Departments of Immunopathology, University of Amsterdam, Amsterdam, the Netherlands.
J Thromb Haemost. 2012 Jun;10(6):1165-71. doi: 10.1111/j.1538-7836.2012.04712.x.
Factor VII-activating protease (FSAP) is a serine protease that circulates in plasma in its inactive single-chain form and can be activated upon contact with dead cells. When activated by apoptotic cells, FSAP leads to the release of nucleosomes. The serpins C1-inhibitor and α(2) -antiplasmin are reported to be the major inhibitors of FSAP. However, regulation of FSAP activity by Kunitz-type inhibitors is not well studied.
To compare the inhibition of FSAP activity and FSAP-induced nucleosome release from apoptotic cells by tissue factor pathway inhibitor (TFPI) with that of C1-inhibitor and α(2) -antiplasmin.
Apoptotic cells were incubated with plasma or FSAP in presence of the inhibitor, and nucleosome release was analyzed with flow cytometry. Monoclonal antibodies against TFPI and altered forms of TFPI were used to investigate which domains of TFPI contribute to FSAP inhibition.
We show that TFPI abrogates FSAP activity and nucleosome release from apoptotic cells. TFPI is a much more efficient inhibitor than C1-inhibitor or α(2) -antiplasmin. The active site of K2 is required for inhibition of FSAP. A direct binding interaction between FSAP and the C-terminal domain of TFPI is also required for efficient inhibition. Inhibition of FSAP-induced nucleosome release by recombinant TFPI might, in part, explain the anti-inflammatory effects of recombinant TFPI infusion observed in animal and human sepsis.
VII 因子激活蛋白酶（FSAP）是一种丝氨酸蛋白酶，以无活性的单链形式在血浆中循环，在与死亡细胞接触时可被激活。当被凋亡细胞激活时，FSAP 导致核小体的释放。丝氨酸蛋白酶抑制剂 C1 抑制剂和 α2-抗纤溶酶被报道为 FSAP 的主要抑制剂。然而，Kunitz 型抑制剂对 FSAP 活性的调节尚未得到很好的研究。
比较组织因子途径抑制剂（TFPI）对 FSAP 活性的抑制作用以及对凋亡细胞中 FSAP 诱导的核小体释放的抑制作用与 C1 抑制剂和 α2-抗纤溶酶的抑制作用。
将凋亡细胞与血浆或 FSAP 在抑制剂存在的情况下孵育，并通过流式细胞术分析核小体释放。使用针对 TFPI 的单克隆抗体和 TFPI 的改变形式来研究 TFPI 的哪些结构域有助于 FSAP 抑制。
我们表明 TFPI 可阻断 FSAP 从凋亡细胞中发挥作用并释放核小体。TFPI 比 C1 抑制剂或 α2-抗纤溶酶更有效。K2 的活性部位是抑制 FSAP 所必需的。FSAP 和 TFPI 的 C 末端结构域之间的直接结合相互作用也是有效抑制所必需的。重组 TFPI 对 FSAP 诱导的核小体释放的抑制作用可能部分解释了在动物和人类脓毒症中观察到的重组 TFPI 输注的抗炎作用。
