Neutralization of a Low Molecular Weight Heparin (LHN-1) and Conventional Heparin by Protamine Sulfate in Rats

Summary

The neutralization of a low molecular weight heparin (LHN-1) and conventional heparin (CH) by protamine sulfate has been studied in vitro and in vivo. In vitro, the APTT activity of CH was completely neutralized in parallel with the anti-Xa activity. The APTT activity of LHN-1 was almost completely neutralized in a way similar to the APTT activity of CH, whereas the anti-Xa activity of LHN-1 was only partially neutralized.

In vivo, CH 3 mg/kg and LHN-1 7.2 mg/kg was given intravenously in rats. The APTT and anti-Xa activities, after neutralization by protamine sulfate in vivo, were similar to the results in vitro. In CH treated rats no haemorrhagic effect in the rat tail bleeding test and no antithrombotic effect in the rat stasis model was found at a protamine sulfate to heparin ratio of about 1, which neutralized APTT and anti-Xa activities. In LHN-1 treated rats the haemorrhagic effect was neutralized when APTT was close to normal whereas higher doses of protamine sulfate were required for neutralization of the antithrombotic effect. This probably reflects the fact that in most experimental models higher doses of heparin are needed to induce bleeding than to prevent thrombus formation. Our results demonstrate that even if complete neutralization of APTT and anti-Xa activities were not seen in LHN-1 treated rats, the in vivo effects of LHN-1 could be neutralized as efficiently as those of conventional heparin. The large fall in blood pressure caused by high doses of protamine sulfate alone was prevented by the prior injection of LHN-1.

Key words

LMW heparin - Protamine sulfate

• References

• 1 Shapira N, Schaff HV, Piehler JM, White RD, Sill JC, Pluth JR. Cardiovascular Effects of Protamine Sulfate in Man. J Thorac Cardiovasc Surg 1982; 84: 505-514
• 2 Best N, Sinosich MJ, Teisner B, Grudzinskas JG, Fisher MM. Complement Activation During Cardiopulmonary Bypass by Hepa-rin-Protamine Interaction. Br J Anaesth 1984; 56: 339-343
• 3 Kakkar VV, Murray W JG. Efficacy and Safety of Low-Molecular-Weight Heparin (CY 216) in Preventing Postoperative Venous Thrombo-Embolism: A Cooperative Study. Br J Surg 1985; 72: 786-791
• 4 Bara L, Billaud E, Gramond G, Kher A, Samama M. Comparative Pharmacokinetics of a Low Molecular Weight Heparin (PK 10169) and Unfractioned Heparin after Intravenous and Subcutaneous Administration. Thromb Res 1985; 39: 631-636
• 5 Hubbard AR, Jennings CA. Neutralization of Heparan Sulphate and Low Molecular Weight Heparin by Protamine. Thromb Haemostas 1985; 53: 86-89
• 6 Holmer E, Soderstrom G. Neutralization of Unfractionated Heparin and a Low Molecular Weight (LMW) Heparin Fragment by Protamine. Thromb Haemostas 1983; 50: 103 (Abstr)
• 7 Harenberg J, Gnasso A, de Vries JX, Zimmerman R, Augustin J. Inhibition of Low Molecular Weight Heparin by Protamine Chloride in vivo. Thromb Res 1985; 38: 11-20
• 8 Massonnet-Castel S, Pelissier E, Dreyfus G, Deloche A, Abry B, Guibourt P, Terrier E, Passelecq J, Jaulmes B, Carpentier A. Low-Molecular-Weight Heparin in Extracorporeal Circulation. Lancet May 26 1984; 1182-1183
• 9 Diness V, Nielsen JI, Pedersen PC, Wolffbrandt KH, Østergaard PB. A Comparison of the Antithrombotic and Haemorrhagic Effects of a Low Molecular Weight Heparin (LHN-1) and Conventional Heparin. Thromb Haemostas 1986; 55: 410-414
• 10 Teien AN, Lie M, Abildgaard U. Assay of Heparins in Plasma Using a Chromogenic Substrate. Thromb Res 1976; 8: 413-416
• 11 Reyers I, Mussoni L, Donati MB, de Gaetano G. Failure of Aspirin at Different Doses to Modify Experimental Thrombosis in Rats. Thromb Res 1980; 18: 669-674
• 12 Doutremepuich C, Bonini F, Toulemonde F, Bertrand H, Bayrou B, Quilichini R. In vivo Neutralization of Low-Molecular Weight Heparin Fraction CY 216 by Protamine. Semin Thromb Hemostas 1985; 11: 318-322
• 13 Cade JF, Buchanan MR, Boneu B, Ockelford P, Carter CJ, Cerskus AL, Hirsh J. A Comparison of the Antithrombotic and Haemorrhagic Effects of Low Molecular Weight Heparin Fractions: The Influence of the Method of Preparation. Thromb Res 1984; 35: 613-625
• 14 Pangrazzi J, Abbadini M, Zametta M, Naggi A, Torri G, Casu B, Donati MB. Antithrombotic and Bleeding Effects off a Low Molecular Weight Heparin Fraction. Biochem Pharmacol 1985; 34: 3305-3308
• 15 Cumming AM, Jones GR, Wensley RT, Cundall RB. In Vitro Neutralization of Heparin in Plasma Prior to the Activated Partial Thromboplastin Time Test: An Assessment of four Heparin Antagonists and Two Anion Exchange Resins. Thromb Res 1986; 41: 43-56
• 16 Michalski R, Lane DA, Pepper DS, Kakkar VV. Neutralization of Heparin in Plasma by Platelet Factor 4 and Protamine Sulphate. Br J Haematol 1978; 38: 561-571
• 17 Thomas DP, Barrowcliffe TW, Merton RE, Stocks J, Dawes J, Pepper DS. In vivo Release of Anti-Xa Clotting Activity by a Heparin Analogue. Thromb Res 1980; 17: 831-840
• 18 Bianchini P, Osima B, Parma B, Nader HB, Dietrich CP. Lack of Correlation between “in vitro” and “in vivo” Antithrombotic Activity of Heparin Fractions and Related Compounds. Heparan Sulfate as an Antithrombotic Agent “in vivo”. Thromb Res 1985; 40: 597-607