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Relations between factor VIIa binding and expression of factor VIIa/tissue factor catalytic activity on cell surfaces
D T Le1, S I Rapaport, L V Rao
1Department of Pathology, University of California, San Diego, La Jolla 92093.
The Journal of Biological Chemistry
|August 5, 1992
Summary
Recombinant factor VIIa (rVIIa) binds tissue factor (TF) slowly, but forms active complexes rapidly. Two populations of VIIa/TF complexes exist, differing in activity and inhibition by TFPI.
Area of Science:
- Biochemistry
- Molecular Biology
- Hemostasis
Background:
- Tissue factor (TF) initiates the extrinsic coagulation pathway.
- Recombinant factor VIIa (rVIIa) is used to treat bleeding disorders.
- Understanding VIIa/TF complex kinetics is crucial for hemostasis research.
Purpose of the Study:
- To investigate the binding kinetics of rVIIa to cell surface TF.
- To characterize the expression and activity of VIIa/TF complexes.
- To determine the role of TFPI in modulating VIIa/TF activity.
Main Methods:
- Radioligand binding assays using 125I-rVIIa.
- Enzyme activity assays measuring factor X activation.
- Experiments with active-site inhibited rVIIa.
- Inhibition studies using TFPI/FXa complexes.
Main Results:
- rVIIa binding to TF reached saturation in 30-60 min, but VIIa/TF activity was expressed within 1 min.
- Two populations of VIIa/TF complexes were identified: a minor active population and a major inactive population on intact cells.
- Freeze-thawing monolayers increased VIIa/TF activity significantly.
- TFPI/FXa inhibited both populations, with differing potencies.
Conclusions:
- VIIa/TF complex formation and activity expression are distinct processes.
- Cell integrity influences VIIa/TF complex activity.
- TFPI differentially regulates active and latent VIIa/TF complexes.