Factor XIII-A Transglutaminase Contributes to Neutrophil Extracellular Trap (NET)-mediated Fibrin(ogen) Network Formation and Crosslinking

Abstract

Background

Neutrophil extracellular traps can contribute to thrombosis via stabilization of fibrin network, which is normally conducted by plasma transglutaminase, Factor XIII-A as part of coagulation cascade. The possible presence and activity of FXIII-A in neutrophils or during NETosis are unknown. Here, we investigated potential presence of FXIII-A in neutrophils and participation in NET-fibrin(ogen) interaction in vitro.

Methods

Data mining of human and mouse F13A1/F13a1 mRNA expression in whole-body scRNA sequence atlases was conducted. F13a1 mRNA and protein expression was assessed in isolated mouse bone marrow neutrophils. NETosis was induced using 12-phorbol 13-myristate acetate (PMA), and the transglutaminase activity was assessed with 5-(biotinamido)pentylamine incorporation to plasma fibronectin and a fluorescence-fibrin(ogen)-based activity assay using ATTO488-Cadaverine. Externalization of FXIII-A and its interaction with neutrophil extracellular trap (NET) markers, namely, decondensed DNA, CitH3, and MPO, were examined with immunofluorescence microscopy. NET–fibrin(ogen) interaction was investigated with and without serum and/or transglutaminase inhibitor, NC9. Effect of soluble fibrinogen and fibrin(ogen) network on NETosis was also assessed.

Results

Data mining of RNAseq atlases showed F13A1/F13a1 expression in adipose tissue, blood, and bone marrow neutrophils. mRNA expression and protein production were confirmed in isolated neutrophils where expression was comparable to that of macrophages and monocytes. FXIII-A was externalized and active as a transglutaminase and colocalized with NET markers during NETosis. FXIII-A transglutaminase activity promoted NET–fibrin(ogen) interaction and entrapment of neutrophils within fibrin(ogen) matrix. Soluble fibrinogen or fibrin(ogen) network did not induce NETosis.

Conclusion

This study identifies neutrophils as a source of FXIII-A and suggests its role in stabilizing NET–fibrin(ogen) matrix structures.

Keywords

Factor XIII-A - transglutaminase - neutrophil - fibrin(ogen) - fibrin - neutrophil extracellular traps

Authors' Contribution

M.T.K. designed the study and revised the manuscript. F.S. performed experiments, analyzed data, and wrote the manuscript. S.E. conducted the monocyte and osteoclast isolation. M.W. conducted the platelet-related experiments. M.L. and M.W. revised the intellectual content. All authors have read and approved the final version of the manuscript.

Publication History

Received: 26 June 2024

Accepted: 16 December 2024

Accepted Manuscript online:
18 December 2024

Article published online:
20 January 2025

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