Monitoring Hypocoagulant Conditions in Rat Plasma: Factors Determining the Endogenous Thrombin Potential of Tissue Factor-Activated Plasma
Cécile M. A. Nieuwenhuys
1
From the Department of Human Biology, The Netherlands
,
Marion A. H. Feijge
2
The Department of Biochemistry, University of Maastricht, The Netherlands
,
Suzette Béguin
2
The Department of Biochemistry, University of Maastricht, The Netherlands
,
Johan W. M. Heemskerk
1
From the Department of Human Biology, The Netherlands
2
The Department of Biochemistry, University of Maastricht, The Netherlands
› InstitutsangabenWe thank R. F. G. Offermans and R. van Oerle for analytical assistance. We thank Drs. H. C. Hemker, G. Hornstra and G. Tans for valuable comments and discussions. We acknowledge Dr. D. Rijkers for preparation of MVA-pNA. This study was supported by Hoffmann-La Roche (Basel, Switzerland).
Automated human plasma, continuous monitoring of the formation and inactivation of thrombin during the coagulation process provides an adequate way to detect hypo- and hypercoagulant conditions. Here, we describe an analogous procedure to determine the endogenous thrombin potential (ETP), i. e. the free thrombin concentration-time integral, of coagulating rat plasma. When activated with tissue factor, the ETP of plasma from Wistar rats was comparable to the ETP of human plasma, in spite of a relatively short half-life time of free thrombin in rat plasma. The ETP was highly sensitive to heparin as well as to administration of vitamin K antagonist or feeding of the animals with a vitamin K-deficient diet. In plasma that was activated under sub-optimal conditions (reduced levels of tissue factor or vitamin K-dependent coagulation factors), the ETP increased with the rate of thrombin formation in the first minutes of the coagulation process. Since both parameters are dependent of the prothrombin concentration, it appears that this level plays an important role in determining both the initial and total activity of the coagulation system. Thus, automated measurement of free thrombin during the coagulation process of rat plasma allows a detailed analysis of hypocoagulability in this animal model.
Key words
Endogenous thrombin potential -
hypocoagulability -
rat -
tissue factor -
vitamin K
References
1
Mann KG.
Biochemistry and physiology of blood coagulation. Thromb Haemost 1999; 82: 165-74.
3
Hemker HC,
Willems GM,
Béguin S.
A computer assisted method to obtain the prothrombin activation velocity in whole plasma independent of thrombin decay processes. Thromb Haemost 1986; 56: 9-17.
5
Hemker HC,
Wielders S,
Kessels H,
Béguin S.
Continuous registration of thrombin generation in plasma: its use for the determination of the thrombin potential. Thromb Haemost 1993; 70: 33-41.
6
Wielders S,
Mukherjee M,
Michiels J,
Rijkers DTS,
Cambus JP,
Knebel RWC,
Kakkar V,
Hemker HC,
Béguin S.
The routine determination of the endogenous thrombin potential, first results in different forms of hyperand hypocoagulability. Thromb Haemost 1997; 77: 629-36.
7
Bernat A,
Mares AM,
Defreyn JP,
Herbert JM.
Effect of various antiplatelet agents on acute arterial thrombosis in the rat. Thromb Haemost 1993; 70: 812-6.
8
Eriksson BI,
Carlsson S,
Halvarsson M,
Risberg B,
Mattsson C.
Antithrombotic effect of two low molecular weight thrombin inhibitors and a low-molecular weight heparin in a caval vein thrombosis model in the rat. Thromb Haemost 1997; 78: 1404-7.
9
Andriamampandry MD,
Leray C,
Freund M,
Cazenave JP,
Gachet C.
Antithrombotic effects of (n-3) polyunsaturated fatty acids in rat models of arterial and venous thrombosis. Thromb Res 1999; 93: 9-16.
10
Nieuwenhuys CMA,
Hornstra G.
The effects of purified eicosapentaenoic and docosahexaenoic acids on arterial thrombosis tendency and platelet function in rats. Biochim Biophys Acta 1998; 1390: 313-22.
12a
Tans G,
Janssen-Claessen T,
Hemker HC,
Zwaal RFA,
Rosing J.
Meizothrombin formation during factor Xa-catalyzed prothrombin activation. Formation in a purified system and plasma. J Biol Chem 1991; 266: 21864-73.
13
Rijkers DTS,
Wielders SJH,
Béguin S,
Hemker HC.
Prevention of the influence of fibrin and α2-macroglobulin in continuous measurement of the thrombin potential: implications for an endpoint determination of the optical density. Thromb Res 1998; 89: 161-9.
15
Groenen-van Dooren MMCL,
Ronden JE,
Soute BAM,
Vermeer C.
Bioavailability of phylloquinone and menaquinones after oral and colorectal administration in vitamin K-deficient rats. Biochem Pharmacol 1995; 50: 797-801.
17
Pratt SK,
Win MJ,
Park BK.
The disposition of enantiomers of warfarin following chronic administration to rats: relationship to anticoagulant response. J Pharm Pharmacol 1989; 41: 743-6.
18
Béguin S,
Kessels H,
Dol F,
Hemker HC.
The consumption of antithrombin III during coagulation, its consequences for the calculation of prothrombinase activity and the standardisation of heparin activity. Thromb Haemost 1992; 68: 136-42.
19
Gauthier F,
Genell S,
Mouray H,
Ohlsson K.
Interactions in vitro and in vivo between rat serum protease inhibitors and anodal and cathodal rat trypsin and chymotrypsin. Biochim Biophys Acta 1979; 566: 200-10.
21
Craciun AM,
Groenen-van Dooren MMCL,
Thijssen HHW,
Vermeer C.
Induction of prothrombin synthesis by K-vitamins compared in vitamin K-deficient and in brodifacoum-treated rats. Biochim Biophys Acta 1998; 1380: 75-81.
22
Vainieri H,
Wingard LJ.
Effect of warfarin on the kinetics of the vitamin K-dependent coagulation factors in rats. J Pharmacol Exp Ther 1977; 201: 507-17.
25
Nicolaes GAF,
Thomassen MCLGD,
van Oerle R,
Hamulyàk K,
Hemker HC,
Tans G,
Rosing J.
A prothrombinase-based assay for detection of resistance to activated protein C. Thromb Haemost 1996; 76: 404-10.
