The Effect of Dimethylbiguanide on Thrombin Activity, FXIII Activation, Fibrin Polymerization, and Fibrin Clot Formation

The Effect of Dimethylbiguanide on Thrombin Activity, FXIII Activation, Fibrin Polymerization, and Fibrin Clot Formation Kristina F. Standeven 1 , Robert A.S. Ariëns 1 , Paul Whitaker 1 , Alison E. Ashcroft 2 , John W. Weisel 3 and Peter J. Grant 1 1 Academic Unit of Molecular Vascular Medicine, University of Leeds, Leeds, U.K. 2 School of Biochemistry and Molecular Biology, University of Leeds, Leeds, U.K. 3 Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania Abstract The antihyperglycemic drug dimethylbiguanide (DMB, also known as metformin) reduces the risk of cardiovascular complications in type 2 diabetes, although the mechanism(s) involved are unclear. DMB reduces glycosylation-related protein cross-linking, a process similar to fibrin cross-linking catalyzed by activated factor XIII (FXIII). To investigate whether the cardioprotective effect of DMB could be related to effects on clot stabilization, we studied the effects of DMB on FXIII, thrombin activity, and cleavage of fibrin(ogen). Activity of purified and plasma FXIII was inhibited by DMB. Analysis by mass spectrometry and FXIII-coupled magnetic particles excluded binding of DMB to FXIII. Thrombin-induced cleavage of the activation peptide from FXIII was inhibited in a dose-dependent manner, as was fibrinopeptide cleavage from fibrinogen. Ancrod-induced cleavage of fibrinopeptide A was not affected. DMB prolonged clotting time of normal plasma. Fiber thickness and pore size of fibrin clots, measured by permeation experiments and visualized by scanning electron microscopy, decreased significantly with DMB. No interactions between DMB and the active site of thrombin were found. Turbidity experiments demonstrated that DMB changed polymerization and lateral aggregation of protofibrils. These results suggest that DMB interferes with FXIII activation and fibrin polymerization, but not only by binding to thrombin on a different location than the active site. In patients on DMB therapy, FXIII antigen and activity levels in vivo were reduced over a 12-week period. These findings indicate that part of the cardioprotective effect of DMB in patients with type 2 diabetes may be attributed to alterations in fibrin structure/function. Footnotes Address correspondence and reprint requests to Robert A.S. Ariëns, Unit of Molecular Vascular Medicine, University of Leeds, G-Floor, Martin Wing, Leeds General Infirmary, Leeds LS1 3EX, U.K. E-mail: r