tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) in patients positive for aPL. METHODS: Seventy-seven Japanese and 82 British patients with aPL were examined for Alu-repeat insertion (I)/deletion (D) polymorphism of the tPA gene by polymerase chain reaction (PCR), and 4G/5G polymorphism in the PAI-1 promoter gene by site-directed mutagenesis-PCR and restriction fragment length polymorphism analysis. Correlations between these polymorphisms and clinical symptoms of antiphospholipid syndrome (APS) (arterial thrombosis, venous thrombosis, miscarriage) were analyzed. RESULTS: Significant differences in the allele frequencies of these genes did not exist between patients and controls. There was no significant correlation between these gene polymorphisms and clinical symptoms of APS in patients with aPL. CONCLUSION: Polymorphisms of the tPA or PAI-1 genes probably do not significantly influence the risk of anerial thrombosis, venous thrombosis, or pregnancy morbidity in patients with aPL.     

Interpretation and recommended testing for antiphospholipid antibodies

The antiphospholipid syndrome (APS) is defined by the association of arterial and/or venous thrombosis and/or pregnancy complications with the presence of at least one of the main laboratory-detected antiphospholipid antibodies (aPL) (i.e., lupus anticoagulants [LA], IgG and/or IgM anticardiolipin antibodies [aCL], and IgG and/or IgM anti-β2-glycoprotein I antibodies [aβ2GPI]). During the last decade efforts have been made to improve the harmonization and reproducibility of laboratory detection of aPL and guidelines have been published. The prognostic significance of aPL is being clarified through the fine elucidation of their antigenic targets and pathogenic mechanisms. Several clinical studies have consistently reported that LA is a stronger risk factor for both arterial and venous thrombosis compared with aCL and aβ2GPI. In particular, LA activity dependent on the first domain of β2-glycoprotein I and triple aPL positivity are prognosticators of the thrombotic and obstetric risks. Hopefully, this increasing knowledge will help improve diagnostic and treatment strategies for APS.     

Heterogeneity of antibodies to beta2-glycoprotein 1 from patients with systemic lupus erythematosus

We studied antibodies to beta2-glycoprotein 1 (anti-beta2GP1) from 72 patients with systemic lupus erythematosus (SLE) with or without antiphospholipid syndrome (APS) or with or without anticardiolipin antibodies (aCL). Fifteen patients had APS and positive antiphospholipid antibodies [clinical APS(+)/aPL(+)], 12 patients had APS, negative serum IgG and IgM aCL, antiphosphatidylethanolamine, anti-phosphatidylserine and no lupus anticoagulant [clinical APS(+)/ aPL(-)]. A third group included 16 patients without APS but high aCL levels [clinical APS(-)/ aPL(+)]. In a fourth group we studied 29 patients without clinical manifestations of APS or aCL [clinical APS(-)/aPL(-)]. One hundred anticardiolipin and VDRL-negative normal sera were studied as controls. IgG antibodies to cardiolipin proper in a bovine beta2GP-free system, to human beta2GP1 immobilized on cardiolipin or to human beta2GP1 alone were detected in all sera by ELISA using irradiated and nonirradiated plates from two manufacturers. Sera from APS(+)/aPL(+) patients showed IgG binding to CL, CL + beta2GP1 and beta2GP1 in irradiated and nonirradiated plates. APS(+)/ aPL(-) sera had more significant IgG binding to beta2GP1 than normal controls when studied in both irradiated or nonirradiated plates (P = 0.001). This binding was inhibited by solid-phase cardiolipin in a dose-dependent manner. Sera from the APS(-)/aPL(+) subgroup had comparable IgG activity in both the CL and CL + beta2GP1 assays, while no anti-beta2GP1 activity was detected in these sera. Sera from the clinical APS(-)/aPL(-) patients were negative in the three ELISA systems. Antibodies to human beta2GP1 from SLE patients recognize various epitopes. Those from APS(+)/ aPL(+) patients appear to react with an epitope boosted by cardiolipin in addition to another one present in the native protein. In contrast, anti-beta2GP1 from patients with APS(+)/aPL(-) are blocked by cardiolipin, suggesting that their epitope is the phospholipid-binding site.     

Recent advances in antiphospholipid antibodies and antiphospholipid syndromes in pediatric populations

In recent years, antiphospholipid antibodies (aPL) and their associated clinical features have been recognized increasingly in various pediatric autoimmune and non-autoimmune diseases. Pathogenic mechanisms involved in pediatric antiphospholipid syndrome (APS) appear to be the same as in adults. However, since pediatric patients do not have prothrombotic risk factors present in adults, there clearly are differences in the spectrum of clinical findings. The frequency of aPL-related thrombotic events is generally low in pediatric populations. On the other hand, various commonly acquired infections are likely to be responsible for higher percentage of non-pathogenic and transient aPL in childhood. Such points have to be considered in clinical judgment of elevated aPL in children. In this review we summarize the recent data on the prevalence and clinical significance of aPL in neonates, children and adolescents.     

Antiphospholipid syndrome: endocrinologic manifestations and organ involvement

Antiphospholipid syndrome (APS) is a systemic autoimmune disease characterized by thrombotic tendency that affects most organ systems in the human body. In this report, we present a review of the endocrinologic manifestations associated with APS by evaluating the medical literature from 1968 to 2009 using MEDLINE and these keywords: APS, antiphospholipid syndrome, antiphospholipid antibodies, anticardiolipin antibodies, lupus anticoagulant, anti β-2 glycoprotein I, pituitary, adrenal, thyroid, parathyroid, ovary, testes, diabetes mellitus, and diabetes insipidus. Adrenal insufficiency was found to be the most common endocrine manifestation associated with APS. Autoimmune thyroid disease was associated with increased titers of antiphospholipid antibodies (aPL) without any APS clinical manifestations. In addition, hypopituitarism and Sheehan syndrome are increasingly being reported in association with aPL. Data regarding the prevalence and significance of aPL in diabetic patients remains uncertain. Finally, only a few cases of ovarian and testicular derangements have been reported. APS should be considered in any patient with adrenal insufficiency even in the absence of other thrombotic manifestations. It is also advisable to assess aPL in the sera of patients presenting with pituitary insufficiency. Further studies are needed to clarify the relationship between aPL and thyroid disorders and diabetes mellitus.     

Identification of an Fcγ receptor–independent mechanism by which intravenous immunoglobulin ameliorates antiphospholipid antibody–induced thrombogenic phenotype

Objective

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 (Fcγ receptors [FcγR]). We examined the potential mechanisms of action of IVIG in an in vivo murine model of antiphospholipid antibody (aPL)–induced thrombosis and endothelial cell activation.

Methods

Mice infused with IgG containing human anticardiolipin antibodies (aCL) were treated with IVIG (36 μg IV), 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 FcγR are required for the effects of IVIG, we treated mice deficient in stimulatory FcγR. To examine the effects of IVIG on endogenously generated antibody, we treated mice immunized with β₂‐glycoprotein I (β₂GPI).

Results

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 β₂GPI. The thrombophilic effects of aPL were evident in FcγR‐deficient mice.

Conclusion

Treatment with IVIG inhibits the thrombogenic effects of aPL in vivo and reduces the levels of aCL in the circulation. Blockade of stimulatory FcγR 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.

     

Circulating levels of tissue factor and proinflammatory cytokines in patients with primary antiphospholipid syndrome or leprosy related antiphospholipid antibodies

The antiphospholipid syndrome (APS) is characterized by the presence of antiphospholipid antibodies (aPL) in patients with thromboembolic complications. In APS, most aPL are autoantibodies to beta2-glycoprotein I and prothrombin, which play a major role in the APS pathogenesis. Nevertheless, antibodies with the same antigen specificity are also found in aPL patients with leprosy, in whom thromboembolic complications are uncommon. The in vivo upregulation of the tissue factor (TF) pathway and the imbalance of cytokines have been proposed as potential mechanisms of thrombosis in the APS. We measured the circulating levels of TF, interleukin 6 (IL-6), IL-6 receptor (sIL-6R), tumor necrosis factor (TNF-alpha) and interferon gamma (IFN-gamma) in 83 patients with autoimmune aPL (42 with and 41 without clinical features of definite primary APS), 48 leprosy patients (33 with aPL) and 48 normal controls. There was a trend (P = 0.06) to higher median sTF in patients with autoimmune aPL (139 pg/mL) compared with leprosy patients (103.5 pg/mL) and controls (123 pg/mL). In addition, the frequency of raised sTF levels (> 187 pg/mL) was significantly higher in the group with autoimmune aPL [22.9% (APS 21.4%, non-APS 24.4%)] but not in leprosy (10.4%) compared with controls (4.2%). Elevated levels of IL-6 and TNF-alpha and a trend to lower IFN-gamma were found in patients with definite APS. Leprosy patients with aPL, however, had increased TNF-alpha and IFN-gamma but normal IL-6 levels. Levels of sIL-6R did not differ between controls and either patients with autoimmune aPL or leprosy. The different cytokine profiles as well as differences in circulating levels of TF might contribute to the high thrombotic risk found in patients with autoimmune aPL but not in leprosy related aPL patients.