Intravascular Tissue Factor (TF) Is Predominantly Platelet-Associated during the Aplastic Phase of Hematopoietic Stem Cell Transplantation (HSCT).
Authors: Markova Marketa , Slungaard Arne , Aras Omer , Hysjulien Jessica L. , Long Janel , Shet Arun , Bach Ronald R. , Key Nigel S. 
Published: October 2019

Abstract
TF is a 47 kDa cell-bound glycoprotein that initiates coagulation upon contact with factor VII(a). Both monocytes and cells derived from the vessel wall (such as endothelial cells) are capable of TF synthesis, and both intact cells and microparticles (MPs) derived from them are considered to be the principal source(s) of intravascular TF. Data from in vitro systems and animal models suggest that MP-borne TF may be transferred to activated platelets via a P-selectin dependent pathway. We previously demonstrated that during the aplastic phase of HSCT, about 55% of baseline (pre-transplant) levels of circulating whole blood TF procoagulant activity (PCA) remained detectable (Thromb & Haemost2001;85:250). This study was therefore undertaken to identify whether this residual TF activity is accounted for by intact cells or MPs, and to determine whether cells derived from the vessel wall or the hematopoietic compartment are responsible. MP-associated TF PCA was measured using a recently described MP capture assay (Blood2004;103:4545). MP-associated TF PCA declined by 89% at day 0 and by 77% at day +7 compared to baseline levels measured at day −7, with a subsequent normalization of activity paralleling engraftment. TF PCA was assayed on fractionated platelets and mononuclear cells obtained on days −7,0,7,14 and 21 from 11 patients undergoing HSCT. Mononuclear cell and platelet fractions were obtained by differential centrifugation and exposed to calcium ionophore to maximally ‘de-encrypt’ TF PCA. Following pre-incubation with FVIIa, the clotting time of added plasma was recorded, and the amount of TF calculated by reference to a standard curve generated using Innovin™. In parallel with the drop in circulating monocyte counts, a significant decline in TF PCA in both the mononuclear cell (1855±690 at day −7 vs. 74±37 pg/ml at day +7, p=0.003) and platelet (260±45 at day −7 vs. 51±16 pg/ml at day +7, p=0.05) fractions was noted. TF PCA in these fractions subsequently returned towards baseline levels with engraftment. Notably, while platelet-associated TF PCA accounted for only 12% of the total cell-associated TF PCA at baseline (day −7), it accounted for fully 59% at day 0 and 41% at day +7. This led us to postulate that transfused platelets might contribute to circulating TF during the aplastic phase. Within 1 hour of receiving leuco-depleted platelet transfusion, total cell-associated (platelet + mononuclear cell) TF PCA had increased significantly (142 ± 43 pg/ml to 317 ± 55 pg/ml, p=0.015), confirming that transfused platelets are indeed a source of TF. While this augmentation of cell-associated TF PCA was primarily seen in the platelet fraction, an increase in mononuclear cell-associated TF PCA was also noted. Conclusion: The majority of cell-associated TF PCA is associated with the mononuclear cell fraction in whole blood. However, with the onset of severe leucopenia during HSCT, the platelet pool becomes the dominant location for TF. Our data are in agreement with recent transgenic mouse model data demonstrating that the bone marrow - not the vessel wall - is the dominant source of both cell- and MP-associated intravascular TF.