Mandal Samir K, Iakhiaev Alexei, Pendurthi Usha R, Rao L Vijaya Mohan
Biomedical Research Division, The University of Texas Health Center at Tyler, Tyler, TX 75708, USA.
Blood. 2005 Jan 1;105(1):153-60. doi: 10.1182/blood-2004-03-0990. Epub 2004 Aug 24.
Cholesterol, in addition to providing rigidity to the fluid membrane, plays a critical role in receptor function, endocytosis, recycling, and signal transduction. In the present study, we examined the effect of membrane cholesterol on functional expression of tissue factor (TF), a cellular receptor for clotting factor VIIa. Depletion of cholesterol in human fibroblasts (WI-38) with methyl-beta-cyclodextrin-reduced TF activity at the cell surface. Binding studies with radiolabeled VIIa and TF monoclonal antibody (mAB) revealed that reduced TF activity in cholesterol-depleted cells stems from the impairment of VIIa interaction with TF rather than the loss of TF receptors at the cell surface. Repletion of cholesterol-depleted cells with cholesterol restored TF function. Loss of caveolar structure on cholesterol removal is not responsible for reduced TF activity. Solubilization of cellular TF in different detergents indicated that a substantial portion of TF in fibroblasts is associated with noncaveolar lipid rafts. Cholesterol depletion studies showed that the TF association with these rafts is cholesterol dependent. Overall, the data presented herein suggest that membrane cholesterol functions as a positive regulator of TF function by maintaining TF receptors, probably in noncaveolar lipid rafts, in a high-affinity state for VIIa binding.
胆固醇除了为流体膜提供刚性外，在受体功能、内吞作用、再循环和信号转导中也起着关键作用。在本研究中，我们检测了膜胆固醇对组织因子（TF）功能表达的影响，TF是凝血因子VIIa的细胞受体。用甲基-β-环糊精处理人成纤维细胞（WI-38）使胆固醇耗竭，导致细胞表面TF活性降低。用放射性标记的VIIa和TF单克隆抗体（mAB）进行的结合研究表明，胆固醇耗竭细胞中TF活性降低源于VIIa与TF相互作用受损，而非细胞表面TF受体丢失。用胆固醇补充胆固醇耗竭的细胞可恢复TF功能。去除胆固醇时小窝结构的丧失并非TF活性降低的原因。在不同去污剂中对细胞TF进行溶解表明，成纤维细胞中相当一部分TF与非小窝脂筏相关。胆固醇耗竭研究表明，TF与这些脂筏的结合是依赖胆固醇的。总体而言，本文提供的数据表明，膜胆固醇通过将TF受体（可能存在于非小窝脂筏中）维持在对VIIa结合具有高亲和力的状态，从而作为TF功能的正向调节因子发挥作用。
