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Novel mechanisms of fibrinolysis in health and disease

University of British Columbia

This is a two-part thesis, focusing first on a clinical and then on a biochemical aspect of fibrinolysis, the process that dissolves blood clots. Hyperfibrinolysis: An explanation for reduced cardiovascular disease in hemophilia patients Hemophilia is a coagulation disorder where factor (F) VIII or FIX deficiency results in prolonged bleeding. Therapeutic FVIII or FIX replacement has lengthened hemophilia patient life expectancy. Interestingly, retrospective studies have demonstrated a lower standard mortality risk from cardiovascular disease (CVD) compared to the normal population and enhanced fibrinolytic capability has been proposed as an explanation for this relative protection from CVD. Through the analysis of tissue-type plasminogen activator (tPA), the initiator of fibrinolysis and two inhibitors of fibrinolysis: plasminogen activator inhibitor-1 (PAI-1) and thrombin activatable fibrinolysis inhibitor (TAFI), the current study showed that some hemophilia patients have reduced inhibition of the fibrinolytic pathway compared to age and cardiovascular risk matched controls. A trend of hyperfibrinolysis was also seen in hemophilia patients through accelerated plasma clot lysis with 50% of the patients having at least a 2-fold enhancement, and 22% with at least a 10-fold enhancement. Regulation of Clotting Factor Xa Auxiliary Cofactor Function in Fibrinolysis through β-Peptide Excision We have shown that clotting factor Xa (FXa) cleaved by the fibrinolysis protease, plasmin (Pn), produces consecutive fragments, called FXaβ and Xa33/13. These both have newly exposed C-terminal lysines (Lys) that accelerate tPA in purified clot lysis assays. However, in plasma Xa33/13 rapidly loses this fibrinolytic function due to degradation. It is therefore important to define the role of four possible basic amino acid residues in generating FXaβ; Lys(K)427, Arg(R)429, K433 and K435. Using site directed mutagenesis, K435 was defined as the preferred cleavage site, while R429 was unfavourable. K433 and K435 were found to be important for RVV-X activation and β-peptide cleavage facilitates the production of Xa33/13. Interestingly, when all four basic residues were mutated to Gln (Q), preventing production of FXaβ, an unexpected distal cleavage site was demonstrated to also enhance Pn generation. Replacing the R429 with lysine generated a hyperfibrinolytic species suggesting a potential novel therapeutic approach.

Text

2014-05-16

Attribution-NoDerivs 2.5 Canada

http://hdl.handle.net/2429/46746

Experimental Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nd/2.5/ca/