Impact of complement regulator factor H on thrombin’s role in fibrin clot formation and the anticoagulant protein C pathway

McCluskey, Genevieve 2021. Impact of complement regulator factor H on thrombin’s role in fibrin clot formation and the anticoagulant protein C pathway. PhD Thesis, Cardiff University. Item availability restricted.

	PDF (PhD Thesis) - Accepted Post-Print Version Download (6MB)
	PDF (Cardiff University Electronic Publication Form) - Supplemental Material Restricted to Repository staff only Download (76kB)

Abstract

Complement is part of innate immunity in blood plasma and contributes to eliminating pathogens and cellular debris from the host system. Coagulation is involved in haemostasis, enabling clotting after injury to a blood vessel. Both pathways are evolutionarily linked and composed of effector proteins called zymogens and regulators. Recently, increasing evidence has demonstrated the molecular interactions, or crosstalk, between complement and coagulation. Understanding the importance of the molecular crosstalk is key to further determine the impact of its dysregulation in diseases. A key step in the coagulation system is the generation of the enzyme thrombin which further enhances the pathway, and cleaves fibrinogen into fibrin to form a clot, preventing fluid loss. Thrombin is also tightly regulated to prevent excess thrombi formation, for instance through its interaction with endothelial membrane bound cofactor thrombomodulin, enabling protein C activation which downregulates upstream coagulation factors. Complement factor H, a key regulator of the alternative pathway in complement activation in the fluid phase and on cell surfaces, has been shown to interact with thrombomodulin, as well as other coagulation components such as factor XII, factor XIII, von Willebrand factor and platelets. However, the role and involvement of factor H in coagulation activation and regulation remains poorly understood. The aim of this work was to analyse the impact of factor H on thrombin’s anticoagulant role in protein C activation, in the presence and absence of thrombomodulin, as well as on its procoagulant role, in the cleavage of fibrinogen into fibrin. Therefore, I developed biochemical assays to assess activated protein C generation, and thrombin-mediated fibrin clot generation in a pure protein system and in plasma, in the presence of factor H. Finally, I investigated the binding sites on factor H using SPR and binding assays supported by computational modelling. Factor H enhanced protein C activation by the thrombin/thrombomodulin complex but also by thrombin alone. It also enhanced the rate of fibrin clot formation and altered the structure of the clot. Absence of factor H in plasma increased clotting time and restoration of physiological levels decreased it significantly. Importantly, it was determined that thrombomodulin, and primarily thrombin, are ligands for factor H, and these interactions mediated these functional effects. It was also showed that the C-terminal domain of factor H is one binding site involved in the interaction with thrombin. To conclude, factor H could be a potential novel ligand for thrombomodulin and thrombin, regulating its pro and anticoagulant roles. This is relevant in iv diseases such as atypical hemolytic uremic syndrome (aHUS) where complement and coagulation are dysregulated due to mutations in factor H.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Pharmacy
Subjects:	Q Science > Q Science (General)
Date of First Compliant Deposit:	29 September 2021
Last Modified:	10 Dec 2022 02:23
URI:	https://orca.cardiff.ac.uk/id/eprint/144514

Actions (repository staff only)

Edit Item