Factor XIIIa-dependent retention of red blood cells in clots is mediated by fibrin α-chain crosslinking.
Factor XIII(a) [FXIII(a)] stabilizes clots and increases resistance to fibrinolysis and mechanical disruption. FXIIIa also mediates red blood cell (RBC) retention in contracting clots and determines venous thrombus size, suggesting FXIII(a) is a potential target for reducing thrombosis. However, the mechanism by which FXIIIa retains RBCs in clots is unknown. We determined the effect of FXIII(a) on human and murine clot weight and composition. Real-time microscopy revealed extensive RBC loss from clots formed in the absence of FXIIIa activity, and RBCs exhibited transient deformation as they exited the clots. Fibrin band-shift assays and flow cytometry did not reveal crosslinking of fibrin or FXIIIa substrates to RBCs, suggesting FXIIIa does not crosslink RBCs directly to the clot. RBCs were retained in clots from mice deficient in α2-antiplasmin, thrombin-activatable fibrinolysis inhibitor, or fibronectin, indicating RBC retention does not depend on these FXIIIa substrates. RBC retention in clots was positively correlated with fibrin network density; however, FXIIIa inhibition reduced RBC retention at all network densities. FXIIIa inhibition reduced RBC retention in clots formed with fibrinogen that lacks γ-chain crosslinking sites, but not in clots that lack α-chain crosslinking sites. Moreover, FXIIIa inhibitor concentrations that primarily block α-, but not γ-, chain crosslinking decreased RBC retention in clots. These data indicate FXIIIa-dependent retention of RBCs in clots is mediated by fibrin α-chain crosslinking. These findings expose a newly recognized, essential role for fibrin crosslinking during whole blood clot formation and consolidation and establish FXIIIa activity as a key determinant of thrombus composition and size.
Department of Pathology and Laboratory Medicine and McAllister Heart Institute, University of North Carolina at Chapel Hill, Chapel Hill, NC;
James R
JR
Byrnes
Division of Cardiovascular and Diabetes Research, Leeds Institute of Cardiovascular and Metabolic Medicine, University of Leeds, Leeds, United Kingdom;
Cédric
C
Duval
Canadian Blood Services, and Department of Laboratory Medicine and Pathobiology, St. Michael's Hospital, University of Toronto, Toronto, Canada;
Yiming
Y
Wang
School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, GA; Wallace C. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA; Aflac Cancer and Blood Disorders Center, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA;
Caroline E
CE
Hansen
School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, GA; Wallace C. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA; Aflac Cancer and Blood Disorders Center, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA;
Byungwook
B
Ahn
Department of Medicine and.
Micah J
MJ
Mooberry
Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, NC;
Martha A
MA
Clark
Research Institute, Bloodworks Northwest, Seattle, WA; Department of Medicine, University of Washington, Seattle, WA;
Jill M
JM
Johnsen
Department of Pathology and Laboratory Medicine and McAllister Heart Institute, University of North Carolina at Chapel Hill, Chapel Hill, NC;
Susan T
ST
Lord
School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, GA; Wallace C. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA; Aflac Cancer and Blood Disorders Center, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA;
Wilbur A
WA
Lam
Department of Experimental Vascular Medicine, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands; and Department of Plasma Proteins, Sanquin Research, Amsterdam, The Netherlands.
Joost C M
JC
Meijers
Canadian Blood Services, and Department of Laboratory Medicine and Pathobiology, St. Michael's Hospital, University of Toronto, Toronto, Canada;
Heyu
H
Ni
Division of Cardiovascular and Diabetes Research, Leeds Institute of Cardiovascular and Metabolic Medicine, University of Leeds, Leeds, United Kingdom;
Robert A S
RA
Ariëns
Department of Pathology and Laboratory Medicine and McAllister Heart Institute, University of North Carolina at Chapel Hill, Chapel Hill, NC;
Alisa S
http://orcid.org/0000-0002-2845-2303
AS
Wolberg
D000818:Animals; D001777:Blood Coagulation / Q000502:physiology*; D001779:Blood Coagulation Factors / Q000235:genetics / Q000378:metabolism*; D025901:Carboxypeptidase B2 / Q000235:genetics / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism*; D005353:Fibronectins / Q000235:genetics / Q000378:metabolism; D006474:Hemorrhagic Disorders / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D000979:alpha-2-Antiplasmin / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D005723:gamma-Glutamyltransferase / Q000235:genetics / Q000378:metabolism*
D001779:Blood Coagulation Factors; D005353:Fibronectins; D000979:alpha-2-Antiplasmin; D005723:gamma-Glutamyltransferase; D025901:Carboxypeptidase B2