CC BY-NC-ND 4.0 · Thromb Haemost 2019; 19(10): 1624-1631
DOI: 10.1055/s-0039-1693701
Coagulation and Fibrinolysis
Georg Thieme Verlag KG Stuttgart · New York

Decoration of Fibrin with Extracellular Chaperones

Simone Talens
1  Department of Hematology, Erasmus MC, University Medical Center Rotterdam, Erasmus University Rotterdam, Rotterdam, The Netherlands
,
Frank W. G. Leebeek
1  Department of Hematology, Erasmus MC, University Medical Center Rotterdam, Erasmus University Rotterdam, Rotterdam, The Netherlands
,
Robert Veerhuis
2  Clinical Chemistry Department, Amsterdam Neuroscience, Amsterdam UMC, Amsterdam, The Netherlands
3  Psychiatry Department, Amsterdam Neuroscience, Amsterdam UMC, Amsterdam, The Netherlands
,
Dingeman C. Rijken
1  Department of Hematology, Erasmus MC, University Medical Center Rotterdam, Erasmus University Rotterdam, Rotterdam, The Netherlands
› Author Affiliations
Further Information

Publication History

26 March 2019

12 June 2019

Publication Date:
22 August 2019 (eFirst)

  

Abstract

Background Many proteins bind to fibrin during clot formation in plasma. We previously identified by mass spectrometry the most abundant proteins that noncovalently bind to fibrin clots. Several of these proteins (e.g., apolipoprotein J/clusterin, haptoglobin, α2-macroglobulin, α1-antitrypsin) can act as extracellular chaperones.

Objective We hypothesize that clot-binding proteins may interact with fibrin as chaperones. The goal of this study is to test this hypothesis and to investigate the origin of the cross-β or amyloid structures in fibrin clots, which are associated with protein unfolding.

Methods and Results A thioflavin T assay was used to detect cross-β structures. A steadily increasing amount was measured in the fibrinogen fraction of plasma during heat stress, a standard treatment to induce unfolding of proteins. Heat-stressed plasma was clotted and clot-bound proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The results showed that the amounts of the clot-bound proteins were related to the duration of the heat stress. This indicates that cross-β structures in unfolded fibrin(ogen) are involved in clot binding of the proteins, which supports our chaperone hypothesis. A contributing role of fibrin formation itself was studied by clotting purified fibrinogen with thrombin in the presence of thioflavin T. The fluorescence intensity increased in time in the presence of thrombin, but did not increase in its absence. This provides evidence for the generation of cross-β structures during fibrin formation.

Conclusion Fibrin clots generated in plasma are decorated with extracellular chaperones. The binding of these chaperones involves cross-β structures originating both from unfolded fibrinogen and from fibrin formation.

Authors' Contributions

S.T. designed the research, performed the laboratory experiments, analyzed and interpreted data, and wrote the manuscript; F.W.G.L. designed the research and interpreted data; R.V. provided critical support about amyloid structures and for the experiments with thioflavin T; D.C.R. designed the research, analyzed and interpreted data, and wrote the manuscript. All authors critically reviewed the manuscript and gave their consent.