Research of hemostasis system

The hemostatic system is a set of functional-morphological and biochemical mechanisms ensuring the preservation of the liquid state of the blood, preventing and stopping bleeding, and the integrity of blood vessels.

In a complete organism, in the absence of any pathological effects, the liquid state of the blood is a consequence of the balance of the factors that cause the processes of coagulation and hinder their development. Violation of this balance can be caused by very many factors, but regardless of etiological reasons, thrombotic formation in the body occurs according to unified laws with the inclusion of certain cellular elements, enzymes and substrates in the process.

There are two links in blood clotting: cellular( vascular-trom-botsitic) and plasma( coagulation) hemostasis.

■ Cellular hemostasis means the adhesion of cells( that is, the interaction of cells with a foreign surface, including cells of a different kind), aggregation( gluing of the same blood cells among themselves), as well as the release of elements that activate plasma hemostasis from the elements.

■ Plasma( coagulation) hemostasis is a cascade of reactions involving clotting factors that result in the formation of fibrin. The resulting fibrin is further destroyed by plasmin( fibrinolysis).

It is important to note that the division of haemostatic reactions into cellular and plasma is conditional, however, it is valid in the in vitro system and significantly facilitates the selection of adequate techniques and the interpretation of the results of laboratory diagnostics of the pathology of hemostasis. In the body, these two links of the coagulating blood system are closely related and can not function separately.

A vascular wall plays a very important role in the implementation of hemostasis reactions. Endothelial vascular cells are capable of synthesizing and / or expressing on their surfaces various biologically active substances that modulate thrombus formation. These include von Willebrand factor, endothelial relaxation factor( nitric oxide), pro-stacycline, thrombomodulin, endothelin, tissue type tissue-type plasminogen activator, tissue-type plasminogen activator inhibitor, tissue factor( thromboplastin), tissue factor pathway inhibitor and some others. In addition, membranes of endotheliocytes carry receptors, which under certain conditions mediate binding to molecular ligands and cells freely circulating in the bloodstream.

In the absence of any damage, the lining vessels of the endothelial cells have thrombose-resistant properties that the

method maintains the liquid state of the blood. The endothelial thromboreal resistance provides:

■ contact inertness of the inner( turned into the lumen of the vessel) surface of these cells;

■ synthesis of a potent platelet aggregation inhibitor - prostacyclin;

■ presence of thrombomodulin on the membrane of endotheliocytes, which binds thrombin;while the latter loses the ability to cause blood clotting, but retains the activating effect on the system of two most important physiological anticoagulants - proteins C and S;

■ high content of mucopolis-harids on the inner surface of vessels and fixation of heparin-antithrombin III( ATH) complex on the endothelium of the complex;

■ the ability to secrete and synthesize a tissue plasminogen activator that provides fibrinolysis;

■ the ability to stimulate fibrinolysis through the protein system C and S.

Violation of the integrity of the vascular wall and / or changes in the functional properties of endotheliocytes may contribute to the development of prothrombotic reactions - the antithrombotic potential of the endothelium is tris-formed into thrombogenic. The causes leading to vascular injury are very diverse and include both exogenous( mechanical damage, ionizing radiation, hyper and hypothermia, toxic substances, including drugs, etc.), and endogenous factors. The latter include biologically active substances( thrombin, cyclic nucleotides, a number of cytokines, etc.), capable of exhibiting membrane-aggressive properties under certain conditions. Such a mechanism of involvement of the vascular wall is typical for many diseases, accompanied by a tendency to thrombosis.

All cellular elements of the blood take part in thrombogenesis, but for platelets( unlike red blood cells and leukocytes) the procoagulant function is the main one. Platelets not only act as the main participants in the process of thrombus formation, but also have a significant effect on other links of hemocoagulation, providing activated phospholipid surfaces necessary for the realization of plasma hemostasis processes, releasing a number of coagulation factors into the blood, modulating fibrinolysis and disrupting hemodynamic constants both by transient vasoconstriction, caused by the generation of thromboxane A2, and by the formation and isolation of mitogenic factors contributing toiperplazii vascular wall. When thrombogenesis is initiated, platelets are activated( i.e., activation of platelet glycoproteins and phospholipases, metabolism of phospholipids, formation of secondary mediators, protein phosphorylation, arachidonic acid metabolism, actin and myosin interaction, Na + / H + exchange, fibrinogen receptor expression and calcium ion redistribution) and inductionprocesses of their adhesion, release and aggregation reaction;the adhesion precedes the release and platelet aggregation reaction and is the first step in the haemostatic process.

When the endothelial lining is violated, the subendothelial components of the vascular wall( fibrillar and nonfibrillar collagen,

elastin, proteoglycans, etc.) come in contact with blood and form a surface for binding the von Willebrand factor, which not only stabilizes factor VIII in plasma, but also plays a keyrole in the process of adhesion of platelets, binding subendothelial structures to cell receptors [Barkagan ZS, 1998].

Adhesion of platelets to the thrombogenic surface is accompanied by their spreading. This process is necessary to achieve a more complete interaction of platelet receptors with fixed ligands, which contributes to the further progression of thrombosis, since, on the one hand, it provides stronger adhesion of the cells to the vascular wall, and on the other hand, immobilized fibrinogen and von Willebrand factorare able to act as platelet agonists, promoting further activation of these cells.

In addition to interacting with a foreign( including damaged vascular) surface, platelets are able to adhere to each other, that is, aggregate. Aggregation of platelets is caused by substances of different nature, for example, thrombin, collagen, ADP, arachidonic acid, thromboxane A2 prostaglandins G2 and H2, serotonin, adrenaline, platelet activation factor and others. Proagregantami can be exogenous substances( not in the body), such as latex.

Both adhesion and aggregation of platelets can lead to the development of a release reaction - a specific Ca2 + -dependent secretory process in which platelets release a number of substances into the extra-cellular space. Induced release reaction of ADP, adrenaline, subendothelial connective tissue and thrombin. First, the contents of dense granules are released: ADP, serotonin, Ca2 +;to release the contents of a-granules( platelet factor 4, P-thromboglobulin, platelet-derived growth factor, von Willebrand factor, fibrinogen and fibronectin) requires more intensive platelet stimulation. Liposomal granules containing acid hydrolases are released only in the presence of collagen or thrombin. It should be noted that the released from the platelet factors contribute to the closure of the defect of the vascular wall and the development of the hemostatic plug, but with a sufficiently pronounced vessel damage, further activation of platelets and their adhesion to the injured portion of the vascular surface forms the basis for the development of a widespread thrombotic process followed by vessel occlusion.

In any case, the result of damage to endotheliocytes is the acquisition of intimal vessels with procoagulant properties, which is accompanied by the synthesis and expression of tissue factor( thromboplastin), the main initiator of the blood clotting process. Thromboplastin itself does not possess enzymatic activity, but can act as a cofactor of activated factor VII.The thromboplastin / factor VII complex is capable of activating both factor X and factor XI, thereby generating thrombin generation, which in turn induces further progress in the reactions of both cellular and plasma haemostasis.

Hemo-static reactions, a combination of which is commonly referred to as plasma( coagulation) hemostasis, ultimately result in

activation;- transition from one state to another

Fig. Thrombocytic hemostasis of

to fibrin formation;these reactions are mainly realized by proteins called plasma factors. In Table.the list of the factors participating in blood coagulation is resulted.

Table International Nomenclature of Coagulation Factors

Table International Nomenclature of Coagulation Factors

* Synthesized in the liver.

* Synthesized in the liver.

The process of plasma hemostasis can be conditionally divided into 3 phases.

Phase I - formation of prothrombinase or contact-kallikrein-kinin-cascade activation. Phase I is a multi-step process, as a result of which a complex of factors capable of converting prothrombin to thrombin accumulates in the blood, therefore this complex is called prothrombinase. There are internal and external ways of protrombinase formation. On the internal pathway, the coagulation of the blood is initiated without the involvement of tissue thromboplastin;Plasma factors( XII, XI, IX, VIII, X), kallikrein-kinin system and platelets take part in the formation of prothrombinase. As a result of the initiation of reactions of the internal pathway, a complex of factors Xa and V is formed on the phospholipid surface( platelet factor 3) in the presence of ionized calcium. This entire complex acts as a prothrombinase, converting prothrombin to thrombin. The triggering factor of this mechanism is XII, which is activated either by the contact of blood with a foreign surface, or by the contact of blood with subendothelium( collagen) and other components of connective tissue in damage to the walls of the vessels;either factor XII is activated by enzymatic cleavage( kallikre-

in other, plasmin, other proteases).In the external pathway for the formation of prothrombinase, the main factor is the tissue factor( factor III), which is expressed on cell surfaces with tissue damage and forms a complex with factor VIIa and calcium ions capable of translating factor X into factor Xa, which activates prothrombin. In addition, factor Xa retrogradely activates the complex of tissue factor and factor VIIa. Thus, the inner and outer paths are connected on the coagulation factors. The so-called "bridges" between these paths are realized through the mutual activation of factors XII, VII and IX.This phase lasts from 4 minutes 50 seconds to 6 minutes 50 seconds.

II phase - the formation of thrombin. In this phase, prothrombinase, together with coagulation factors V, VII, X and IV, transfers the inactive factor II( prothrombin) to the active factor IIa-thrombin. This phase lasts 2-5 s.

III phase - the formation of fibrin. Thrombin cleaves two peptides A and B from the fibrinogen molecule, converting it to fibrin monomer. The molecules of the latter are polymerized first into dimers, then into still soluble, especially acidic, oligomers, and eventually into fibrin-polymer. In addition, thrombin promotes the conversion of factor XIII to the factor XIII.The latter, in the presence of Ca2 +, changes the fibrin-polymer from a labile, readily soluble fibrinolysin( plasmin) form into a slowly and boundedly soluble form, which forms the basis of the blood clot. This phase lasts 2-5 s.

In the process of formation of a hemostatic thrombus, the distribution of thrombus formation from the site of damage to the vessel wall along the vasculature does not occur, as this is impeded by the rapidly increasing anticoagulation potential of the blood and the activation of the fibrinolytic system after the coagulation.

The preservation of blood in a liquid state and the regulation of the rates of interaction of factors in all phases of coagulation are largely determined by the presence in the bloodstream of natural substances with anticoagulant activity. The liquid state of blood provides a balance between the factors that induce blood clotting and the factors that prevent its development, the latter not being allocated to a separate functional system, since the realization of their effects is most often impossible without the participation of procoagulation factors. Therefore, the allocation of anticoagulants, preventing the activation of clotting factors and neutralizing active forms, is very arbitrary. Substances that have anticoagulant activity are constantly synthesized in the body and released into the bloodstream at a certain rate. These include ATH, heparin, proteins C and S, the recently discovered tissue clotting inhibitor-TFPI( inhibitor of the factor factor VIIa-Ca2 + complex), a2-macroglobulin, antitrypsin, etc. In the process of blood clotting, fibrinolysis from clotting factors andother proteins also produce substances with anticoagulant activity. Anticoagulants have a pronounced effect on all phases of blood coagulation, so the study of their activity in the case of blood clotting disorders is very important.

Fig. Plasma hemostasis. VMK - high molecular weight kininogen;RFMK - soluble fibrin-monomer complexes;fp A and B - fibrinopeptides A and B;С3, С5В, С9 - factors of complement system

Fig. Plasma hemostasis. VMK - high molecular weight kininogen;RFMK - soluble fibrin-monomer complexes;fp A and B - fibrinopeptides A and B;C3, C5B, C9 - factors of the complement system

Fig. III phase of blood coagulation( fibrin formation)

After stabilization of fibrin, together with the shaped elements of the primary red blood clot forming, two main processes of the postcoagulable phase begin - spontaneous fibrinolysis and retraction, resulting in the formation of a hemostatically complete final thrombus. Normally, these two processes proceed in parallel. Physiological spontaneous fibrinolysis and retraction contribute to tightening the thrombus and performing hemostatic functions. In this process, an active part is taken by the plasmin( fibrinolytic) system and fibrinase( factor XIIIa).Spontaneous( natural) fibrinolysis reflects a complex reaction between the components of the plasmin system and fibrin. The plasmin system consists of four main components: plasminogen, plasmin( fibrinolysin), activators of fibrinolysis proenzymes and its inhibitors. Violation of the ratios of the components of the plasmin system leads to pathological activation of fibrinolysis.

In clinical practice, the study of the hemostasis system has the following objectives:

■ diagnosis of hemostasis system disorders;

■ elucidation of the admissibility of surgical intervention for revealed violations in the hemostasis system;

■ monitoring of anticoagulant treatment of direct and indirect effects, as well as thrombolytic therapy.