From the Departments of Physiology and Pharmacology (B.C.T., O.A., K.D., P.R.K., J.Y., J.H.Z.) and Neurosurgery (J.H.Z.), Loma Linda University School of Medicine, CA.
Stroke. 2013 Nov;44(11):3189-94. doi: 10.1161/STROKEAHA.113.001574. Epub 2013 Sep 5.
Neuronal apoptosis is a key pathological process in subarachnoid hemorrhage (SAH)-induced early brain injury. Given that recombinant osteopontin (rOPN), a promising neuroprotectant, cannot pass through the blood-brain barrier, we aimed to examine whether nasal administration of rOPN prevents neuronal apoptosis after experimental SAH.
Male Sprague-Dawley rats (n=144) were subjected to the endovascular perforation SAH model. rOPN was administered via the nasal route and neurological scores as well as brain water content were evaluated at 24 and 72 hours after SAH induction. The expressions of cleaved caspase-3, phosphorylated focal adhesion kinase (FAK), and phosphorylated Akt were examined using Western blot analysis. Neuronal cell death was demonstrated with terminal deoxynucleotid transferase-deoxyuridine triphosphate (dUTP) nick end labeling. We also administered FAK inhibitor 14 and phosphatidylinositol 3-kinase inhibitor, Wortmannin, prior to rOPN to establish its neuroprotective mechanism. ELISA was used to measure rOPN delivery into the cerebrospinal fluid.
Cerebrospinal fluid level of rOPN increased after its nasal administration. This was associated with improved neurological scores and reduced brain edema at 24 hours after SAH. rOPN increased phosphorylated FAK and phosphorylated Akt expressions and decreased caspase-3 cleavage, resulting in attenuation of neuronal cell death within the cerebral cortex. These effects were abolished by FAK inhibitor 14 and Wortmannin.
Nasal administration of rOPN decreased neuronal cell death and brain edema and improved the neurological status in SAH rats, possibly through FAK-phosphatidylinositol 3-kinase-Akt-induced inhibition of capase-3 cleavage.
神经元凋亡是蛛网膜下腔出血（SAH）诱导早期脑损伤的关键病理过程。由于重组骨桥蛋白（rOPN）是一种有前途的神经保护剂，无法穿过血脑屏障，因此我们旨在研究鼻内给予 rOPN 是否可以预防实验性 SAH 后的神经元凋亡。
雄性 Sprague-Dawley 大鼠（n=144）接受血管内穿孔 SAH 模型。通过鼻内途径给予 rOPN，并在 SAH 诱导后 24 和 72 小时评估神经功能评分和脑水含量。使用 Western blot 分析检测裂解的 caspase-3、磷酸化粘着斑激酶（FAK）和磷酸化 Akt 的表达。使用末端脱氧核苷酸转移酶-脱氧尿苷三磷酸（dUTP）缺口末端标记法显示神经元细胞死亡。我们还在给予 rOPN 之前给予 FAK 抑制剂 14 和磷脂酰肌醇 3-激酶抑制剂 Wortmannin，以确定其神经保护机制。ELISA 用于测量 rOPN 递送至脑脊液中的水平。
鼻内给予 rOPN 后，脑脊液中的 rOPN 水平增加。这与 SAH 后 24 小时神经功能评分改善和脑水肿减少有关。rOPN 增加了磷酸化 FAK 和磷酸化 Akt 的表达，并减少了 caspase-3 的裂解，从而减轻了大脑皮质内的神经元细胞死亡。这些作用被 FAK 抑制剂 14 和 Wortmannin 所消除。
鼻内给予 rOPN 可减少神经元细胞死亡和脑水肿，并改善 SAH 大鼠的神经状态，可能通过 FAK-磷脂酰肌醇 3-激酶-Akt 诱导的 caspase-3 裂解抑制作用。
