Combined administration of barbourin--albumin and hirudin--albumin fusion proteins limits fibrin(ogen) deposition on the rabbit balloon-injured aorta.
BACKGROUND
There are continuing needs for new antithrombotic agents and procedures. We hypothesized that the slowly cleared recombinant fusion proteins barbourin--albumin (BLAH6) and hirudin--albumin (HLAH6) would be effective in limiting fibrin(ogen) and/or platelet deposition in a rabbit model of arterial injury.


METHODS
Recombinant fusion proteins were expressed in Pichia pastoris fermenter cultures and purified by nickel-chelate affinity chromatography. They were injected intravenously into rabbits prior to blood sampling and platelet aggregometry, assessment of deposition of 125I-fibrin(ogen) and 51Cr-platelet onto the balloon-injured thoracic aorta, electron microscopy (EM) and immunohistochemistry of aortic sections, and determination of bleeding time following a standardized ear incision.


CONCLUSIONS
BLAH6 administration elicited a dose- and time-dependent inhibition of platelet aggregation in post-injection whole blood samples, and reduced both fibrin(ogen) and platelet deposition on the injured aorta, although the former effect was both more durable and more significant than the latter. In contrast, HLAH6 injection reduced fibrin(ogen) but not platelet deposition. Doses of the two proteins ineffective in preventing fibrin(ogen) deposition when given alone were effective when combined, suggesting at least additive effects. Immunohistochemistry and EM supported the radioactive deposition studies, while bleeding times were decreased with combined BLAH6 and HLAH6 administration compared to HLAH6 alone in a rabbit ear bleeding model. The data show that these fusion proteins exert an antithrombotic effect in vivo and may indicate that combined low-dose administration of antiplatelet and antithrombin agents could offer safety advantages in the treatment of thrombosis.
McMaster University, Department of Pathology and Molecular Medicine, HSC 4N66, 1200 Main Street West, Hamilton, Ontario, Canada L8N 3Z5. sheffiel@mcmaster.ca
William P
WP
Sheffield
Sharon
S
Gataiance
Louise J
LJ
Eltringham-Smith
D000418:Albumins / Q000008:administration & dosage / Q000494:pharmacology*; D000818:Animals; D001011:Aorta / Q000293:injuries; D001780:Blood Coagulation Tests; D002404:Catheterization / Q000009:adverse effects; D003435:Crotalid Venoms / Q000008:administration & dosage / Q000494:pharmacology*; D004195:Disease Models, Animal; D004359:Drug Therapy, Combination; D005337:Fibrin / Q000187:drug effects* / Q000378:metabolism; D005343:Fibrinolytic Agents / Q000008:administration & dosage / Q000494:pharmacology*; D006629:Hirudins / Q000008:administration & dosage / Q000494:pharmacology*; D007150:Immunohistochemistry; D008854:Microscopy, Electron; D010973:Platelet Adhesiveness / Q000187:drug effects; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000138:chemical synthesis / Q000627:therapeutic use; D013927:Thrombosis / Q000188:drug therapy
D000418:Albumins; D003435:Crotalid Venoms; D005343:Fibrinolytic Agents; D006629:Hirudins; D011993:Recombinant Fusion Proteins; C068858:barbourin; D005337:Fibrin