Logo
DeutschClear Cookie - decide language by browser settings
Reinhardt, Christoph (2007): Analyses on molecular mechanisms of activation of intravascular Tissue Factor. Dissertation, LMU München: Faculty of Medicine
[img]
Preview
PDF
Reinhardt_Christoph.pdf

973Kb

Abstract

Both activated platelets and circulating microparticles were described to express tissue factor (TF), the principal initiator of coagulation, on their cell surface (intravascular TF). It is still not clear whether TF is functionally active on activated platelets. TF expressed on activated monocytes and various other cell types has been described to be functionally inactive (encrypted or latent TF). In the present study, cellular mechanisms are analyzed that could release the TF procoagulant activity of blood components. Tissue factor pathway inhibitor-1 (TFPI) represents the main physiologic inhibitor of the coagulation start. It inhibits the ternary initiator complex of the extrinsic coagulation pathway by first binding the circulating factors X / Xa and subsequently interacting with VII / VIIa. We found that after stimulation with thrombin and collagen type I, TFPI was recovered in the platelet releasate and it was degraded by neutrophil elastase (NE) released from activated neutrophils. TFPI degradation was also induced by NE on neutrophil microparticles. We found that NE is bound to negatively charged macromolecules (proteoglycans, RNA) on the surface of activated neutrophils by polar interactions. Overall, we could provide substantial experimental evidence that upon interaction of activated platelets with PMN a microenvironment is formed, which allows the efficient degradation of TFPI by the PMN-associated serine protease NE. This cross talk between the innate immune system and the coagulation system might be of general importance in pathologies, such as sepsis, arterial and venous thrombosis and myocardial infarction. TF-encryption has represented an unsolved problem for several decades. We reveal that thiol-disulfide exchange in the extracellular C186/C209 disulfide pair of TF triggers the TF procoagulant activity. Formation of the intramolecular C186/C209 disulfide activates TF procoagulant function, whereas reduction of the disulfide to the appropriate sulfhydryls and the formation of mixed disulfides (protein S-glutathionylation of TF) were found to suppress its procoagulant function. TF activation is supported by the thiol isomerase protein disulfide isomerase (PDI) and it is facilitated by the reactive oxygen species (ROS) detoxifying enzyme glutaredoxin (GRX). Protein S-glutathionylation of TF was uncovered as one reversible mechanism preventing the intravascular coagulation start. We suggest that thiol isomerases are injury-responsive signals driving coagulation through posttranslational cysteine modifications of TF. This mechanism could help to explain the augmented occurrence of vasoocclusive pathologies during ageing, where increasing concentrations of ROS might favour TF oxidation.