PubMed · 5721946
The source did not provide an abstract. Follow the original record for more information.
Explore connections, maps & timelines
I Maxwell. 1968-11-30. Prothrombin estimations.. https://pubmed.ncbi.nlm.nih.gov/5721946/
Cite the original work for its findings. Save a collection to share your selection of sources.
KEEP EXPLORING
Contact system activation, in vitro, is triggered by activation of factor XII (FXII) on binding to an activator, such as negatively charged surfaces. A putative surface-binding site of FXII has been located within the amino acid residues 1-28 by identifying the epitope recognized by a monoclonal antibody (MoAb), B7C9, which inhibits kaolin-induced clotting activity. To further elucidate the role of the amino terminal binding site in the regulation of FXII activation, we have characterized a FXII recombinant protein (rFXII-triangle up19) deleted of the amino acid residues 3-19, which are encoded by the second exon of FXII gene. A plasmid encoding for rFXII-triangle up19 was constructed and expressed in HepG2 cells by using vaccinia virus. Purified rFXII-triangle up19 migrated as a single band of Mr 77,000 on sodium dodecyl sulfate (SDS)-polyacrylamide gel, did not bind to MoAb B7C9 immobilized on Protein A-Sepharose, thus confirming that it lacked the epitope for this MoAb, and had no amidolytic activity towards the chromogenic substrate S-2302 in the absence of activator. rFXII-triangle up19 specific clotting activity was lower (44%) than that of native FXII. The activation rate of rFXII-triangle up19 by kallikrein in the absence of dextran sulfate was about four times higher than that of full-length FXII and was increased in the presence of dextran sulfate. However, rFXII-triangle up19 underwent autoactivation in the presence of dextran sulfate. Labeled rFXII-triangle up19 bound to kaolin, which binding was equally well inhibited by either, rFXII-triangle up19 or full-length FXII (IC50 = 7.2 +/- 2.2 nmol/L for both proteins). Accordingly, a synthetic peptide corresponding to FXII amino acid residues 3-19 did not inhibit the binding of labeled full-length FXII to kaolin. rFXII-triangle up19 generated a similar amount of FXIIa- and kallikrein-C1-inhibitor complexes in FXII-deficient plasma in the presence of kaolin, as did full-length FXII; but generated less factor XIa-C1-inhibitor complexes (50%) than full-length FXII. This impaired factor XI activation by rFXII-triangle up19a was also observed in a purified system and was independent of the presence of high molecular weight kininogen. Furthermore, the synthetic peptide 3-19, preincubated with factor XI, inhibited up to 30% activation of factor XI both in the purified system as well as in plasma. These results together indicate that amino acid residues 3-19 of FXII are involved in the activation of factor XI and do not contribute to the binding of FXII to negatively charged surfaces.
We examined the spatial dynamics of in vitro clot growth in human blood and plasma and found that initially, a clot grows at a constant speed, then abruptly stops and becomes surrounded by an 'inhibition zone' in which coagulation is strongly suppressed. We also observed the formation of 'stratified structures' (target patterns) in which solid layers alternated with liquid plasma. These and other spatial regimes of clotting are explained in terms of two interacting concentration waves propagating without attenuation. The experimental results are consistent with a hypothesis that blood is a bi-excitable medium, a new type of excitable medium.
The dynamics of clot formation was studied in a two-compartment chamber designed to allow free diffusion of thrombin according to its concentration gradient into nonstirred citrate plasma or fibrinogen solution. Fibrin clots in fibrinogen solutions increased progressively until the substrate was depleted. In plasma, the clot weight dynamics significantly depended on the concentration of thrombin in the thrombin compartment. When the thrombin concentrations were extremely low (25-40 nM), the clot weight increased throughout the experiment (sometimes 20-24 h). At higher thrombin concentrations, the clot weight increased for 1-2 h and then stopped growing for the following 3-4 h. The clot weight observed at the plateau varied only slightly in the range of thrombin concentrations of 50-770 nM. In this range, high thrombin concentrations (250-770 nM) caused a second increase in the clot weight 4-8 h after the start of diffusion, which was followed by the second plateau in the curve of clot weight against time. The time to the plateau and the plateau duration decreased with increasing thrombin concentrations. The abundant plasma inhibitors of thrombin cannot account for these results. It was hypothesized that an as yet unknown mechanism is responsible for the inhibition of clot growth.