Laboratory of Clinical Chemistry, and Vesicle Observation Centre, Academic Medical Centre of the University of Amsterdam, Amsterdam, The Netherlands.
Department of Immunology and Microbial Science, Scripps Research Institute, La Jolla, California, United States.
Thromb Haemost. 2018 Jun;118(6):990-1000. doi: 10.1055/s-0038-1642031. Epub 2018 Apr 21.
Tissue factor (TF) can be present in a non-coagulant and coagulant form. Whether the coagulant activity is affected by the plasma membrane microenvironment is unexplored.
This article studies the presence and coagulant activity of human TF in plasma membrane micro-domains.
Plasma membranes were isolated from human MIA PaCa2 cells, MDA-MB-231 cells and human vascular smooth muscle cells by Percoll gradient ultracentrifugation after cell disruption. Plasma membranes were fractionated by OptiPrep gradient ultracentrifugation, and the presence of TF, flotillin, caveolin, clathrin, protein disulphide isomerase (PDI), TF pathway inhibitor (TFPI) and phosphatidylserine (PS) were determined.
Plasma membranes contain two detergent-resistant membrane (DRM) compartments differing in density and biochemical composition. High-density DRMs (DRM-H) have a density () of 1.15 to 1.20 g/mL and contain clathrin, whereas low-density DRMs (DRM-L) have a density between 1.09 and 1.13 g/mL and do not contain clathrin. Both DRMs contain TF, flotillin and caveolin. PDI is detectable in DRM-H, TFPI is not detectable in either DMR-H or DRM-L and PS is detectable in DRM-L. The DRM-H-associated TF (> 95% of the TF antigen) lacks detectable coagulant activity, whereas the DRM-L-associated TF triggers coagulation. This coagulant activity is inhibited by lactadherin and thus PS-dependent, but seemed insensitive to 16F16, an inhibitor of PDI.
Non-coagulant and coagulant TF are present within different types of DRMs in the plasma membrane, and the composition of these DRMs may affect the TF coagulant activity.
组织因子 (TF) 可以以非凝血和凝血两种形式存在。凝血活性是否受质膜微环境的影响尚未可知。
本文研究了人 TF 在质膜微区中的存在和凝血活性。
用 Percoll 梯度超速离心法破坏细胞后，从人 MIA PaCa2 细胞、MDA-MB-231 细胞和人血管平滑肌细胞中分离质膜。用 OptiPrep 梯度超速离心法对质膜进行分级，检测 TF、绒毛蛋白、小窝蛋白、网格蛋白、蛋白二硫键异构酶 (PDI)、组织因子途径抑制剂 (TFPI) 和磷脂酰丝氨酸 (PS) 的存在。
质膜包含两个去污剂抗性膜 (DRM) 区室，它们在密度和生化组成上存在差异。高密度 DRM (DRM-H) 的密度为 1.15 至 1.20 g/mL，含有网格蛋白，而低密度 DRM (DRM-L) 的密度在 1.09 至 1.13 g/mL 之间，不含有网格蛋白。两种 DRM 均含有 TF、绒毛蛋白和小窝蛋白。PDI 可在 DRM-H 中检测到，TFPI 既不在 DRM-H 中也不在 DRM-L 中检测到，PS 可在 DRM-L 中检测到。DRM-H 相关的 TF(>95%的 TF 抗原)缺乏可检测的凝血活性，而 DRM-L 相关的 TF 触发凝血。这种凝血活性被乳凝集素抑制，因此依赖于 PS，但似乎对 PDI 的抑制剂 16F16 不敏感。
非凝血和凝血 TF 存在于质膜的不同类型的 DRM 中，这些 DRM 的组成可能影响 TF 的凝血活性。
