Abstract

Background: Amniotic fluid (AF) embolism is one of the most catastrophic complications during pregnancy. While studies primarily attributed the procoagulability of AF to circulating tissue factor (TF), the exact mechanism remains unclear. Aims: Define the procoagulant mechanisms of AF. Methods: Clotting time assays were performed using normal human plasma or plasma depleted of factor (F)XII, XI, X, IX, VII, V, or prekallikrein (PK). Chromogenic substrate assays were used to quantify FX activation by the TF/FVIIa complex and prothrombin activation by FXa. Phosphatidylserine (PS) levels were quantified using a fluorometric kit. Results: AF shortened the clotting time of normal plasma from 572±83 to 160±2 seconds; AF-induced clotting times were equivalent for FXII, PK, FXI, FX, FIX, or FV-depleted plasma or in the presence of contact pathway inhibitors as compared to normal plasma. In contrast, the procoagulant effects of AF were diminished or absent for FVII-depleted plasma or in the presence of a blocking anti-TF antibody or low-molecular-weight heparin. In a purified system, AF reduced FX activation by the TF/FVIIa complex. The addition of a blocking anti-tissue factor pathway inhibitor (TFPI) antibody reversed the effect of AF on FX activation. Western blotting confirmed the existence of both TF and TFPI in AF. AF enhanced prothrombin activation by FXa. The addition of annexin V, or lactadherin, which bind to PS, or the use of Gla-domainless prothrombin mitigated the effect of AF on prothrombin activation. PS levels in AF and normal plasma were 811±376 and 10.4±3.8 μM, respectively. Conclusions: The procoagulability of AF is independent of the contact pathway while being sensitive to both TF and TFPI levels. Furthermore, PS in AF significantly contributes to its procoagulability. This knowledge is important when developing strategies to lessen the procoagulability of AF without compromising hemostasis in the setting of AF embolism-associated coagulopathy.