title: Disintegration and reorganization of fibrin networks during tissue-type plasminogen activator-induced clot lysis.
abstract: In this study, we investigated tissue-type plasminogen activator (tPA)-induced lysis of glutamic acid (glu)-plasminogen-containing or lysine (lys)-plasminogen-containing thrombin-induced fibrin clots. We measured clot development and plasmin-mediated clot disintegration by thromboelastography, and used scanning electron microscopy (SEM) to document the structural changes taking place during clot formation and lysis. These events occurred in three overlapping stages, which were initiated by the addition of thrombin, resulting first in fibrin polymerization and clot network organization (Stage I). Autolytic plasmin cleavage of glu-plasminogen at lys-77 generates lys-plasminogen, exposing lysine binding sites in its kringle domains. The presence of lys-plasminogen within the thrombin-induced fibrin clot enhanced network reorganization to form thicker fibers as well as globular complexes containing fibrin and lys-plasminogen having a greater level of turbidity and a higher elastic modulus (G) than occurred with thrombin alone. Lys-plasminogen or glu-plasminogen that had been incorporated into the fibrin clot was activated to plasmin by tPA admixed with the thrombin, and led directly to clot disintegration (Stage II) concomitant with fibrin network reorganization. The onset of Stage III (clot dissolution) was signaled by a sustained secondary rise in turbidity that was due to the combined effects of lys-plasminogen presence or its conversion from glu-plasminogen, plus clot network reorganization. SEM images documented dynamic structural changes in the lysing fibrin network and showed that the secondary turbidity rise was due to extensive reorganization of severed fibrils and fibers to form wide, occasionally branched fibers. These degraded structures contributed little, if anything, to the structural integrity of the residual clot, and eventually collapsed completely during the course of progressive clot dissolution. These results provide new perspectives on the major structural events that occur in the fibrin clot matrix during fibrinolysis.
authors.affiliation: The Blood Research Institute of The Blood Center of Southeastern Wisconsin, Milwaukee, Wisconsin 53201-2178, USA.
authors.forename: D A
authors.initials: DA
authors.lastname: Meh
authors.forename: M W
authors.initials: MW
authors.lastname: Mosesson
authors.forename: J P
authors.initials: JP
authors.lastname: DiOrio
authors.forename: K R
authors.initials: KR
authors.lastname: Siebenlist
authors.forename: I
authors.initials: I
authors.lastname: Hernandez
authors.forename: D L
authors.initials: DL
authors.lastname: Amrani
authors.forename: L
authors.initials: L
authors.lastname: Stojanovich
mesh_terms: D005337:Fibrin / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D005342:Fibrinolysis / Q000187:drug effects*; D006801:Humans; D007700:Kinetics; D008855:Microscopy, Electron, Scanning; D009391:Nephelometry and Turbidimetry; D010446:Peptide Fragments / Q000494:pharmacology / Q000502:physiology; D010958:Plasminogen / Q000494:pharmacology / Q000502:physiology; D013916:Thrombelastography; D013917:Thrombin / Q000494:pharmacology; D010959:Tissue Plasminogen Activator / Q000494:pharmacology* / Q000502:physiology
chemical_list: D010446:Peptide Fragments; C018998:lysyl-plasminogen; D005337:Fibrin; D010958:Plasminogen; D013917:Thrombin; D010959:Tissue Plasminogen Activator