The purpose of secondary hemostasis is to generate a fibrin network to “cement” the primary platelet plug.

Secondary hemostasis, also known as coagulation, con sists of a complex series of events involving a large number of molecules (Table 1). The traditional model of secondary hemostasis is that of a “cascade,” which describes coagulation as a sequential series of enzymatic or proteolytic reactions in each of which an inactive precursor of a proteolytic enzyme (termed zymogen) is converted to its active form (protease), which in turn activates the next zymogen in the series. Since the individual sequential reactions are catalyzed by an enzyme, and since each enzyme can catalyze the generation of a large number of other active molecules, it is easy to understand how this model is characterized by an enormous amplifying potential, hence the name “cascade.” It is possible to summarize the main features of the cascade model in a few essential points.

Table1. Numbering of coagulation factors (F)

• Each step in the cascade includes a serine protease and sometimes a cofactor (to form an enzyme “complex”).

 • Enzyme complexes consist of similar components with similar characteristics with respect to their assembly and function.

• The formation of the complex results in a considerable increase (about 100-fold) of the catalytic efficiency towards the substrate.

• To function effectively, individual zymogens and enzyme complexes require the binding of one of their terminal carboxyl groups to an appropriate surface consisting of negatively charged phospholipids (generally those present on the membrane of platelets), a binding mediated by the indispensable presence of calcium ions (Ca²⁺) (Fig. 1). In the absence of the carboxylic group on the coagulation factors or as a result of the subtraction of calcium ions, the binding between the coagulation factor and the platelet occurs with much less efficiency, overall, also altering the effectiveness of the coagulation cascade.

Fig1. Role of calcium in secondary hemostasis. (Copyright EDISES 2021. Reproduced with permission)

During the last two decades, the model of the coagulation cascade has undergone considerable interpretative revisions, conditioned by the progressive discovery of new coagulation factors and new and complex biochemical interactions. The historical paradigm of coagulation is the one that, for years, was based on the existence of three different “pathways,” also known as extrinsic, intrinsic, and common. Over the years, however, it has been demonstrated that this model, while maintaining stringent validity in the interpretation of laboratory abnormalities of the coagulation cascade, has no real feedback in vivo. In this case, it has been concluded that the “parallel” model, according to which extrinsic and intrinsic pathways concur in parallel to the activation of the common pathway, has no real biological correspondence since the physiological mechanism of activation of coagulation is represented only by the extrinsic pathway. According to the most current model, therefore, coagulation can be described in three sequential phases (Fig. 2):

• Activation phase (“trigger”) of the coagulation cascade: mediated by the extrinsic and common pathways. The exposure of the TF is the key event.

• Propagation phase (burst) of thrombin: mediated by an intrinsic and common pathway, it is characterized by the formation of enzymatic complexes aimed at the creation of a stable clot.

• Coagulation shutdown phase: supported by a discrete number of natural anticoagulants, aimed at circumscribing procoagulant events and preventing the formation of excess clots.

Fig2. Secondary hemostasis. The dotted lines define the critical processes of the thrombin burst. AT antithrombin, FB fibrin, FBG fibrinogen, FDP fibrin degradation products, FII prothrombin, FIIa thrombin, PC protein C, PG plasminogen, PN plasmin, PS protein S, TF tissue factor, tPA tissue plasminogen activator, TFPI inhibitor of the tissue factor pathway. (Copyright EDISES 2021. Reproduced with permission)

In the following paragraphs, we will try to fully describe the sequence of events; for the sake of synthesis, the term “coagulation factor” will be abbreviated with the letter “F” for all the different factors.

Coagulation Activation Phase

The physiological activation (“trigger”) of coagulation is supported only and exclusively by the extrinsic pathway and, subsequently, by the common pathway. This evidence is supported not only by the physiological mechanism that will be described later but also by the clear and indisputable clinical evidence that total deficits of FXII (the factor upstream of the activation of the intrinsic pathway) are fully compatible with life and, indeed, do not cause any type of bleeding diathesis. As will be seen later, the task of the intrinsic pathway is not to constitute a mechanism of parallel activation of coagulation but rather to amplify the formation of thrombin and fibrin, generated in small quantities by the activation of the extrinsic pathway.

Activation of the extrinsic pathway thus originates from the rupture of the vascular wall, which directly results in damage to endothelial cells and exposure or release of the TF contained therein. Immediately after its release, TF binds with high affinity and specificity to FVII, thus triggering coagulation. Since TF is essentially a transmembrane protein, its propagation in circulation is very limited, except in the presence of considerable tissue damage. This is easily justified by the need for the coagulation process to be limited to the site where the endothelial damage has occurred without excessive propagation to other districts in which the coagulation process is not required (intact endothelium) and in which the excessive activation of coagulation could, on the other hand, generate the paradoxical phenomenon of thrombosis. It is interesting to note how the excessive propagation of TF is contrasted not only by the layer of platelets that have adhered to the collagen at the site of endothelial damage but also by the release, by the same endothelial cells damaged, of the inhibitor of the tissue factor pathway (TFPI), also known as EPI (extrinsic pathway inhibitor), actively released by endothelial cells and, probably, also by platelets. Unlike what was thought years ago, a small amount of TF is always present in the blood, even in the absence of endothelial damage. Although its origin remains controversial, it is legitimate to assume that it originates from cellular microparticles, tiny membrane vesicles released from stimulated cells, or cells undergoing apoptosis. The essential function of TF is to bind to FVII (zymogen) and activate it to FVIIa (active enzyme) by proteolytic cutting at the level of Arg152, which determines the conversion of FVII from a single chain to a double chain. Recent evidence shows that a modest amount of FVIIa (1–2% of total FVII) is nevertheless present in the circulation, despite its concentration being too modest to trigger the coagulation process, especially in the absence of TF (only the interaction between FVIIa and TF is able to “trigger” the active site of FVIIa, increasing its catalytic activity towards the next factor in the coagulation cascade, i.e., FX). Although the origins and function of this small amount of physiologically circulating FVIIa are still uncertain, it seems plausible to assume that the serine protease factor VII-activating protease (FSAP) can produce the physiological activation of a modest amount of FVII in order to make coagulation more ready and efficient when necessary.

The function of the TF-FVIIa complex (also known as the extrinsic complex) is primarily to activate FX to FXa and, to a small extent, also to activate FXI to FXIa. FXa, together with its essential cofactor FVa, forms the so-called tenase complex, which has the function of activating prothrombin (FII) to thrombin (FIIa). Thrombin, in turn, converts fibrinogen into fibrin, which, as it polymerizes, forms a “mesh work,” or “reticulum,” aimed at stabilizing the thrombus, according to a series of mechanisms described below. The quantities of thrombin and fibrin produced by the mechanisms now described are in the order of nanomoles, which are concretely too scarce (3–5% of the necessary amount) for a fibrin lattice large enough to stabilize the platelet plug (in practice, this is an “ineffective” clot). The main function of this small amount of thrombin produced is to activate a secondary “burst” (also known as the thrombin propagation phase), which is the most functional link between the extrinsic and intrinsic pathways.

Thrombin Propagation Phase

As repeatedly described, the initiation of coagulation by the extrinsic pathway produces small amounts of thrombin (3–5% of the total), which are nevertheless sufficient to accelerate the coagulation process by a series of mechanisms involving platelet activation, FXIII activation, and activation of FV and FVIII cofactors. Most thrombin (95–97%) is instead generated in the so-called propagation phase of coagulation (thrombin burst; Fig. 2).

The sequential series of events in the thrombin propagation phase involves the activation of FXI (to FXIa), which in turn catalyzes the conversion of FIX to FIXa. This latter fac tor, together with its essential cofactor FVIII (FVIIIa in activated form), forms a molecular complex called intrinsecase. The FIXa-FVIIIa complex can activate FX to FXa, with kinetics almost 100-fold higher than the single action of FIXa and 50- to 100-fold higher than the action of the TF-FVIIa complex. It is, therefore, evident how the maxi mum part of the activation of FX to FXa is sustained by the intrinsecase complex (FIXa-FVIIIa) and not by the extrinsic complex (TF-FVIIa), which justifies the onset of a severe hemorrhagic syndrome related to the deficiency of FVIII (hemophilia A) and FIX (hemophilia B).

In the final phase of coagulation, as described above, thrombin acts enzymatically on the fibrinogen molecule, determining the cutting of part of the α and β chains (releasing the corresponding fibrinopeptides A and B) and generating the so-called fibrin monomer. This peptide has a characteristic shape, consisting of a central domain (called E) and two side domains (called D). The fibrinopeptide A cleavage exposes a binding sequence on the central E domain that associates, by noncovalent bonding, with a complementary site on the lateral D region of an adjacent fibrin mono mer. The overlap of fibrin molecules, where the central domain of one binds to the lateral domain of the other, results in the formation of double-stranded protofibrils. The release of fibrinopeptide B exposes an additional binding site on the central E domain, which in turn binds to a complementary site on the lateral D region of another adjacent fibrin monomer, thereby promoting the lateral association of the protofibrils and the formation of thicker fibers. The deposition of this first fibrin lattice on the platelet plug is essentially governed by weak forces, which are not sufficient to ensure the long-term stability of the clot. Its stabilization is, therefore, dependent on the intervention of FXIIIa. In summary, FXIIIa associates with fibrin to promote the formation of a series of stable bonds in the clot, initially between chains of two adjacent fibrin molecules and subsequently forming covalent bonds between two overlapping fibrin chains (Fig. 3). Once this process has been completed, the clot is sufficiently stable to provide an effective barrier against the leakage of blood from the injured vessel.

Fig3. Fibrinoformation and fibrinolysis. (Copyright EDISES 2021. Reproduced with permission)

As a corollary to the description of the coagulation cascade, it is necessary to define, at least briefly, the role of VWF. This factor is a large adhesive protein synthesized mainly by endothelial cells and megakaryocytes and is therefore present in the α granules of platelets. It is present in the circulation in the form of multimers (aggregations of monomers), which, according to their extent, are classified as low, intermediate, and high molecular weight multimers. It per forms two essential functions. The first, already described regarding primary hemostasis, is that of mediating platelet aggregation or adhesion. The second function, equally essential, is to act as a carrier of FVIII in the circulation, thus protecting it from early degradation. Deficiency of this factor generates Von Willebrand disease (VWD), whose clinical characterization is very complex (types 1 and 3 are due to partial or virtually complete VWF deficiency, while type 2 is due to a qualitative defect), depending on the nature of the deficiency. In general, however, it is a disease with highly variable symptoms; the dual function of VWF in primary and secondary hemostasis determines the appearance of “mixed” symptoms between the two conditions. The high molecular weight multimers of VWF are much more efficient in the process of platelet aggregation. The efficiency of the process is guaranteed by the catalytic action of the enzyme ADAMTS13, which acts by cutting large VWD into small fragments. Alterations in the action of ADAMTS13 lead to a polymerization deficit, as, for example, in thrombotic thrombocytopenic purpura, in which ADAMTS13 deficiency pre vents the degradation of multimers that remain anchored to the endothelium, favoring the adhesion of platelets by means of gp Ib and gp IIb/IIIa and thus favoring the formation of potentially occlusive thrombi in terminal arterioles and capillaries of many organs (heart, pancreas, kidneys, adrenals, brain, spleen, and liver).

Coagulation Shutdown Phase

The fine regulation of the hemostatic process is guaranteed by the intervention, at various levels and with different mechanisms, of a series of physiological inhibitors of coagulation, essentially represented by the thrombin activatable fibrinolysis inhibitor (TAFI), antithrombin (AT), and protein C (PC)–protein S (PS) complex.

Antithrombin

Antithrombin (AT) is by far the most potent and important inhibitor of coagulation. Present in the blood at a much higher concentration than any other coagulation factor generated by the TF pathway, it appears to be capable of inhibiting multiple coagulation factors, although its activity is directed primarily at inhibition of FXa and thrombin (Fig. 2). The inhibitory efficacy of antithrombin is strongly conditioned by the presence of physiological (heparan sulfates lining the inner wall of vessels) or pharmacological (heparin) heparin-like substances, in the presence of which its inhibitory activity on FXa and thrombin is increased almost 1000-fold. The importance of the inhibitory effect of antithrombin is reflected in the association between deficiency and venous thrombosis, from the incompatibility with life to the state of homozygous deficiency and the effective ness of heparin therapy in preventing thrombotic events.

Protein C (PC) and Protein S (PS) System

 The protein C (PC) and protein S (PS) system is the second most biologically and clinically important inhibitory mechanisms of coagulation. PC is a vitamin K-dependent protein whose activity is conditioned by binding to negatively charged surfaces (phospholipids), thrombomodulin, and the endothelial protein C receptor (EPCR). The activity of protein C is then strictly dependent on the presence of protein S, which therefore represents its essential cofactor, increasing the activity of protein C about 20-fold. Protein S is also vita min K-dependent and circulates in the blood in a free form (30–40% of the total) or complexed with C4BP (60–70%), a complement regulatory protein. Only the free form of protein S can carry out its activity as a cofactor of protein C, hence the importance in the laboratory of effectively distinguishing the free circulating quantity (the “effective” one) from the total one. To better understand how the PC–PS system works, it is essential to briefly illustrate the structure of the two cofactors of coagulation (FV and FVIII) that represent the molecular target. The two proteins have considerable structural homology, which involves an arrangement into sequential domains (A1A2-B-A3-C1-C2). Thrombin activates both FV and FVIII by proteolytic cutting, causing the release of the B domain. The PC–PS system then inactivates FV by making an additional cut at the levels of Arg306, Arg506, and Arg679. Proteolytic cutting at the level of Arg506 results in the formation of an intermediate form of FVa that retains some of its procoagulant properties, whereas cutting at the level of Arg306 causes the total loss of the fac tor’s procoagulant activity. The inactivation of FVIIIa by the PC–PS complex occurs instead by proteolytic cutting at the levels of Arg336 and Arg562, although only the latter (Arg562) determines the complete inactivation of the factor.

اخر الاخبار

اخبار العتبة العباسية المقدسة

فرقة العباس تنظم مجلس عزائها السنوي بذكرى شهادة النبي (صلى الله عليه وآله) في النجف

قسم الشؤون الفكرية وجامعة الكوفة يعقدان شراكة علمية لتدريب الطلبة على صيانة المخطوطات وحفظها

بالتزامن مع ذكرى شهادته.. قسم الشؤون الفكرية يصدر كتابًا عن النبي الأعظم (صلى الله عليه وآله)

بالصور.. جموع المؤمنين تحيي ذكرى شهادة الرسول الأكرم عند مرقد الإمام علي (صلوات الله عليهما)

المزيد

الآخبار الصحية

لون أسنانك يتحدث عن صحتك.. إليك ما يقول

عادة يومية بسيطة تخفض الكوليسترول وتحمي القلب

الزنجبيل.. دواء سحري لحماية القلب

المشروبات شديدة السخونة.. "خطر خفي" يهدد صحتك

المزيد

الآخبار التكنلوجية

كيف يمكن لعادات القيادة السيئة أن تؤثر على قيمة سيارتك

في طوفان الذكاء الاصطناعي.. نصائح لحجز "تذكرة النجاة"

هكذا يسرق "قراصنة الذكاء الاصطناعي" المليارات بطرق بسيطة

فيديو لانفجار كرة نارية في سماء اليابان.. ما تفسير ذلك؟

المزيد

مواضيع ذات صلة

Fibrinolysis

Primary Hemostasis and Platelets

Primary Hemostasis and Vessel Participation

Phenotypic and Functional Properties of Erythroid Progenitor Cells

GRAPHICAL ANALYSIS OF VENTRICULAR PUMPING

The Heart Valves Prevent Backflow of blood during Systole

Excitation-Contraction Coupling— function of calcium ions and the transverse tubules

PHYSIOLOGIC ANATOMY OF CARDIAC MUSCLE

الطحال Spleen

الثايمس Thumus

اللوزتان Tonsils

العقد الليمفاوية