Contact activation of coagulation depends on the maximal lipophilic trigger concentration

Contact activation of hemostasis is of great clinical importance. Intrinsic coagulation starts upon blood matrix change, resulting in activation of factor XII (F XII) to F XIIa and/or of prekallikrein to kallikrein. The intrinsic system is very complex. There are many reactions that can increase or decrease the generation of F Xa/thrombin. Currently, there are two main trigger types that activate the intrinsic system: (delta)-negatively charged molecules and lipophilic molecules. Recently, it was shown that the stimulation of thrombin generation by (delta)-negatively charged molecules depends on their maximal plasma concentration prior to plasma dilution. The questions arise whether this is also true for lipophilic triggers. Fifty-microliter frozen/thawed pooled normal platelet-poor citrated plasma (PNP) were supplemented with up to 5% (final concentration) hexane, followed by repetitive 1 + 1 dilutions on polystyrene U-wells microtiter plates of high quality (Brand, Wertheim, Germany; article number 781600). Immediately thereafter, the recalcified coagulation activity assay was started and the approximate 200% stimulatory concentrations (approx. SC200s) on intrinsic thrombin generation were determined, the 100% control being unsupplemented PNP. The higher the maximal concentration of hexane prior to dilution, the higher the approx. SC200. If the maximal hexane concentration prior to dilution was higher than 2%, PNP had an approx. SC200 of 0.1-0.2% (8-15 mmol/l) hexane. At maximal hexane concentrations prior to dilution in the range of 0.1-1%, the approx. SC200 decreased 10-fold to about 0.01-0.02%. Up to about 0.1% maximal hexane concentration prior to dilution the corresponding approx. SC200 of hexane on intrinsic thrombin generation was 0.003-0.01% (0.2-0.8 mmol/l). Both main types of contact triggers - negatively charged or lipophilic molecules - have a peculiar behavior respective to maximal plasma concentration/thrombin generation: if the maximal plasma concentration of the trigger prior to plasma dilution is high, then the thrombin generating system needs rather high amounts of triggers to reach approx. SC200; and, if the maximal plasma concentration of the trigger prior to dilution is low, then thrombin is easily generated with a low approx. SC200. This means that the plasmatic F XII/prekallikrein/HMWK system could be inhibited by high plasma concentrations of any trigger and this inhibition cannot be reversed by plasma dilution. To study the action of drugs on the intrinsic system of plasma, plasma should be supplemented with the respective drug at maximal concentrations that are in the range of the maximal blood concentrations obtained in clinical medicine.