Coagulation factor XI induces Ca2+ response and accelerates cell migration in vascular smooth muscle cells via proteinase-activated receptor 1
Coagulation factor XI induces Ca2+ response and accelerates cell migration in vascular smooth muscle cells via proteinase-activated receptor 1
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DOI：
10.1152/ajpcell.00426.2018
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发表时间：
2019-03-01
影响因子：
5.5
通讯作者：
Hirano, Katsuya
中科院分区：
文献类型：
--
作者：
Liu, Wenhua;Hashimoto, Takeshi;Hirano, Katsuya
活化凝血因子XI （Activated coagulation factor XI, FXIa）是一种丝氨酸蛋白酶，在内在凝血途径中起关键作用。FXI基因敲除小鼠的分析表明FXI在动脉粥样硬化发病机制中的作用。然而，其潜在机制尚不清楚。我们假设FXIa通过蛋白酶激活受体1 （PAR1）发挥血管平滑肌作用。Fura-2荧光测定显示FXIa诱导大鼠胚胎主动脉平滑肌A7r5细胞内Ca2+信号。细胞外Ca2+的内流比细胞内Ca2+的释放在产生Ca2+信号中发挥更大的作用。fxia诱导的Ca2+信号被PAR1拮抗剂阿托帕沙（atopaxar）或蛋白酶抑制剂4-氨基苯基甲基磺酰氟（p-APMSF）预处理消除，同时在PAR(1)（-/-）小鼠衍生的胚胎成纤维细胞中也丢失。FXIa在含有PAR1细胞外区与凝血酶（一种典型的PAR1激动剂）相同的位置（R45/S46）切割重组蛋白。fxia诱导的Ca2+内流可被l型Ca2+通道阻滞剂地尔硫卓（diltiazem）和靶向CaV1.2的siRNA抑制。FXIa诱导的Ca2+内流也被蛋白激酶c的抑制剂GF109203X和rottlerin抑制。在伤口愈合实验中，FXIa使细胞迁移率增加了2.46倍，部分被阿托帕沙或地尔硫平抑制。综上所述，FXIa主要通过PAR(1)/Ca(V)1.2介导的Ca2+内流引发Ca2+信号，加速血管平滑肌细胞的迁移。本研究首次提供了FXIa对血管平滑肌产生直接细胞效应的证据。
Activated coagulation factor XI (FXIa) is a serine proteinase that plays a key role in the intrinsic coagulation pathway. The analysis of FXI-knockout mice has indicated the contribution of FXI to the pathogenesis of atherosclerosis. However, the underlying mechanism remains unknown. We hypothesized that FXIa exerts vascular smooth muscle effects via proteinase-activated receptor 1 (PAR1). Fura-2 fluorometry revealed that FXIa elicited intracellular Ca2+ signal in rat embryo aorta smooth muscle A7r5 cells. The influx of extracellular Ca2+ played a greater role in generating Ca2+ signal than the Ca2+ release from intracellular stores. The FXIa-induced Ca2+ signal was abolished by the pretreatment with atopaxar, an antagonist of PAR1, or 4-amidinophenylmethanesulfonyl fluoride (p-APMSF), an inhibitor of proteinase, while it was also lost in embryonic fibroblasts derived from PAR(1)(-/-) mice. FXIa cleaved the recombinant protein containing the extracellular region of PAR1 at the same site (R45/S46) as that of thrombin, a canonical PAR1 agonist. The FXIa-induced Ca2+ influx was inhibited by diltiazem, an L-type Ca2+ channel blocker, and by siRNA targeted to CaV1.2. The FXIa-induced Ca2+ influx was also inhibited by GF109203X and rottlerin, inhibitors of protein kinase C. In a wound healing assay, FXIa increased the rate of cell migration by 2.46-fold of control, which was partly inhibited by atopaxar or diltiazem. In conclusion, FXIa mainly elicits the Ca2+ signal via the PAR(1)/Ca(V)1.2-mediated Ca2+ influx and accelerates the migration in vascular smooth muscle cells. The present study provides the first evidence that FXIa exerts a direct cellular effect on vascular smooth muscle.