Modification of Fibrinogen by Homocysteine Thiolactone Increases Resistance to Fibrinolysis:  A Potential Mechanism of the Thrombotic Tendency in Hyperhomocysteinemia

Abstract

We have previously shown functional differences in fibrinogen from hyperhomocysteinemic rabbits compared to that in control rabbits. This acquired dysfibrinogenemia is characterized by fibrin clots that are composed of abnormally thin, tightly packed fibers with increased resistance to fibrinolysis. Homocysteine thiolactone is a metabolite of homocysteine (Hcys) that can react with primary amines. Recent evidence suggests that Hcys thiolactone-lysine adducts form in vivo. We now demonstrate that the reaction of Hcys thiolactone with purified fibrinogen in vitro produces fibrinogen (Hcys fibrinogen) with functional properties that are strikingly similar to those we have observed in homocysteinemic rabbits. Fibrinogen purified from homocysteinemic rabbits and Hcys fibrinogen are similar in that (1) they both form clots composed of thinner, more tightly packed fibers than their respective control rabbit and human fibrinogens; (2) the clot structure could be made to be more like the control fibrinogens by increased calcium; and (3) they both form clots that are more resistant to fibrinolysis than those formed by the control fibrinogens. Further characterization of human fibrinogens showed that Hcys fibrin had similar plasminogen binding to that of the control and an increased capacity for binding tPA. However, tPA activation of plasminogen on Hcys fibrin was slower than that of the control. Mass spectrometric analysis of Hcys fibrinogen revealed twelve lysines that were homocysteinylated. Several of these are close to tPA and plasminogen binding sites. Lysines are major binding sites for fibrinolytic enzymes and are also sites of plasmin cleavage. Thus, modification of lysines in fibrinogen could plausibly lead to impaired fibrinolysis. We hypothesize that the modification of lysine by Hcys thiolactone might occur in vivo, lead to abnormal resistance of clots to lysis, and thereby contribute to the prothrombotic state associated with homocysteinemia.

Ablation of MMP9 Gene Ameliorates Paracellular Permeability and Fibrinogen–Amyloid Beta Complex Formation during Hyperhomocysteinemia

“…Presence of high level of Hcy can further increase the complex's resistance to fibrinolysis. 36 We previously found that elevated level of Hcy decreased EC layer integrity (defined by changes in transendothelial electrical resistance) suggesting the wider opening of gaps between the ECs. 37 The present data confirmed the results of our other study where we showed that HHcy increased overall pial venular protein leakage but were unable to differentiate paracellular from transcellular transport.…”

“…It has been shown that clots formed from homocysteinylated Fg are more resistant to lysis than clots formed from native Fg. 36,45 Therefore, activation of MMP9 has lesser effect on Fg clots or Fg-Ab complexes during HHcy than during normal level of Hcy. In addition, Hcy potentiates Ab-induced neurodegeneration affecting cytosolic calcium.…”

Coagulology, biochemical profile and muscle pathology in calves diagnosed with nutritional muscular dystrophy

“…Studies in humans with Duchenne muscular dystrophy did not reveal changes in fibrinogen concentrations (Saito et al 2001). Other authors (Sauls et al 2006) observed that homocysteine thiolactone (HTL) can modify the properties of fibrinogen and disrupt its biological functions through N-homocysteinylation. The results of an in vitro study demonstrated that fibrinogen is easily modified by HTL.…”

Homocysteine and its thiolactone-mediated modification of fibrinogen affect blood platelet adhesion

“…Moreover, lysines are major binding sites for fibrinolystic enzymes and also sites of plasmin cleavage [18][19][20]. Sauls et al [20] suggest that homocysteinylation of lysine residues in fibrinogen may be linked to three important functional consequences.…”

“…Sauls et al [18] have reported the same process, but in their experiments they used rabbit fibrinogen treated only with Hcys, and the authors did not write what form of Hcys was used. Clot structure analysed by scanning electron microscopy showed also no differences in either the general architecture or the fibrin fiber diameter at different Hcys concentrations (13,19, and 52 mM) [17]. However, mass spectrometric analysis of fibrinogen treated with Hcys revealed 12 lysines that were homocysteinylated.…”