Lupu Cristina, Hu Xiaohong, Lupu Florea
Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, 825 NE 13th Street, Oklahoma City, OK 73104, USA.
J Biol Chem. 2005 Jun 10;280(23):22308-17. doi: 10.1074/jbc.M503333200. Epub 2005 Apr 6.
Tissue factor pathway inhibitor (TFPI) blocks tissue factor-factor VIIa (TF-FVIIa) activation of factors X and IX through the formation of the TF-FVIIa-FXa-TFPI complex. Most TFPI in vivo associates with caveolae in endothelial cells (EC). The mechanism of this association and the anticoagulant role of caveolar TFPI are not yet known. Here we show that expression of caveolin-1 (Cav-1) in 293 cells keeps TFPI exposed on the plasmalemma surface, decreases the membrane lateral mobility of TFPI, and increases the TFPI-dependent inhibition of TF-FVIIa. Caveolae-associated TFPI supports the co-localization of the quaternary complex with caveolae. To investigate the significance of these observations for EC we used RNA interference to deplete the cells of Cav-1. Functional assays and fluorescence microscopy revealed that the inhibitory properties of TFPI were diminished in EC lacking Cav-1, apparently through deficient assembly of the quaternary complex. These findings demonstrate that caveolae regulate the inhibition by cell-bound TFPI of the active protease production by the extrinsic pathway of coagulation.
组织因子途径抑制物（TFPI）通过形成组织因子 - 因子VIIa - 因子Xa - TFPI复合物来阻断组织因子 - 因子VIIa（TF - FVIIa）对因子X和因子IX的激活。体内的大多数TFPI与内皮细胞（EC）中的小窝相关联。这种关联的机制以及小窝TFPI的抗凝作用尚不清楚。在这里，我们表明293细胞中小窝蛋白 - 1（Cav - 1）的表达使TFPI暴露于质膜表面，降低了TFPI的膜侧向流动性，并增强了TFPI对TF - FVIIa的依赖性抑制作用。与小窝相关的TFPI支持四元复合物与小窝的共定位。为了研究这些观察结果对内皮细胞的意义，我们使用RNA干扰来耗尽细胞中的Cav - 1。功能测定和荧光显微镜检查显示，在缺乏Cav - 1的内皮细胞中，TFPI的抑制特性减弱，显然是由于四元复合物组装不足。这些发现表明，小窝调节细胞结合的TFPI对凝血外源性途径产生活性蛋白酶的抑制作用。
