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Factor XIII-A: An Indispensable “Factor” in Haemostasis and Wound Healing
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Factor XIII (FXIII) is a transglutaminase enzyme that catalyses the formation of ε-(γ-glutamyl)lysyl isopeptide bonds into protein substrates. The plasma form, FXIIIA2B2, has an established function in haemostasis, with fibrin being its principal substrate. A deficiency in FXIII manifests as a severe bleeding diathesis emphasising its crucial role in this pathway. The FXIII-A gene (F13A1) is expressed in cells of bone marrow and mesenchymal lineage. The cellular form, a homodimer of the A subunits denoted FXIII-A, was perceived to remain intracellular, due to the lack of a classical signal peptide for its release. It is now apparent that FXIII-A can be externalised from cells, by an as yet unknown mechanism. Thus, three pools of FXIII-A exist within the circulation: plasma where it circulates in complex with the inhibitory FXIII-B subunits, and the cellular form encased within platelets and monocytes/macrophages. The abundance of this transglutaminase in different forms and locations in the vasculature reflect the complex and crucial roles of this enzyme in physiological processes. Herein, we examine the significance of these pools of FXIII-A in different settings and the evidence to date to support their function in haemostasis and wound healing.
British Society of Haemostasis and Thrombosis (BSHT) virtual scientific conference 2021
Monocytes expose factor XIII-A and stabilize thrombi against fibrinolytic degradation
Background: Factor XIII (FXIII) is a transglutaminase (TG) that promotes thrombus stability by cross-linking fibrin. FXIII exists as a cellular form, a homodimer of the A subunits (FXIII-A) and it is found in abundance within circulating platelets and monocytes.
Aim: To investigate externalization of FXIII-A on monocytes and its role in extracellular crosslinking reactions.
Methods: Isolated human monocytes or THP-1 cells (6 x 105cells/ml) were treated ± 20 ng/ml interleukin-4 (IL-4); 20 ng/ml interleukin-10 (IL-10); or 100 ng/ml lipopolysaccharide (LPS) for 24 h. FXIII-A exposure and activity were detected using a FITC-labelled anti-FXIII-A antibody (40 μg/ml) or the fluorescent amine donor substrate, TAMRA (40 μg/ml) by flow cytometer and imaged by confocal microscopy. Pooled normal plasma or factor XIII deficient plasma + FITC-labelled fibrinogen (45 µg/ml) was re-calcified with CaCl2 (10.9 mM) ± stimulated monocytes or THP-1 cells (3 x 105 cells/ml) ± 1 mM a TG inhibitor. Thrombi were bathed in tissue plasminogen activator (1 µg/ml) and sampled every 30 min for 4 h and fluorescence read (Ex 485 nm Em 528 nm).
Results: IL-10 stimulation significantly increased the number of FXIII-A positive monocytes compared to unstimulated cells (21.2 ± 3.4% vs 7.9 ± 2.1% P < 0.05). FXIII-A activity was augmented 2.5-fold with IL-4 and IL-10 stimulation compared to resting cells. Basal levels of FXIII-A antigen expression were higher in THP-1 cells with a negligible impact of stimulation. However, FXIII-A activity on the surface of THP-1 cells was significantly increased with all stimuli compared to resting cells. FXIII-A antigen and activity were expressed on the external membrane of stimulated monocytes and THP-1 cells. However, the distribution with LPS stimulation produced a distinct punctate pattern around the cell periphery. IL-4 or IL-10 activated monocytes and THP-1 cells stabilized FXIII-depleted thrombi against fibrinolytic degradation, via a transglutaminase-dependent mechanism.
Summary: Activated monocytes and THP-1 cells externalize and retain active FXIII-A on their membrane thereby stabilizing thrombi against fibrinolytic degradation.