One of the main considerations in thrombolytic enzyme therapy with plasmin, streptokinase, and urokinase is the creation of a high local enzyme concentration in the vicinity of the thrombus. Different approaches have been suggested to accomplish this. Angiographic methods are used, making it possible to infuse the enzyme solution directly to the thrombus.' Targeting of enzyme to the thrombus using microcontainers, such as microcapsules, liposomes, erythrocytes, or other cells, also can be useful, but requires the creation of a special system for drug release in the target zone. On the other hand, "polymeric affinity drugs," developed principally in the late 1970s by H. Ringsdorf, E. Goldberg, and others, can be of practical importan~e.~.' These affinity drugs consist of a polymeric carrier with coimmobilized enzyme and a vector molecule capable of recognizing and binding to a target site in the organism. Normally, specific antibodies are used as vector molecules; but some other compounds also can be used. This affinity approach also can be of practical use in the case of thrombolytic (fibrinolytic) enzymes.In in vitro experiments, we have studied the fibrinolytic activity of a model system toward fibrin clots. The model system consists of a dextran carrier, activated by periodate oxidation, and a proteolytic enzyme, a-chymotrypsin, and a polyclonal antibody towards fibrinogen coimmobilized on the carrier (FIG. 1). These antibodies participate in immunochemical reactions with fibrinogen, fibrin, and their degradation products and bind with common antigenic determinants.Protein binding studies with oxidized dextran (molecular weight 35,00Cr50,000) were performed as described in Torchilin et aL4 Both the enzyme and antibody immobilization reactions can be performed simultaneously, using an enzyme-toantibody mixture in a molar ratio of 1O:l. Or, the immobilization can be done in two stages. In the first stage the enzyme is bound, and in the second stage the antibody is bound via the remaining aldehyde groups. In our work, the total number of aldehyde groups was 20-24 per 100 glycoside units. Unbound proteins were separated by gel chromatography. As a result, macromolecular conjugates were obtained with molecular weights of about 250,000 and possessing both enzymatic and immunological activity. The enzymatic activity was measured by following the initial hydrolysis rate of a low-molecular-weight specific substrate, N-acetyl-L-tyrosine ethyl ester, on a pH-stat. It was demonstrated that the high-molecular-weight conjugates contained -10% of the initial enzymatic activity introduced into the immobilization reaction.This corresponded to about 50 mg of active enzyme per gram of carrier.The following preparations were used in experiments on fibrin clot lysis: native a-chymotrypsin, dextran-chymotrypsin conjugate, and dextran-chymotrypsin-antibody conjugate. Fibrin clots were prepared by mixing 0.5 ml of 10 mg/ml fibrinogen 289
The role of plasma fibronectin in hemostasis system in erysipelas, depending on the disease gravity and pronounced hemorrhagic syndrome, is studied. The results obtained indicate that different phases of disseminated intravascular blood coagulability syndrome are determined, depending directly on the disease gravity in patients with erysipelas at the height of the disease in the presence of the decrease of plasma fibronectine. The decrease of plasma fibronectine in heavy infectious disease is connected with its consumption in the process of intravascular microthrombogenesis and presents a unique protective response of the macroorganism to the action of pathogene toxins.
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