淫羊藿苷对脑缺血再灌注损伤大鼠的脑保护作用机制研究

The protective mechanism of icariin on cerebral ischemia reperfusion injury in rats

目的 探讨磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路参与淫羊藿苷对脑缺血再灌注损伤(CIRI)大鼠的作用机制。方法 SD大鼠按随机数字表法将分成为正常组、模型组和实验组,每组15只。向大鼠大脑右侧中动脉插入线栓模拟缺血2 h后缓慢拔出制备CIRI模型。模型组和实验组大鼠构建CIRI模型,正常组大鼠仅切开颈部皮肤暴露右侧中动脉。术前30 min,实验组灌胃100 mg·kg^(-1)淫羊藿苷,正常组和模型组灌胃等体积的生理盐水。再灌注24 h后,用Zea-Longa’s 5级标准评分法对3组大鼠进行神经功能的缺损性评价,测定脑组织的干湿重评价脑含水量,用原位末端转移酶标记技术(TUNEL)进行神经元凋亡检测,用蛋白质印迹法检测患侧脑组织PI3K/Akt蛋白的表达水平。结果 正常组、模型组和实验组的神经功能缺损评分分别为(0.00±0.00),(2.53±0.36)和(2.04±0.31)分;这3组脑含水量分别为(61.33±5.69)%,(89.34±5.24)%和(75.34±3.19)%;这3组TUNEL染色细胞凋亡率分别为(0.00±0.00)%,(87.11±13.48)%和(48.98±11.28)%;这3组p-P13K蛋白表达水平分别为0.63±0.03,0.11±0.02和0.36±0.03;这3组p-Akt蛋白表达水平分别为1.77±0.19,0.91±0.08和1.35±0.06。上述指标,模型组与正常组相比,或实验组与模型组相比,差异均有统计学意义(均P<0.05)。结论 淫羊藿苷发挥神经保护作用可能与激活PI3K/Akt信号通路,抑制细胞凋亡有关。

Objective To study the effect and mechanism of phosphatidylinositol 3-kinase/protein kinase B(PI3 K/Akt) signaling pathway involved in icariin effects on cerebral ischemia/reperfusion injury(CIRI) in rats. Methods SD rats were randomly divided into normal group, model group and experimental group, with 15 rats in each group. A suture was inserted into the right middle cerebral artery of rats to simulate ischemia for 2 h and then slowly pulled out to prepare CIRI model. The CIRI model was established in the model group and the experimental group, and the rats in the normal group were only exposed to the right middle artery by cutting the neck skin. The rats in experimental group was gavaged with icariin at a dose of 100 mg·kg^(-1) at 30 min before plugging. The rats in normal group and model group were given equal volume normal saline by gavage.At 24 h after reperfusion, neurologic impairment was evaluated by Zea-longa’s Standard Grade 5 scoring method;cerebral water content was evaluated by measuring wet and dry weight of brain tissue;neuronal apoptosis was tested by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay( TUNEL);PI3K/Akt were tested by Western blotting. Results Neurological deficit score in normal group,model group,and experimental group were( 0. 00 ± 0. 00),( 2. 53 ± 0. 36) and( 2. 04 ± 0. 31) scores;cerebral water content in the three groups were( 61. 33 ± 5. 69) %,( 89. 34 ± 5. 24) % and( 75. 34 ± 3. 19) %;apoptosis rate in the three groups were( 0. 00 ± 0. 00) %,( 87. 11 ± 13. 48) % and( 48. 98 ± 11. 28) %;the expression levels of p-PI3K in the three groups were 0. 63 ± 0. 03,0. 11 ± 0. 02,and 0. 36 ± 0. 03;the expression levels of p-Akt in the three groups were 1. 77 ± 0. 19,0. 91 ± 0. 08 and 1. 35 ± 0. 06. Compared between model group and normal group,the difference of the above indexes were significant( all P < 0. 05);compared between experimental group and model group,the difference of the above indexes were significant( all P < 0. 05). Conclusion The neuroprotective effect of icariin may be related to the activation of PI3K/Akt signaling pathway and the inhibition of cell apoptosis.