Identification and characterization of murine alternatively spliced tissue factor.

Tissue factor (TF) is a transmembrane glycoprotein that initiates coagulation and plays a critical role in regulating hemostasis and thrombosis. We have recently reported a naturally occurring, soluble form of human tissue factor (asTF) generated by alternative splicing. This splice variant has a novel C-terminus with no homology to that of the full-length TF (flTF), lacks a transmembrane domain, and is active in the presence of phospholipids. Mouse models offer unique opportunities to examine the relative importance of flTF and asTF in mediating thrombosis, the response to arterial injury, and ischemic damage. To that end, we have identified and characterized murine asTF (masTF). Like the human splice variant, masTF lacks a transmembrane domain and has a unique C-terminus. We have generated antibodies specific to masTF and murine flTF (mflTF) to examine the expression of both forms of TF. masTF antigen is widely and abundantly expressed, with a pattern similar to that of mflTF, in adult tissues, in experimentally induced thrombi, and during development. These studies demonstrate that masTF contributes to the pool of total TF and may thus play an important role in mediating TF-dependent processes.

Active tissue factor shed from human arterial smooth muscle cells adheres to artificial surfaces.

Through a series of in vitro assays, this study outlines a flow-mediated process by which active tissue factor (TF), the prime initiator of coagulation, may be transferred from the plasma membrane of vascular smooth muscle cells (VSMCs) to that of artificial surfaces such as those typically associated with intravascular implants. Studies with quiescent and activated rat VSMCs demonstrated that pathologically high shear stresses (tau(w) = 250 dyn cm(-2)) resulted in the loss of TF activity from the cell surface. Subsequent experiments with human VSMCs showed that VSMCs continuously release active TF into their extracellular medium, presumably in the form of lipid vesicles or microparticles, and that fluid shear stress (tauw = 50 dyncm(-2)) or chemical agonists (A23187) can significantly accelerate this release. Experiments with a wide array of polymeric and metallic materials showed that the TF shed from VSMCs was able to adhere to these surfaces and promote the activation of coagulation factor X (FX) at the material surface. Extracellular TF bound strongly to both uncoated and human plasma coated surfaces under a wide range of hemodynamic shear stresses (0-20 dyncm(-2)). When an extracellular, VSMC-derived TF mixture was perfused over Ti 6-4 surfaces, the adhesion of TF was found to be time-dependent, gradually accumulating on the material surface over time. Thus an important criterion in the design or success of intravascular devices may be related to their ability to interact with TF, shed from cell surfaces. This is especially important as TF may lead to thrombotic complications, the products of which may also increase cellular proliferation.