Dual role of collagen in factor XII–dependent thrombus formation

Abstract

In vivo mouse models have indicated that the intrinsic coagulation pathway, initiated by factor XII, contributes to thrombus formation in response to major vascular damage. Here, we show that fibrillar type I collagen provoked a dosedependent shortening of the clotting time of human plasma via activation of factor XII. This activation was mediated by factor XII binding to collagen. Factor XII activation also contributed to the stimulating effect of collagen on thrombin generation in plasma, and increased the effect of platelets via glycoprotein VI activation. Furthermore, in flow-dependent thrombus formation under coagulant conditions, collagen promoted the appearance of phosphatidylserine-exposing platelets and the formation of fibrin. Defective glycoprotein VI signaling (with platelets deficient in LAT or phospholipase C␥2) delayed and suppressed phosphatidylserine exposure and thrombus formation. Markedly, these processes were also suppressed by absence of factor XII or XI, whereas blocking of tissue factor/factorVIIa was of little effect. Together, these results point to a dual role of collagen in thrombus formation: stimulation of glycoprotein VI signaling via LAT and PLC␥2 to form procoagulant platelets; and activation of factor XII to stimulate thrombin generation and potentiate the formation of platelet-fibrin thrombi. (Blood. 2009; 114:881-890) IntroductionVascular injury leads to exposure of hemostatically active components, such as tissue factor and collagen, to the bloodstream. For a long time, it has been considered that exposed tissue factor is the main vascular activator of the coagulation cascade. In the extrinsic pathway of coagulation, de-encrypted tissue factor complexes with circulating factor (F)VII(a), which activates a multistep cascade of serine proteases to form thrombin and fibrin. 1 The exposed collagen is thought to function as a substrate for the adhesion and activation of platelets via the glycoprotein VI (GPVI) receptor, which evokes an important signal transduction pathway in platelets. 2 Independently of tissue factor, the second intrinsic coagulation pathway is initiated by activation of plasma FXII (Hageman factor). Proteolytically active FXIIa mediates the sequential activation of the serine proteases, FXI and FIX, which also lead to thrombin generation. Until recently, the physiologic relevance of this pathway has remained obscure, as the initial trigger in vivo was not known. On the other hand, in vitro the intrinsic system is easily triggered in plasma using negatively charged materials, such as kaolin, glass, and ellagic acid. Common clotting tests, such as the activated partial thromboplastin time (aPTT), rely on activation of the intrinsic coagulation pathway. In the 1980s, it was proposed that polyanionic vascular components, such as cerebroside sulfates and glycosaminoglycans, function to stimulate FXII activation. 3,4 A similar role for collagen was suggested, 5,6 but this was refuted by other authors. [7][8][9] Recent studies with mice have greatly increased the inte...

Suboptimal Activation of Protease-activated Receptors Enhances α2β1 Integrin-mediated Platelet Adhesion to Collagen

“…Mangin et al (40) reported that thrombin was able to compensate for the effect of GPVI/FcR␥ deficiency in mouse platelets in in vivo models of thrombosis. The mechanism we describe, selective enhancement of the adhesive activity of the ␣ 2 ␤ 1 integrin via PAR4 activation, is the likely explanation for the observations of van der Meijden et al (39) and Mangin et al (40).…”

“…This mechanism is further supported by the recent findings regarding the role of thrombin in platelet adhesion to collagen under conditions of flow. Van der Meijden et al (39) described separate but complimentary roles of thrombin produced subsequent to the contact activation of factor XII by collagen, and GPVI/FcR␥ signaling on human platelet adhesion to collagen. Furthermore, inhibition of thrombin resulted in diminished platelet adhesion to collagen.…”

Rate-limiting roles of the tenase complex of factors VIII and IX in platelet procoagulant activity and formation of platelet-fibrin thrombi under flow

“…15,38 In order to investigate the role of FVIII in this process, we used citrate-anticoagulated blood from F8 -/-or F8 +/+ mice and made this flow during recalcification over collagen at an intermediate wall-shear rate (1000 s -1 ). The blood samples were supplemented with a surplus of 10 pM tissue factor to stimulate the extrinsic pathway.…”

“…37 Samples of citrate-anticoagulated blood (400-1500 μL) were made to flow at defined wall-shear rate, as described for murine 38 …”

Counting and Tracking: Development and Use of New Methods for Detailed Analysis of Thrombus Formation

“…Three interesting possibilities have been discussed but not fully validated. Fibrillar collagen is thought to activate FXII in addition to activating platelets (15), neutrophil extracellular traps (NETs) are negatively charged and could thereby trigger FXII activation (16) and activated platelets may induce coagulation through FXII by the release of polyphosphates. However the platelet-induced activation has been debated, it was originally postulated by Müller et al (17) and has thereafter been contradicted by our research group, Faxälv et al (18).…”