The Characterisation of Thrombus Development in an Improved Model of Arterio-Venous Shunt Thrombosis in the Rat and the Effects of Recombinant Desulphatohirudin (CGP 39393), Heparin, and lloprost

Summary

An existing arterio-venous shunt thrombosis model in the rat has been modified to increase its usefulness for the testing of antithrombotic agents and characterised using morphological and radiometric techniques. The thrombus formed on the cotton thread held in the shunt was found to be composed of platelet aggregates surrounded by red thrombus. After 30 min of blood flow there was a 15-fold increase in the platelet content of the thrombus and a 4-fold increase in the fibrin(ogen) content compared with an equivalent weight of whole blood.

Use of the anticoagulants recombinant desulphatohirudin (CGP 39393, 4 mg/kg, s.c.) and unfractionated heparin (800 IU/ kg, s.c.) showed that approx. 90% inhibition of thrombus weight and approx. 80% inhibition of fibrin(ogen) content could be achieved without significant effect on the platelet content. Conversely, using the platelet inhibitor lloprost (1 Μg kg⁻¹ min⁻¹), a reduction in thrombus weight of 50% was associated with 75% inhibition of platelet content and only 20% inhibition of fibrinogen). These observations suggest that the growth of this type of thrombus is largely the result of continued fibrin formation rather than continued platelet recruitment and activation.

• REFERENCES

• 1 Smith JR, White AM. Fibrin, red cell and platelet interactions in an experimental model of thrombosis. Br J Pharmacol 1982; 77: 29-38
• 2 Shand RA, Smith JR, Wallis RB. Expression of the platelet procoagulant activity in vivo in thrombus formation in an extra-corporeal shunt in the rat. Thromb Res 1984; 36: 223-232
• 3 Talbot MD, Ambler J, Butler KD, Findlay VS, Mitchell KA, Peters RF, Tweed MF, Wallis RB. Recombinant desulphatohirudin (CGP 39393). Anticoagulant and antithrombotic properties in vivo. Thromb Haemostas 1989; 61: 77-80
• 4 Jaques LB. Heparins - anionic polyelectrolyte drugs. Pharmacol Rev 1980; 31: 99-166
• 5 Stürzebecker C-S, Losert W. Effect of lloprost on platelet activation in vitro. In: Prostacyclin and its Stable Analogue lloprost Gryglewski RJ, Stock G. (eds.). Springer-Verlag, Berlin: 1987: 39-45
• 6 Lendrum AC, Fraser DS, Slidders W, Henderson R. Studies on the character and staining of fibrin. J Clin Pathol 1962; 15: 401-413
• 7 Clauss A. Gerinnungsphysiologische Schnellmethode zur Bestim-mung des Fibrinogens. Acta Haematol 1957; 17: 237-246
• 8 Doumas B, Watson W, Biggs H. Albumin standards and the measurement of serum albumin with bromocresol green. Clin Chim Acta 1971; 31: 87-96
• 9 Maguire ED, Wallis RB. In vivo redirection of prostaglandin endoperoxides into 6-keto PGF la formation by thromboxane synthetase inhibitors in the rat. Thromb Res 1983; 32: 15-27
• 10 Dacie JV, Lewis SM. Practical Haematology. Churchill Livingstone, London 1984; 24-25
• 11 Smith LF, Schaible KL, Fessler RG, Rachlin JR, Brown FD. Examination of the utility of the rat as an animal model for human anticoagulation. Haemostasis 1987; 17: 206-210
• 12 Brown J, Goddard P, Petrak K. Soluble polymeric carriers for drug delivery. Part one: free radical polymerisation of N-(2-hydroxypropyl) methacrylamide with N-(2-[hydroxyphenyl] ethyl) acrylamide. J Polymer Sci, Polymer Letts 1989 (in press).
• 13 Packham MA. The behaviour of platelets at foreign surfaces. Proc Soc Exp Biol Med 1988; 189: 261-274
• 14 Davey MG, Luscher EF. Actions of thrombin and other coagulant and proteolytic enzymes on blood platelets. Nature London 1967; 216: 857-858
• 15 Hoffmann A, Markwardt F. Inhibition of the thrombin-platelet reaction by hirudin. Haemostasis 1984; 14: 164-169
• 16 Talbot MD. The biology of recombinant hirudin (CGP 39393): a new prospect in the treatment of thrombosis. Semin Thromb Hemostas 1989; 15: 293-301
• 17 Markwardt F, Kaiser B, Nowak G. Studies on antithrombotic effects of recombinant hirudin. Thromb Res 1989; 54: 377-388
• 18 Kelly AB, Hanson SR, Marzec U, Harker LA. Recombinant hirudin interruption of platelet-dependent thrombus formation. Circulation 1988; 78: 1242
• 19 Heras M, Chesebro JH, Penny WJ, Bailey KR, Badimon L, Fuster V. Effects of thrombin inhibition on the development of acute platelet-thrombus deposition during angioplasty in pigs. Heparin versus recombinant hirudin, a specific thrombin inhibitor. Circulation 1989; 79: 657-665