Department of Physiology, Loma Linda University School of Medicine, Loma Linda, Calif 92354, USA.
Stroke. 2010 Aug;41(8):1783-90. doi: 10.1161/STROKEAHA.110.586537. Epub 2010 Jul 8.
Osteopontin (OPN) is an inducible, multifunctional, extracellular matrix protein that may be protective against blood-brain barrier (BBB) disruption after subarachnoid hemorrhage (SAH). However, the protective mechanisms remain unclear.
We produced the endovascular perforation model of SAH in rats and studied the time course of OPN induction in brains by Western blotting and immunofluorescence (n=50). Then, 34 rats were randomly assigned to sham (n=3), sham+OPN small interfering RNA (siRNA, n=3), SAH+negative control siRNA (n=14), and SAH+OPN siRNA (n=14) groups, and 109 rats were allocated to sham+vehicle (n=17), sham+recombinant OPN (n=17), SAH+vehicle (n=33), SAH+recombinant OPN (n=31), and SAH+recombinant OPN+L-arginyl-glycyl-L-aspartate motif-containing hexapeptide (n=11) groups. The effects of OPN siRNA or recombinant OPN on BBB disruption and related proteins were studied.
OPN was significantly induced in reactive astrocytes and capillary endothelial cells, peaking at 72 hours after SAH, during the recovery phase of BBB disruption. Blockage of endogenous OPN induction exacerbated BBB disruption and was associated with a reduction of angiopoietin-1 and mitogen-activated protein kinase (MAPK) phosphatase-1 (an endogenous MAPK inhibitor), activation of MAPKs, and induction of vascular endothelial growth factor-A at 72 hours after SAH, whereas recombinant OPN treatment improved it and was associated with MAPK phosphatase-1 induction, MAPK inactivation, and vascular endothelial growth factor-A reduction, which was blocked by L-arginyl-glycyl-L-aspartate motif-containing hexapeptide at 24 hours after SAH. Vascular endothelial growth factor-B and angiopoietin-2 levels were unchanged.
OPN may increase MAPK phosphatase-1 that inactivates MAPKs, upstream and downstream of vascular endothelial growth factor-A, by binding to L-arginyl-glycyl-L-aspartate-dependent integrin receptors, suggesting a novel mechanism of OPN-induced post-SAH BBB protection.
骨桥蛋白（OPN）是一种诱导型、多功能的细胞外基质蛋白，可能对蛛网膜下腔出血（SAH）后血脑屏障（BBB）的破坏具有保护作用。然而，其保护机制尚不清楚。
我们在大鼠中制作了血管内穿孔模型，通过 Western blot 和免疫荧光法研究了脑内 OPN 诱导的时间过程（n=50）。然后，34 只大鼠被随机分为假手术（n=3）、假手术+OPN 小干扰 RNA（siRNA，n=3）、SAH+阴性对照 siRNA（n=14）和 SAH+OPN siRNA（n=14）组，109 只大鼠被分配到假手术+载体（n=17）、假手术+重组 OPN（n=17）、SAH+载体（n=33）、SAH+重组 OPN（n=31）和 SAH+重组 OPN+L-精氨酰-甘氨酰-L-天冬氨酸基序六肽（n=11）组。研究了 OPN siRNA 或重组 OPN 对 BBB 破坏及相关蛋白的影响。
OPN 在反应性星形胶质细胞和毛细血管内皮细胞中明显诱导，在 SAH 后 72 小时达到峰值，此时 BBB 破坏的恢复阶段。内源性 OPN 诱导阻断加重了 BBB 破坏，并与血管生成素-1 和丝裂原活化蛋白激酶（MAPK）磷酸酶-1（一种内源性 MAPK 抑制剂）减少、MAPK 激活以及 SAH 后 72 小时血管内皮生长因子-A 的诱导有关，而重组 OPN 治疗则改善了这一点，并与 MAPK 磷酸酶-1 的诱导、MAPK 的失活以及血管内皮生长因子-A 的减少有关，SAH 后 24 小时 L-精氨酰-甘氨酰-L-天冬氨酸基序六肽阻断了这一作用。血管内皮生长因子-B 和血管生成素-2 水平无变化。
OPN 可能通过与 L-精氨酰-甘氨酰-L-天冬氨酸依赖性整合素受体结合，增加 MAPK 磷酸酶-1，使 MAPK 失活，从而使血管内皮生长因子-A 的上下游信号转导，提示 OPN 诱导的 post-SAH BBB 保护的一种新机制。
