Displacement of Fibrin-bound Thrombin by r-Hirudin Precludes the Use of ¹³¹l-r-Hirudin for Detecting Pulmonary Emboli in the Rabbit

 

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Summary

Pulmonary emboli are detectable by filling defects in the pulmonary vasculature upon pulmonary angiography. Emboli derived from venous thrombi are rich in fibrin to which thrombin remains bound. Hirudin, a specific thrombin inhibitor, binds to thrombin to yield a 1:1 stoichiometric complex. We examined whether ¹³¹I-recombinant hirudin (r-hirudin) could be used to detect pulmonary emboli in rabbits. Clots were formed by re-calcifying rabbit plasma in vitro, and then injected (0.034 ml) into a femoral vein to lodge in the lungs. ¹³¹I-r-hirudin (29 ± 4 μCi/kg) was injected intravenously but emboli could not be detected by gamma camera in real time. Post-mortem analysis of lung tissue showed that ¹³¹I-r-hirudin did not associate with emboli prepared with ¹²⁵I-fibrin. Because of these findings, we used different techniques to look at the binding of hirudin to plasma clots. Clots formed in vitro were incubated with ¹³¹I-r-hirudin in the presence of equimolar amounts of ¹²⁵I-albumin; specific binding of ¹³¹I-r-hirudin was not observed. Experiments with immobilized fibrin(ogen) showed that ¹²⁵I-r-hirudin did not bind to and remain with fibrin-bound ¹³¹I-thrombin but did lead to the inactivation and displacement of up to 70% of bound thrombin as r-hirudin-thrombin complex; residual thrombin bound to fibrin remained active. Thus, released r-hirudin-thrombin complex is probably cleared rapidly from the region of the embolus in vivo; radioiodinated r-hirudin may not, therefore, be useful as a marker for detecting emboli.

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