Endothelial protein C receptor-assisted transport of activated protein C across the mouse blood-brain barrier.
Endothelial protein C receptor-assisted transport of activated protein C across the mouse blood-brain barrier.
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DOI：
10.1038/jcbfm.2008.117
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发表时间：
2009-01
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活化蛋白C（APC）是一种丝氨酸蛋白酶，具有抗凝、抗炎和细胞保护活性，具有神经保护作用，并具有治疗不同神经系统疾病的潜力。目前尚不清楚APC是否能穿过血脑屏障（BBB）到达脑中的治疗靶点。通过使用脑血管灌注技术，我们表明，125 I标记的血浆来源的小鼠APC进入脑血管循环的浓度依赖性机制。125 I-APC（0.1 nM）在不同前脑区域的渗透性表面积积范围为3.11 - 4.13μL/min-g脑。这比14 C-菊粉（一种同时输注的参比示踪剂）高约80-110倍。APC BBB皮质转运的Km值为1.6 ± 0.2 nM。具有降低的抗凝活性的重组APC变体，5A-APC和3 K3 A-APC，而不是蛋白C，对APC BBB转运系统表现出高亲和力。阻断内皮蛋白C受体（EPCR）上的APC结合位点，但不阻断其蛋白酶激活受体-1（PAR 1）催化位点，可被> 85%的APC进入脑内所抑制。APC脑摄取在EPCR缺陷小鼠中减少了64%，但在PAR 1缺失小鼠中没有减少。这些数据表明，APC和其变体与降低抗凝活性通过EPCR介导的饱和运输跨越血脑屏障。
Activated protein C (APC), a serine-protease with anticoagulant, anti-inflammatory, and cytoprotective activities, is neuroprotective and holds potential to treat different neurological disorders. It is unknown whether APC crosses the blood-brain barrier (BBB) to reach its therapeutic targets in the brain. By using a brain vascular perfusion technique, we show that 125I-labeled plasma-derived mouse APC enters the brain from cerebrovascular circulation by a concentration-dependent mechanism. The permeability surface area product of 125I-APC (0.1 nM) in different forebrain regions ranged from 3.11 to 4.13μL/min-g brain. This was approximately 80–110-fold greater than for 14C-inulin, a simultaneously infused reference-tracer. The Km value for APC BBB cortical transport was 1.6 ± 0.2 nM. Recombinant APC variants with reduced anticoagulant activity, 5A-APC and 3K3A-APC, but not protein C, exhibited high affinity for the APC BBB transport system. Blockade of APC binding site on endothelial protein C receptor (EPCR), but not blockade of its protease activated receptor-1 (PAR1) catalytic site, inhibited by > 85% APC entry into the brain. APC brain uptake was reduced by 64% in severely-deficient EPCR mice, but not in PAR1 null mice. These data suggest that APC and its variants with reduced anticoagulant activity cross the BBB via EPCR-mediated saturable transport.
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