Blood Thrombin

Two hypotheses exist for the origin of factor Vila. In one, factor VII is considered to have some proteolytic activity prior to a proteolytic cleavage. In the other view, low concentrations of thrombin and/or factor Xa are considered present at all times in the circulating blood. Thrombin, factor Xa, and factor Vila can activate factor VII to... 

Dismption of the endothehal surface of blood vessels expose coUagen fibers and connective tissue. These provide surfaces that promote platelet adherence, platelet release reaction, and subsequent platelet aggregation. Substances Hberated from the platelets stimulate further platelet aggregation, eg, adenosine diphosphate maintain vasoconstriction, eg, serotonin and participate in blood coagulation, eg, platelet Factors III and IV. In addition, the release reaction modifies platelet membranes in a manner that renders phosphoHpid available for coagulation. The thrombin [9002-04-4] elaborated by the coagulation mechanism is a potent agent in the induction of the platelet release reaction. 

Factor V. High in sialic acid content. Factor V is a large asymmetric single-chain glycoprotein that becomes an active participant in the coagulation cascade when it is converted to its active form by a-thrombin. Approximately 25% of human Factor V is found in the whole blood associated with platelets. Factor V is an essential cofactor along with Factor Xa plus phosphohpid plus Ca " in the conversion of prothrombin to thrombin. 

Factor XIII. Factor XIII circulates in the blood as a zymogen composed of two pairs of different polypeptide chains designated A and B. Inert Factor XIII has a molecular weight of 350,000 daltons and is converted to its active transglutaminase form in the presence of thrombin and calcium. Activated Factor XIII, Xllla, induces an irreversible amide exchange reaction between the y-glutamine and S-lysine side chains of adjacent fibrin... 

Calcium is essential to several steps in the enzyme cascade of the blood clotting process, such as the conversion of prothrombin to thrombin (23). Clotting can be inhibited in stored blood suppHes by addition of complexing agents such as EDTA or citrate which reduce the levels of the free ion, Ca(Il). 

Thrombin (from bovine blood plasma) [9002-04-4] Mj 32,600 [EC 3.4.4.13]. Purified by chromatography on a DEAE-cellulose column, while eluting with O.IM NaCl, pH 7.0, followed by chromatography on Sephadex G-200. Final preparation was free from plasminogen and plasmin. [Yin and Wessler J Biol Chem 243 112 796S.]... 

Thrombin from bovine blood was purified by chromatography using p-chlorobenzylamino-e-aminocaproyl agarose, and gel filtration through Sephadex G-25. [Thompson and Davie Biochim Biophys Acta 250 210 7977.]... 

Prior to May of 1998 when the commercial product was approved by the FDA in the United States, surgeons in this country formed fibrin sealant by using topical bovine thrombin, which is a commercially available product, together with concentrated fibrinogen most frequently obtained from the blood bank. Standard blood bank cryoprecipitate is a good source of concentrated fibrinogen. Also... 

Daniels, T.M. and Fisher et al., PK., Antibodies to bovine thrombin and coagulation factor V associated with the use of topical bovine thrombin or fibrin glue a frequent finding. Blood, 82, 59a (1993). 

FIGURE 15.5 The cascade of activation steps leading to blood clotting. The intrinsic and extrinsic pathways converge at Factor X, and the final common pathway involves the activation of thrombin and its conversion of fibrinogen into fibrin, which aggregates into ordered filamentous arrays that become cross-linked to form the clot. 

The formation of a platelet aggregate requires the recruitment of additional platelets from the blood stream to the injured vessel wall. This process is executed through a variety of diffusible mediators which act through G-protein-coupled receptors. The main mediators involved in this process are adenosine diphosphate (ADP), thromboxane A2 (TXA2), and thrombin (factor Ila). These mediators of the second phase of platelet activation are formed in different ways. While ADP is secreted from platelets by exocytosis, the release of TXA2 follows its new formation in activated platelets. Thrombin can be formed on the surface of activated platelets (see Fig. 2). 

Hirudin is a polypeptide derived from the saliva of the leech Hirudo medicinalis that binds to the blood serine proteinase, thrombins, and thus blocks clot formation. 

The release of NO from the endothelium is induced by various chemical substances, including acetylcholine polypeptides such as substance P, bradykinin, and arginine vasopressin histamine ATP/ADP a2-adrenoceptor agonists thrombin and Ca2+ iono-phores. NO formed in response to mechanical stimuli like shear stress or transmural pressure plays an important role in maintaining basal blood flow. Endothelial NO causes vasodilatation, decreased... 

Heparin inhibits the formation of fibrin clots, inhibits the conversion of fibrinogen to fibrin, and inactivates several of the factors necessary for the clotting of blood. Heparin cannot be taken orally because it is inactivated by gastric acid in the stomach therefore, it must be given by injection. Heparin has no effect on clots that have already formed and aids only in preventing the formation of new blood clots (thrombi). The LMWHs act to inhibit clotting reactions by binding to antithrombin HI, which inhibits the synthesis of factor Xa and the formation of thrombin. 

The serine proteases are the most extensively studied class of enzymes. These enzymes are characterized by the presence of a unique serine amino acid. Two major evolutionary families are presented in this class. The bacterial protease subtilisin and the trypsin family, which includes the enzymes trypsin, chymotrypsin, elastase as well as thrombin, plasmin, and others involved in a diverse range of cellular functions including digestion, blood clotting, hormone production, and complement activation. The trypsin family catalyzes the reaction ... 

Figure 51-1. The pathways of blood coagulation. The intrinsic and extrinsic pathways are indicated. The events depicted below factor Xa are designated the final common pathway, culminating in the formation of cross-linked fibrin. New observations (dotted arrow) include the finding that complexes of tissue factor and factor Vila activate not only factor X (in the classic extrinsic pathway) but also factor IX in the intrinsic pathway, in addition, thrombin and factor Xa feedback-activate at the two sites indicated (dashed arrows). (PK, prekallikrein HK, HMW kininogen PL, phospholipids.) (Reproduced, with permission, from Roberts HR, Lozier JN New perspectives on the coagulation cascade. Hosp Pract [Off Ed] 1992Jan 27 97.)...

The Final Common Pathway of Blood Clotting Involves Activation of Prothrombin to Thrombin... 

This is an insoluble gelatin foam produced by whisking warm gelatin solution to a uniform foam, wtiieh is then dried. It ean be cut into suitable shapes, paeked in metal or paper containers and sterilized by dry heat (150°C for 1 hour). Moist heat destroys the physical properties of the material. Immediately before use, it ean be moistened with normal saline eontaining thrombin. It behaves as a meehanieal haemostat providing the ftamewoik on wtiieh blood elotting ean oeeur. 

This is a dry sponge of human fibrin prepared by elotting a foam of human fibrinogen solution with human thrombin. It is then freeze-dried, cut into shapes and sterilized by dry heat at 130°C for 3 hours. Before use, it is saturated with thrombin solution. Blood coagulation occurs in contact with the thrombin in the interstices of the foam. 

Serum concentrations of D-dimer, a by-product of thrombin generation, is usually elevated. The patient may have an elevated erythrocyte sedimentation rate (ESR) and white blood cell (WBC) count. 

Activated partial thromboplastin time aPTT is performed by adding calcium phospholipids and kaolin to citrated blood and measures the time required for a fibrin clot to form. In this manner, aPTT measures the activity of intrinsic and common pathways. Prolongation of aPTT may be due to a deficiency or inhibitor for factors II, V, VIII, IX, X, XI, and XII. It also may be due to heparin, direct thrombin inhibitors, vitamin K deficiency, liver disease, or lupus anticoagulant. 

The fluidity of blood is a result of the inhibition of a complex series of enzymic reactions in the coagulation cascade (see Fig. 10). When triggered either intrinsically (by contact with foreign surfaces ), or extrinsically (by tissue factors from damaged cells), inactive proenzymes (factors XII, XI, IX, and X) are transformed into activated pro-teinases (XHa, XIa, IXa, and Xa, respectively). Each proteinase catalyzes the activation of the following proenzyme in the sequence, up to formation of thrombin (Factor Ha), another proteinase that catalyzes partial... 

The concept that different structural domains on the heparin chains are principally involved for optimal activity in the foregoing interactions could not be perceived in early work on structure-activity correlations, because the activity of heparin has been most frequently evaluated only with whole-blood-clotting tests (such as the U.S.P. assay). Development of assays for specific clotting-factors (especially Factor Xa and thrombin) has permitted a better insight into the mechanism of action of heparin at different levels of the coagulation cascade. 

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