Florida Hospital Center for Thrombosis Research, Orlando, FL, USA.
J Thromb Haemost. 2014 Feb;12(2):186-96. doi: 10.1111/jth.12475.
Tumor-derived tissue factor (TF) activates coagulation in vitro and in vivo in an orthotopic model of human pancreatic cancer. Here, we further characterized tumor-derived TF in this model.
Conditioned medium (CM) of L3.6pl human pancreatic tumor cells and plasma from nude mice bearing L3.6pl tumors were ultracentrifuged, and the pellets were filtered through membranes with different pore sizes. The size distribution of particles was analyzed in CM or plasma fractions with nanoparticle tracking and dynamic light scattering. Human TF antigen and activity were measured in pellets and supernatants with ELISA and clotting or thrombin generation assays, respectively. Human alternatively spliced TF (asTF) was measured with ELISA. Human TF and thrombin-antithrombin complex (TAT) concentrations were assessed in plasma of mice injected with filtered fractions of CM.
Particles in both CM and plasma were < 0.4 μm. TF antigen and activity in the CM were mainly associated with microparticles (MP). Approximately 50% of antigen and 20% of activity were associated with particles of < 0.1 μm. Injection of < 0.1 μm particles into mice caused a 30% drop in platelet counts and an increase in TAT levels. In contrast, ~ 90% of TF antigen in tumor-bearing mice plasmas was non-sedimentable, whereas TF activity was exclusively associated with MP. Particles of < 0.1 μm and the supernatants of both CM and plasma gained TF activity after addition of exogenous phospholipids. Although asTF was found in MP-free CM supernatants, it was also present in CM and plasma pellets.
Tumor-derived particles of < 0.1 μm and non-sedimentable TF are or can become procoagulant in the presence of phospholipids, and may contribute to the procoagulant potential of circulating TF.
肿瘤来源的组织因子 (TF) 在人胰腺癌细胞的原位模型中体外和体内激活凝血。在这里，我们进一步研究了该模型中的肿瘤来源的 TF。
L3.6pl 人胰腺肿瘤细胞的条件培养基 (CM) 和携带 L3.6pl 肿瘤的裸鼠血浆经超速离心，沉淀用不同孔径的膜过滤。纳米颗粒跟踪和动态光散射分析 CM 或血浆部分中的颗粒大小分布。ELISA 和凝血或凝血酶生成测定分别测定沉淀和上清液中的人 TF 抗原和活性。ELISA 测定人替代剪接 TF (asTF)。用过滤的 CM 滤过部分注射小鼠的血浆中评估人 TF 和血栓素-抗血栓素复合物 (TAT) 浓度。
CM 和血浆中的颗粒均<0.4μm。CM 中的 TF 抗原和活性主要与微颗粒 (MP) 相关。约 50%的抗原和 20%的活性与<0.1μm 的颗粒相关。将<0.1μm 的颗粒注射入小鼠会导致血小板计数下降 30%，TAT 水平升高。相比之下，约 90%的肿瘤荷瘤小鼠血浆中的 TF 抗原是非沉淀的，而 TF 活性仅与 MP 相关。<0.1μm 的颗粒和 CM 及血浆的上清液在添加外源性磷脂后获得 TF 活性。虽然在无 MP 的 CM 上清液中发现了 asTF，但它也存在于 CM 和血浆沉淀中。
<0.1μm 的肿瘤来源颗粒和不可沉淀的 TF 在存在磷脂的情况下具有促凝活性，可能有助于循环 TF 的促凝潜能。
