Molecular mechanisms involved in the resistance of fibrin to clot lysis by plasmin in subjects with type 2 diabetes mellitus

Aims/hypothesis: The aim of this study was to determine the influence of type 2 diabetes on fibrinolysis by assessing interactions between the regulatory components of fibrinolysis and the fibrin clot, using fibrinogen purified from 150 patients with type 2 diabetes and 50 matched controls. Methods: Clot lysis rates were determined by confocal microscopy. Plasmin generation was measured using a plasmin-specific chromogenic substrate. Surface plasmon resonance was used to determine the binding interactions between fibrin, tissue-type plasminogen activator (t-PA) and Glu-plasminogen; cross-linkage of plasmin inhibitor to fibrin by factor XIII was determined using a microtitre plate assay. Results: Lysis of diabetic clots was significantly slower than that of controls (1.35 vs 2.92 mu m/min, p < 0.0001) and plasmin generation was significantly reduced. The equilibrium binding affinity between both t-PA and Glu-plasminogen and fibrin was reduced in diabetic subjects: t-PA, K(D)=0.91 +/- 0.3 mu mol/l (control subjects), 1.21 +/- 0.5 mu mol/l (diabetic subjects), p=0.001; Glu-plasminogen, K(D)=97 +/- 19 nmol/l (control subjects), 156 +/- 66 nmol/l (diabetic subjects), p=0.001. Cross-linkage of plasmin inhibitor to fibrin by factor XIII was enhanced in diabetic subjects, with the extent of in vitro cross-linkage correlating with in vivo glycaemic control (HbA(1c)) (r=0.59, p=0.001). Conclusions/interpretations These results indicate that impairment of the fibrinolytic process in diabetic patients is mediated via a number of different mechanisms; these may be a consequence of post-translational modifications to fibrinogen molecules, resulting from their exposure to the abnormal metabolic milieu associated with diabetes.