An Experimental Method for Quantitative Evaluation of Thrombolysis: Effect of Ca-7, a Protease from Aspergillus Oryzae

Abstract

Numerous methods have been described for experimental induction of intravascular thrombi to facilitate study of thrombolytic agents in vivo.1 Insertion, or production, of radioactive 2, 3 and nonradioactive4 clots in a major blood vessel, disruption of the intima by scratching,5 injection of sclerosing agents,6 and occlusion of a venous segment combined with injection of . homologous serum or thrombin7, 8 have been employed frequently. In general, these methods provide a subjective evaluation of dissolution since clots formed are not of standard size nor consistency. Semiconstricting ligatures are often required to confine thrombi produced by these procedures, thus either completely or partially occluding the blood vessels and imparting a variable which is not easily controlled. Extensive surgical manipulation to insert or form clots and to recover them following treatment is a further limitation. Radioactivity or vital dyes9 have been incorporated into clots permitting a degree of quantitation not possible with other methods, but, in this instance, an atypical clot is formed by addition of p31 fibrinogen or dye. Most experimental methods per se do not provide unequivocal evidence of efficacy which can be translated to application of new agents in man with thrombotic disorders. It is considered of paramount importance, therefore, that several different experimental models of clot systems be utilized for evaluation of new thrombolytic agents.A method has been developed which provides for more rigid standardization of clot formation and ease of insertion and recovery following attempts to demonstrate lysis, without severely traumatizing the animal. More precise quantitation of dissolution can be obtained without addition of exogenous agents to the clot and with the further advantage of an extended period for evaluation and comparison of pharmacological effects of thrombolytic agents during the post-treatment period. The preliminary evaluation of CA-7, a protease without plasminogen activator activity, obtained from the growth of Aspergillus oryzae, was undertaken utilizing this method and evidence of efficacy is presented confirming previous reports describing effects with other clot systems in dogs and cats. 16, 17 METHODS Clot formation, Clots are formed from autologous dog blood on a special framework enclosed within a clotting tube. The frame consists of a length of polyethylene at UQ Library on March 15, 2015 ang.sagepub.com Downloaded from