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目的探讨蛋白酶激活受体-1(protease activated receptor-1,PAR-1)激活剂SFLLRN对体外培养的脑血管内皮细胞的影响,明确凝血酶增加血脑屏障通透性的可能机制。方法将脑微血管内皮细胞在体外进行培养,在细胞的培养液中加入SFLLRN或凝血酶,应用相差显微镜动态观察SFLLRN或TM对内皮细胞形态的影响,应用免疫组织细胞化学技术检测基质金属蛋白酶2(matrix metalloproteinase-2,MMP-2)表达的变化。结果 SFLLRN加入后1h内皮细胞开始收缩,细胞面积变小,细胞间隙增宽,细胞收缩程度具有时间依赖性,至12h时细胞面积仅为处理前的20%。凝血酶加入培养液后,细胞形态学的变化方式与SFLL-TRN组相似。SFLLTRN孵育后6h,内皮细胞MMP-2的表达水平开始增加,24小时达高峰,与对照组比较,存在明显统计学差异(P<0.01),与TM组比较,无明显统计学差异(P>0.05)。结论凝血酶通过激活PAR-1,使内皮细胞发生收缩,促进MMP-2表达,是凝血酶增加BBB通透性的可能机制。
Objective To investigate the effect of protease activated receptor-1 (PAR-1) activator SFLLRN on cultured human cerebrovascular endothelial cells and to explore the possible mechanism by which thrombin increases the permeability of the blood-brain barrier. Methods The cerebral microvascular endothelial cells were cultured in vitro. SFLLRN or thrombin was added to the culture medium of the cells. The effect of SFLLRN or TM on the morphology of endothelial cells was observed by phase-contrast microscopy. The expression of matrix metalloproteinase 2 ( matrix metalloproteinase-2, MMP-2). Results After 1h SFLLRN injection, endothelial cells began to shrink, the cell area became smaller, the cell gap widened and the degree of cell contraction was time-dependent. By 12h, the cell area was only 20% before treatment. Thrombin added to the culture medium, the morphological changes of cells and SFLL-TRN group similar. Six hours after SFLLTRN incubation, the expression of MMP-2 in endothelial cells began to increase and peaked at 24 hours (P <0.01). Compared with the control group, there was no significant difference (P> 0.05). Conclusions Thrombin can activate endothelial cell by contracting PAR-1 and promoting the expression of MMP-2. Thrombin is a possible mechanism of thrombin increasing BBB permeability.