PB1398 - RBC-Derived Transglutaminase-2 Affects Thrombus Formation

Tuesday, June 27, 2023

18:30 – 19:30 ET

Room: Exhibition

Presenting Author(s)

Naoual Ouazzani Chahdi, MSc

PhD student
Department of Molecular Hematology, Sanquin Research, Amsterdam, The Netherlands
Den Haag, Zuid-Holland, Netherlands

Co-Author(s)

Judith J. de Vries, MSc

PhD student
Department of Hematology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, Netherlands
Rotterdam, Zuid-Holland, Netherlands
Hande Eyisoylu, MSc

Phd Student
Department of Hematology, Erasmus MC, Erasmus University Medical Center Rotterdam, Rotterdam, The Netherlands
Rotterdam, Zuid-Holland, Netherlands
HM

Hanke Matlung

Principal Investigator
Department of Molecular Hematology, Sanquin Research, Amsterdam, The Netherlands
Amsterdam, Noord-Holland, Netherlands
CH

Chi Hau

Support Staff
Department of Experimental Clinical Chemistry, Amsterdam UMC, The Netherlands
Amsterdam, Noord-Holland, Netherlands
RN

Rienk Nieuwland

Research Associate
University of Amsterdam
Amsterdam, Noord-Holland, Netherlands
GK

Gijsje H. Koenderink

Professor
Department of Bionanoscience, Kavli Institute of Nanoscience, Delft University of Technology, Delft, the Netherlands
Delft, Zuid-Holland, Netherlands
Moniek P.M. de Maat, PhD

Professor
Department of Hematology, Erasmus University Medical Center Rotterdam, Rotterdam, The Netherlands
Rotterdam, Zuid-Holland, Netherlands
JM

Joost CM Meijers

Biochemist
Department of Molecular Hematology, Sanquin Research, Amsterdam, The Netherlands
Amsterdam, Noord-Holland, Netherlands
RV

Robin Van Bruggen

PI
Department of Molecular Hematology, Sanquin Research, Amsterdam, The Netherlands
Amsterdam, Noord-Holland, Netherlands

Background: A venous clot mainly consists of red blood cells (RBCs), which contribute largely to the size and stability of a venous clot. However, it remains unclear if and how RBCs affect clotting. FXIIIa, an enzyme that crosslinks fibrin, strongly affects the mass and appearance of a clot. Additionally, transglutaminase-2 (TG2), a protein that has a function highly homologous to FXIIIa and is also expressed by RBCs, may play a role in clot formation.

Aims: To investigate whether TG2, expressed by RBCs, affects clot formation.

Methods: Clots were formed by incubating human recalcified citrated blood with tissue factor in the absence or presence of TG2 inhibitors. Moreover, TG2 knock-out (KO) RBCs were generated using CRISPR/Cas9 in hematopoietic stem cells, and used to form clots. Contracting clots were visualized by scanning electron microscopy (SEM) as well as fluorescence microscopy and fibrin formation was studied using time-lapse imaging. Lastly, microparticle flow cytometry was used to investigate RBC-derived vesicles.

Results: Inhibition of TG2 activity resulted in clots that had fewer and thinner fibrin fibers at the exterior of the clot. When using TG2 KO RBCs a less dense fibrin network was formed in the clots, which supported the findings using the TG2 inhibitor. Moreover, our data revealed that most of the fibrin was localized in the inside of the clot, when TG2 was inhibited or absent. Interestingly, when both the TG2 inhibitor and phospholipids, containing phosphatidylserine (PS), were added, the fibrin network seemed to be unaffected.

Conclusion(s): TG2 expressed by RBCs has an effect on the fibrin structure during clot formation.