2 курс / Нормальная физиология / Учебное_пособие_по_физиологии_крови_Авдеева_Е_В_,_Репалова_Н_В_

II. Coagulatory haemostasis.

It is the cascade of biochemical reactions, which result in the formation of fibrin. Plasma factors of coagulation participate coagulative haemostasis:

I – Fibrinogen; II – Prothrombin;

III – Tissue thromboplastin. (tissue factor); IV – Ca2+ ions;

V – Proaccelerin, Ac-globulin, labile factor; VI – Active form of factor V (accelerin); VII – Proconvertin, stable factor;

VIII – Antihaemophilic globulin, antihaemophilic factor A; IX – Christmas’s factor, antihaemophilic factor B;

X – Stuart-Prauer’s factor, prothrombinase;

XI – Plasma antecessor of thromboplastin, antiheamophilic factor C; XII – Hageman’s factor, contact factor;

XIII – Fibrinstabilizing factor, fibrinase.

Coagulative reactions are based on the reactions of hydrolysis, which are performed by proteolytic enzymes. Reactions occur in phospholipids of membranes of destroyed erythrocytes and thrombocytes. The factors of coagulation are fixed on a membrane with the help of Ca2+ ions.

The main stages of blood coagulation are described by Moravic in 1905.

Fig. 39. Coagulation phases.

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There are 3 stages of blood coagulation:

Stage 1 – formation of active prothrombin activator.

In response to rupture of the vessel or damage to the blood itself, a complex cascade of chemical reactions occurs in the blood involving more than a dozen blood coagulation factors. The net result is formation of a complex of activated substances collectively called prothrombin activator.

Protrombin activator is formed with the help of two mechanisms:

1)extrinsic – that is initiated by phospholipids, which are released from damaged cells of vessels or from connective tissue;

2)intrinsic – that is initiated by coagulative factors of blood.

(F. VII). F.VIIa with phospholipids of tissues and calcium form complex, which activate prothrombinase (F. X). F. Xa with phospholipids, calcium and F. Va (proaccelerin) form complex, which is tissue prothrombin activator.

Tissue prothrombinase activates a small amount of thrombin, which:

is used for the thrombocytes aggregation;

provides formation of receptors on membranes of aggregated thrombocytes that bind F. Xa, resulting in its unavailability for anticoagulants, mainly for antithrombin III.

Fig. 40. Extrinsic (tissue) mechanism.

Intrinsic (blood) mechanism (fig. 41) – lasts for 5-7 minutes. Reactions occur on the membranes of the damaged blood cells (thrombocytes, erythrocytes). Collagen fibers initiate the process, they uncover after damage of a vessel. Contact of collagen with F. XII (Hageman’s factor), provides activation of F. XII. F. XIIa, F. XIa and

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phospholipids form complex, which activate F. X. F. Xa together with Ca2+ ions and F. Va also form complex, which is blood prothrombin activator.

Fig. 41. Intrinsic (blood) mechanism.

Stage 2 – formation of thrombin.

The prothrombin activator catalyzes conversion of prothrombin into thrombin (fig. 42).

This is a fast stage (2-5 sec). Blood prothrombinase absorbs prothrombin on its surface and in presence of calcium transforms it to thrombin.

Fig. 42. Formation of thrombin.

Prothrombin is a plasma protein, an alpha – 2-globulin, having a molecular weight of 68.700. Prothrombin is formed continually by the liver. Thrombin is a protein enzyme with the weak proteolytic capabilities.

Stage 3 – formation of fibrin.

Thrombin acts as an enzyme to convert fibrinogen into the fibrin fibers that enmesh platelets, blood cells, and plasma to form the clot itself (fig. 43).

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Fig. 43. Formation of fibrin.

Fibrinogen is a high-molecular-weight protein (MW = 340.000) that occurs in plasma. Fibrinogen is formed in the liver.

Because of its large molecular size, little fibrinogen normally leaks from the blood vessels into the interstitial fluids, and because fibrinogen is one of the essential factors in the coagulation process, interstitial fluids ordinarily do not coagulate.

Fibrin is formed from fibrinogen under the influence of thrombin. Thrombin acts on fibrinogen to remove four low-molecular weight peptides from each molecule of fibrinogen, forming one molecule of fibrin monomer. Polymerization of fibrin monomer starts and fibrin S is formed, which is soluble. With the help of F. XIII (fibrin stabilizing factor) and calcium in S molecule covalent connections are formed and fibrin S turns to insoluble fibrin I.

Fig. 44. Levels of fibrin formation.

So, the result of the third stage is the frormation of fibrin (fig. 44). A clot is composed of a meshwork of the fibrin fibers running in all directions and entrapping blood cells, platelets, and plasma (fig. 45). The fibrin fibers also adhere to the

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damaged surfaces of blood vessels; therefore, the blood clot becomes adherent to any vascular opening and thereby prevents the further blood loss.

Fig. 45. Blood clot.

2.4. Retraction.

The formed blood elements mesh in the fibrin fibers. But this ball of fibers is soft. That’s why the next step of the process is retraction. Packing of thrombus occurs due to contraction of protein thrombocystein. After retraction a clot is packed in 2 times (fig. 46), serum is removed from it and it becomes compact and plasma can not pass through it.

Fig. 46. Retraction of clot.

Platelets are necessary for clot retraction to occur. Electron micrographs of platelets in blood clots show that they become attached to the fibrin fibers in such a way that they actually bond different fibers together. Furthermore, platelets entrapped in the clot continue to release procoagulant substances, one of the most important of which is fibrin-stabilizing factor, which causes more and more cross-linking bonds between adjacent fibrin fibers. In addition, the platelets themselves contribute directly to clot contraction by activating platelet thrombosthenin, actin, and myosin molecules, which are all contractile proteins in the platelets and cause strong contraction of the platelet spicules attached to the fibrin. This also helps compress the fibrin meshwork into a smaller mass. Retraction lasts for 2-3 hours. After some time a clot starts to spring with fibroblasts. This occurs under the influence of thrombocytes growth factor. Integrity of the damage spot of a vessel is restored.

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2.5. Fibrinolysis.

Fibrinolysis is the decomposition of a fibrin clot (fig. 47). It begins simultaneously with retraction, but proceeds at a lower rate. The main function is restoration of a vascular lumen that was stopped with a clot.

The content of fibrinolytic system:

Plasminogen (profibrinolysin) – not-active proteolytic enzyme, which is always in blood plasma.

Plasmin (fibrinolysin) – active enzyme, which is produced in the result of effect of active proteases on plasminogen.

Activators of fibrinolysis – substances that are proteases or provide proteases synthesis.

Inhibitors of fibrinolysis.

There are internal and external mechanisms of fibrinolysis activation. Internal mechanism includes activation of F. XII and formation of kallikrein,

which causes a large amount of fibrinolysis activators to appear in blood.

External mechanism is associated with tissue plasminogen activator (t-PA) and urokinase (enzyme that is found in the urine).

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Fig. 47. Fibrinolysis.

«+» – arrows denote stimulation and «-» − arrows inhibition.

Plasmin is produced in an inactive form, plasminogen, in the liver. When a clot is formed, a large amount of plasminogen is trapped in the clot along with other plasma proteins. This will not become plasmin or cause lysis of the clot until it is activated. The injured tissues and vascular endothelium very slowly release a powerful activator called tissue plasminogen activator (t-PA) that a few days later, after the clot has stopped the bleeding, eventually converts plasminogen to plasmin, which in turn removes the remaining unnecessary blood clot. t-PA and urokinase are

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themselves inhibited by plasminogen activator inhibitor-1 and plasminogen activator inhibitor-2 (PAI-1 and PAI-2). In contrast, plasmin further stimulates plasmin generation by producing more active forms of both tissue plasminogen activator (tPA) and urokinase. Alpha 2-antiplasmin and alpha 2-macroglobulin inactivate plasmin. Plasmin activity is also reduced by thrombin-activatable fibrinolysis inhibitor (TAFI), which modifies fibrin to make it more resistant to the tPA-mediated plasminogen.

The lysis of blood clots allows slow clearing (over a period of several days) of extraneous clotted blood in the tissues and sometimes allows reopening of clotted vessels. An especially important function of the plasmin system is to remove very minute clots from the millions of tiny peripheral vessels that eventually would all become occluded were there no way to cleanse them.

3. Anticoagulation system.

Blood fluidity is maintained by several mechanisms such as:

1)smooth surface of a vessel wall endothelium;

2)negative charge of vessel wall and formed blood elements, so they repel from each other;

3)thin fibrin layer on a vessel wall, which absorbs factors of blood clotting, especially thrombin;

4)synthesis of prostacyclin by endothelium, which is a inhibitor of aggregation;

5)ability of endothelium to synthesize and fix antithrombin III;

6)presence of anticoagulants in the bloodstream.

Fig. 48. Physiological balance.

Classification of anticoagulants (fig. 49):

1.Primary (always present in plasma): antithrombin III, heparin, α1-antithropsin, α2-macroglobulin.

2.Secondary (they are formed in the process of clotting): antithrombin I, fibrin fibers, factor XI, peptides.

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Fig. 49. Anticoagulants.

4. O-A-B blood types.

Agglutinogens – specific antigens, which are situated on the erythrocyte membrane. By chemical nature they are glycoproteins or glycolipids. There is an individual set of specific erythrocyte agglutinogens. Approximately 400 are discovered now. Thirty are mostly common and they can be a reason of reactions after blood transfusion.

Two particular types of antigens are much more likely than the others to cause blood transfusion reactions. They are the O-A-B system of antigens and the Rh system.

There are four major O-A-B blood types (fig. 50), depending on the presence or absence of the two agglutinogens, the A and B agglutinogens. When neither A nor B agglutinogen is present, the blood is type O. When only type A agglutinogen is present, the blood is type A. When only type B agglutinogen is present, the blood is type B. When both A and B agglutinogens are present, the blood is type AB. These combinations of genes are known as the genotypes, and each person is one of the six genotypes.

Agglutinins – specific antibodies, which are dissolved in the blood plasma, they are related to γ-globulins fraction.

When type A agglutinogen is not present in a person’s red blood cells, antibodies known as anti-A agglutinins develop in the plasma. Also, when type B agglutinogen is not present in the red blood cells, antibodies known as anti-B agglutinins develop in the plasma. Thus, note that type O blood, though containing no

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agglutinogens, does contain both anti-A and anti-B agglutinins; type A blood contains type A agglutinogens and anti-B agglutinins; type B blood contains type B agglutinogens and anti-A agglutinins. Finally, type AB blood contains both A and B agglutinogens but no agglutinins (fig. 50).

Fig. 50. ABO blood types.

Agglutinins α and β are formed during the first year of life. Agglutinins are formed to agglutinogens which are absent on the erythrocyte surface (if erythrocytes have agglutinogen A, agglutinin β is formed, if B – α). Agglutinins relate mostly to immunoglobulins M (IgM). They are high-molecular immunoglobulins. IgM are typical hemolysins (when they interact with relative antigens on the erythrocyte membrane, they form substances, which destroy erythrocytes). If similar agglutinogens and agglutinins meet: A with α, B with β – agglutination occurs.

Fig. 51. The prevalence of blood groups in the ABO system.

Agglutination is a process of the clumping together of red blood cells to a particular antibody.

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When blood is mismatched so that anti-A or anti-B plasma agglutinins are mixed with red blood cells that contain A or B agglutinogens, respectively, the red cells agglutinate as a result of the agglutinins attaching themselves to the red blood cells (fig. 52, 53).

Fig. 52. Agglutination process.

Because the agglutinins have two binding sites (IgG type) or 10 binding sites (IgM type), a single agglutinin can attach to two or more red blood cells at the same time, thereby causing the cells to be bound together by the agglutinin. This causes the cells to clump, which is the process of “agglutination.” Then these clumps plug small blood vessels throughout the circulatory system. During ensuing hours to days, either physical distortion of the cells or attack by phagocytic white blood cells destroys the membranes of the agglutinated cells, releasing hemoglobin into the plasma, (hemolysis) of the red blood cells. Sometimes, when recipient and donor blood is mismatched, immediate hemolysis of red cells occurs in the circulating blood. In this case, the antibodies cause lysis of the red blood cells by activating the complement system, which releases proteolytic enzymes that rupture the cell membranes. Immediate intravascular hemolysis is far less common than agglutination followed by delayed hemolysis, because not only does there have to be a high titre of antibodies for lysis to occur, but also a different type of antibody seems to be required, mainly the IgM antibodies; these antibodies are called hemolysins.

Agglutination can be of 2 types:

Direct agglutination.

Inderect agglutination.

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