Tissue Factor (TF) Activity in Trauma Patients Is Suppressed on Cell-Derived Microparticles (MP).

BACKGROUND & AIMS. Traumatic injury is expected to expose or release into circulation large amounts of procoagulant material, including TF from subendothelial tissues, yet thrombosis is rarely an immediate concern. To investigate the basis of this, we measured TF activity and TF antigen (TF:Ag) associated with MP (believed to be a main carrier of circulating TF) in trauma patients.

METHODS. With IRB approval, two tubes of citrated blood were obtained upon admission from each of 18 patients and were processed within 2hr. Four patients were followed up 5–7 days later. MP were isolated from 0.9mL platelet-poor plasma (PPP) by centrifuging at 16,000g for 15min, then resuspending in 0.45mL Tris-saline pH 8.0 with brief (5sec) ultrasonic probe. Then 10uL was added to reaction mix (0.2mL final vol) in 96-well plate on ice containing physiologic amounts of FX, FVII, FVa, prothrombin, phospholipid and chromogenic substrate for thrombin, S-2238, then calcium to start reaction. Plate was then warmed (t=0) on metal block in 37° oven with fan and absorbance (OD) read at intervals from 6 min to 20min at 405nm. Known standards of pure thrombin and TF enabled conversion from OD to units of thrombin per ug TF/min. Flow cytometry was used to measure TF:Ag in PPP by fluorescent mAb.

RESULTS. (i) The apparent TF activity of patient MP was significantly or markedly reduced in 16 of 18 trauma patients, to as little as 10% of controls (mean of the 16 was 45.4%, median 47.7%). This was not seen in any other group of clinical patients. (ii) However, comparing rates of thrombin production by plotting log OD vs. time (giving straight line, R>0.97, as expected since thrombin accumulates in this method) revealed that final rates were often similar to controls. This was explained by prolonged lag phase in patients. Extrapolation back to OD=0.01 showed origin delayed by 2–8min in trauma patients relative to controls. (iii) Neither MP numbers or TF antigen differed significantly or consistently from controls; TF-positive MP in plasma were 529 ±196 vs. 581 ±279 per mL for patients and controls respectively.

CONCLUSIONS. Trauma patients do not exhibit elevated circulating TF activity on MP. The prolonged lag phase might be explained by presence of increased protective TFPI or cryptic state in patients, requiring more time for expression of full activity in assay medium, supported by other experiments showing that anti-TFPI restored normal or above-normal activity in trauma patients.