Background: FSAP is a plasma protease involved in thrombosis, haemostasis and inflammation. A single nucleotide polymorphism (SNP) in the FSAP gene (“Marburg I”), with a strongly reduced proteolytic activity, has been associated with thrombotic events, but a role in venous thrombosis (VT) is uncertain.
Aims: We developed a kinetic assay to measure FSAP generation directly in plasma and demonstrate its suitability in plasma samples of patients with VT.
Methods: FSAP generation was measured with a specific fluorogenic substrate (Ac-Pro-DTyr-Lys-Arg-AMC), using histones as activation trigger. Two-chain FSAP served as calibrator in the FSAP generation assay (FGA), pooled plasma was used as internal control and for normalization. The raw fluorescence curves were transferred into a TGA software package (Technoclone, Vienna) to determine lag phase, time to peak (TtPeak), velocity, peak FSAP and endogenous FSAP potential (EFP).
Results: The histone-triggered FGA is highly FSAP-specific with no interference by other proteases like clotting factors or fibrinolytic enzymes. A good linearity was seen for TtPeak, peak FSAP and EFP. The assay was found suitable for heparinized plasma, when treated with polybrene, and clearly identified Marburg I carrier. Plasma of postmenopausal women with or without a history of VT (each group n=50) showed highly variable results, with only weak, non-significant differences between groups. Marburg I carrier were equally present in both groups and when excluding these from statistical evaluation, a delayed FSAP generation became significantly different between groups.
Conclusion(s): The FGA is robust and reproducible, measuring various aspects of FSAP activity in plasma. In a one-step assay design, the contribution of FSAP antigen, sequence variations and inhibition/inactivation are captured. The Marburg I-SNP did not correlate with history of VT. Focussing on this SNP may hamper risk stratification regarding FSAP activity and thrombosis, and may explain the difficulty in finding a clear consensus on FSAP and VT in the past.
To cite this abstract in AMA style:Etscheid M, Hanschmann K, Sandset P, Kanse S. Measuring Factor VII Activating Protease (FSAP) in venous thrombosis using a newly developed FSAP generation assay [abstract]. https://abstracts.isth.org/abstract/measuring-factor-vii-activating-protease-fsap-in-venous-thrombosis-using-a-newly-developed-fsap-generation-assay/. Accessed February 20, 2024.
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