**4. Pathological mechanisms of the Antiphospholipid syndrome**

Antiphospholipid syndrome (APS) is a systemic autoimmune disorder characterized by venous and/or arterial thrombosis and pregnancy complications in the presence of antiphospholipid antibodies (aPL). aPL are a heterogeneous group of autoantibodies, of which anti-cardiolipin (anti-aCL), anti-β2 glycoprotein I (anti-β2GPI) and lupus anticoagulant (LA), are in the laboratory criteria for the diagnosis of APS [63]. In addition to criteria aPL other, non-criteria aPL, such as antibodies against phosphatidylserine/prothrombin complex, were found to play an important role in APS [64, 65]. These antibodies are, in some patients, the only

elevated aPL. Although aPL are persistent in APS patients, thrombosis occurs only occasionally, suggesting the involvement of other triggers that, together with aPL, turn the hemostatic balance in favor of thrombosis. In the development of APS, a two hit theory has been proposed in which the continuous presence of aPL as the first hit and inflammation, trauma, or surgery as a second hit together lead to thrombus formation [66, 67]. APS pathogenesis clearly involves both inflammatory and coagulation pathways in endothelial cells, monocytes, neutrophils, and platelets. Frequently identified prothrombotic mechanism is inhibition of the natural anticoagulant pathways [68]. It has been shown that aPL inhibit the activation of protein C [69] and its ability to inactivate factors V and VIII [70]. In addition, aPL inhibit the activity of TFPI [71] and activation of antithrombin [72]. They have also been found to be involved in fibrinolysis by neutralizing the ability of anti-β2GPI to stimulate tissue-type plasminogen activator [73]. Furthermore, aPL impair the ability of Annexin A5 to form a network on procoagulant anionic phospholipids [74]. aPL also directly bind to vascular cells and trigger their activation, which in response, release prothrombotic molecules and thus contribute significantly to the pathogenesis of APS. The activation of endothelial cells leads to a disruption of the normally anticoagulant endothelial surface [68]. This is achieved by upregulating adhesion molecules (E-selectin, ICAM-1, VCAM-1) [75], molecules involved in coagulation (TF) [76] and by the decrease in endothelial cell derived nitric oxide [77]. The biochemical pathways are not fully defined, but research has suggested several receptor-mediated mechanisms including, annexin A2, TLR4/NF-κB, TLR2, TLR7 and low-density lipoprotein receptorrelated protein 8 [68]. In addition to endothelial cells, aPL also act on platelets. Increased production of thromboxane B2, increased platelet adhesion to collagen type I and III and increased platelet activation have been described [66]. Among immune cells, monocytes are the most extensively studied in APS. In APS patients, monocytes have been shown to have a proinflammatory and procoagulant phenotype that is mediated by upregulation of NF-κB, MEK-1/ERK, and p38 MAP kinase pathways [78]. The main player of the procoagulant phenotype is increased surface expression, production and activity of TF on monocytes [79]. Stimulation of monocytes with aPL influences the release of IL-1β [80] and TNFα [81], probably by the activation of NLR family pyrin domain containing 3 inflammasome [82]. Monocyte-endothelial interactions are increased by upregulation of adhesion molecules on both cell types, as well as expression of other molecules, such as monocyte chemoattractant protein-1 by the endothelium, which in turn promotes the synthesis of TF by monocytes [83].
