Myeloid cell tissue factor does not contribute to venous thrombogenesis in an electrolytic injury model

Abstract

Introduction: Tissue factor (TF) is a potent initiator of the extrinsic coagulation cascade. The role and source of TF in venous thrombotic disease is not clearly defined. Our study objective was to identify the contribution of myeloid cell TF to venous thrombogenesis in mice.

Materials and methods: The mouse electrolytic inferior vena cava model was used to induce thrombosis. The following groups of mice were used (1) TF(flox/flox)LysMCre(+) mice that have reduced TF expression in myeloid cells, (2) TF(flox/flox)LysMCre(-) littermate controls, (3) Wild type mice given a monoclonal anti-mouse TF antibody (1H1) to inhibit TF activity, and (4) Wild type mice given rat IgG. Evaluations at baseline, day 2, and day 6 post thrombosis included thrombus weight, vein wall inflammatory cell migration, vein wall TF mRNA, and plasma D-dimer levels.

Results: Inhibition of TF significantly decreased thrombus weight 2days post venous thrombosis. In contrast, TF(flox/flox)LysMCre(+) had no change in thrombus weight when compared to littermate controls. The absence of myeloid cell TF did not affect infiltration of neutrophils or monocytes into the vein wall. TF mRNA expression in the vein wall decreased at 2days but then returned to baseline levels by 6days post thrombosis. D-dimer levels peaked at 2days post thrombosis in mice with or without myeloid cell TF.

Conclusions: TF is important in the formation of venous thrombi in the macrovasculature. However, TF expression by myeloid cells does not significantly contribute to venous thrombogenesis in this model.

Figure 1A and Figure 1B. Thrombus Weight Composite

* indicate significant differences, vs. non-thrombosed true controls, **indicate significant differences, vs. 2 days post thrombosis (1A: n=10 for 2D thrombosis and n=5 for 6D thrombosis; 1B: n=6 for TC, n=10 for 2D and 6D thrombosis). TC=Non-thrombosed true controls, D=days post thrombosis. Systemic inhibition of tissue factor significantly decreased thrombus size at 2 days post thrombosis. LysMCre + mice showed no decrease in thrombus size at any time point, compared to LysMCre - littermate controls. Tissue factor derived from myeloid cells did not contribute to thrombus size.

Figure 2A and Figure 2B. Vein Wall Morphometrics Composite

average cell counts for each group (LysMCre +: n=5 for TC, n=10 for 2D and 6D thrombosis; LysMCre -: n=3 for all groups). TC=Non-thrombosed true controls, D=days post thrombosis, HPF=High power field. Both groups of mice (LysMCre + and LysMCre -) showed the same acute (neutrophil) to chronic (monocyte) inflammatory pattern. No differences were seen between the groups at any time-points.

Figure 3. Tissue Factor Quantitative (Real-time) PCR

*indicate significant differences, vs. non-thrombosed true controls and 6 days post thrombosis. (n=6 for TC, n=10 for 2D and 6D thrombosis). TC=Non-thrombosed true controls, D=days post thrombosis. There were no differences in vein wall TF mRNA levels between the LysMCre groups. LysMCre + mice showed no change in vein wall expression of tissue factor.

Figure 4. Plasma D-dimer

* indicate significant differences, vs. non-thrombosed true controls and 6 days post thrombosis. (LysMCre +: n=5 for TC, n=10 for 2D and 6D thrombosis; LysMCre -: n=3 for all groups). TC=Non-thrombosed true controls, D=days post thrombosis. LysMCre + and LysMCre - mice showed a peak in D-dimer levels at day 2 which returned to baseline levels by day 6 post thrombosis. The peak fibrinolysis occurred 2 days post venous thrombosis in both LysMCre groups.