The Fibrin αC Region is Critical for Mechanical Strength, Resistance to Fibrinolysis and the Formation of a Stable Whole Blood Clot
ISTH Academy. McPherson H. Jul 10, 2019; 273873; OC 74.3 Topic: Fibrinogen & Factor XIII
Helen McPherson
Helen McPherson
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OC 74.3

The Fibrin αC Region is Critical for Mechanical Strength, Resistance to Fibrinolysis and the Formation of a Stable Whole Blood Clot

H. McPherson1, C. Duval1, N. Asquith1, S. Baker1, M. Hindle1, M. Domingues2, V. Ridger3, S. Connell1, K. Naseem1, H. Philippou1, R. Ajjan1, R. Ariëns1
1University of Leeds, Leeds, United Kingdom, 2Faculdade de Medicina da Universidade de Lisboa, Lisbon, Portugal, 3University of Sheffield, Sheffield, United Kingdom

Main Topic: Fibrinolysis and Proteolysis
Category: Fibrinogen & Factor XIII

Background: The fibrinogen αC-region contains FXIII, α2-antiplasmin and plasminogen binding sites, and is comprised of a globular domain (αC-domain) and connector region. We previously showed that truncations of the αC-region changed fibrin clot structure in purified systems. The relative contributions of the connector and the αC-domain to alterations in whole blood clot structure and function are unknown.
Aims: To understand the influence of the αC-domain and connector on clot lysis, mechanics and contraction.
Methods: Recombinant fibrinogens wild type (WT), α390 (lacking αC-domain) and α220 (deficient in αC-region) were produced in CHO cells. Fibrinolysis was studied through turbidity and clot mechanics by magnetic microrheology. Ex-vivo investigations of clot contraction, rotational thromboelastometry (ROTEM) and electron microscopy (SEM) were performed using whole blood from fibrinogen-/- mice reconstituted with recombinant fibrinogens. Results are shown as mean±SD.
Results: WT clots showed a 50% lysis time of 74min (±4.78,n=4), which was similar for α390 (69 ±11.97min,n=4), but α220 lysed significantly faster (18±3.63min,n=4). Clot mechanics for α220 could not be studied without FXIII, as clots were exceptionally weak. With FXIII, α220 clots showed markedly reduced storage modulus (G') (0.021±0.071Pa,n=4) compared to WT (2.4±0.357Pa,n=4), while α390 was slightly elevated (3.5Pa±0.737Pa,n=4). No difference in clot weight, contraction or platelet incorporation was observed comparing WT to α390, while α220 showed no platelet-driven contraction, and no visible clots after 120min. ROTEM displayed reduced clot firmness for both truncations, and increased clotting time for α220. SEM of α220 after ROTEM showed clots mainly composed of activated platelets with limited fibrin present.
Conclusions: Our results show that without αC-domains and connector, fibrin is extremely weak, loses resistance to fibrinolysis and does not support stable clot formation in whole blood. These data indicate that the fibrinogen αC-connector provides critical mechanical strength, stabilises the fibrin network and supports normal clot formation in whole blood.

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