内皮素受体拮抗剂BQ-123经PI3-K/Akt信号通路改善蛛网膜下腔出血大鼠早期脑损伤的效果

Effect observation of endothelin A receptor antagonist BQ-123 improves early brain injury of subarachnoid hem orrhage in rats via PI3 K-Akt signaling pathway

目的探讨内皮素受体拮抗剂BQ-123对蛛网膜下腔出血(SAH)后早期脑损伤的治疗作用及其机制。方法按随机数字表法将120只SD大鼠分为假手术组、SAH组、低剂量BQ-123组(50μg/kg)和高剂量BQ-123组(75μg/kg),每组30只。采用枕大池二次注血法制作SAH大鼠模型,每组造模后进一步分为6、24、72、144 h 4个时间点亚组。采用光镜和电镜观察海马区形态结构变化,免疫组化和逆转录聚合酶链反应检测海马区磷脂酰肌醇3-激酶(PI3-K)、蛋白激酶B(PKB/Akt)和雷帕霉素靶蛋白(m TOR)的表达。结果 (1)模型制作过程中,SAH组死亡7只,1只模型不符合标准被剔除;BQ-123高、低剂量组分别死亡6只,各组有1只模型不符合标准被剔除。最终纳入统计学分析:假手术组30只,SAH组22只,BQ-123高、低剂量组均为23只大鼠。(2)与假手术组比较,SAH组各时间点PI3-K、AKt、m TOR的表达增高(均P<0.05)。与SAH组比较,BQ-123低剂量组海马区神经元形态结构损伤减轻,各时间点存活神经元数量增加[(132±18),(110±16),(84±13),(92±10)个/高倍视野](均P<0.05),大鼠抓力值有所升高,学习记忆功能改善;PI3-K和Akt表达进一步升高(均P<0.05),m TOR的表达下降(均P<0.05)。(3)与BQ-123低剂量组各时间点存活神经元比较,BQ-123高剂量组海马区神经元形态结构损伤减轻,各时间点[存活神经元数量(153±20)、(131±18)、(137±19)、(135±17)个/高倍视野]增加(均P<0.05),大鼠抓力值有所升高,动物学习记忆功能改善;PI3-K(3.8±0.8、8.9±2.4、8.6±2.4、6.2±2.0)、Akt(4.86±1.74、8.64±1.62、7.94±1.70、6.48±1.58)表达进一步增多(均P<0.05),m TOR(2.89±0.26、2.14±0.18、1.94±0.17、1.62±0.12)的表达下降(均P<0.05)。结论 BQ-123对SAH后早期脑损伤有较好的治疗作用,其机制可能与调控PI3-K/Akt信号途径有关。

Objectives To investigate the treatment effect of endothclin A receptor antagonist BQ-123 on early brain injury of suharachnoid hemorrhage （SAH） in rats and its mechanism. Methods According to the random number table method, 120 SD rats were divided into four groups： a sham operation （sham） ,a SAH ,a high-dose BQ-123 （ 75 μg/kg） ,and a low-dose BQ-123 （50 μg/kg） （ n = 30 in each group）. A rat model was induced by using the injection of blood into cisterna magna twice. After establishing models at hours 6,24,72, and 144, the rats were further divided into four subgroups. Light and electron microscopes were used to observe the changes of the morphological structure in hippocampal area. Immmunohistocbemistry and RT-PCR were used to detect the expression levels of of phosphoinesifide 3 kina （ PI3- K）,proteinkinaseB （PKB/Akt）,and mammal target of rapmnycin （mTOR）. Resnlts （1） In the process of model making,7 rats died and 1 model did not meet the criteria and was excluded from the SAH group. Six rats died in the high-dese BQ-123 group and the low-dese BQ-123 group respectively. One rat in each group did not meet the criteria and was excluded. The rats were included in the final statistical analysis： 30 in the sham group,22 in the SAH group,23 in the high-dese BQ-123 group,and 23 in the low-dose BQ-123 group. （2）Compared with the sham group,the expression levels of PI3-K,AKt and roTOR were increased signifi- cantly （all P 〈0. 05）. Compared with SAH group,the hippocampal neuronal morphology and structure damage were alleviated in the low-dese BQ-123 group. The number of surviving neurons at each time point was increased （[132±18],[110±16] ,[84±13] ,[92±10] cells/HP,all P〈0.05）. The teasile force values of rats were increased at each time point and the learning and memory function were improved. The expression levels of PI3-K and Akt were further increased （ all P 〈 0. 05）. The expression level of roTOR was decreased （ all P 〈0. 05）. （3）Compared with the low-dose BQ-123 group,the morphological and structural damage of hippocampal neurons were alleviated. The number of smviving neurons at each time point （[153 ±20] ,[131 ± 18] ,[137 ± 19] and [ 135 ± 17] cells/HP） was increased （ all P 〈0.05）. The tensile force values of the rats were increased at each time point. The learning and memory function of the animals were improved. The expression levels of PI3 -K （ 3. 8 ± 0.8,8.9 ± 2.4,8.6 ± 2.4, and 6.2 ± 2.0） and Akt （4.86 ± 1.74, 8.64 ± 1.62,7.94 ± 1.70, and 6.48 ± 1.58） were further increased （all P 〈 0. 05）. The expression levels of roTOR （2.89 ±0.26,2.14 ±0.18,1.94 ±0. 17,and 1.62 ±0. 12） were decreased （all P 〈0.05）. Conclusions BQ-123 has as a good therapeutic effect for early brain injury after SAH. Its mechanism may be associated with the regulation of PI3-K/Akt signaling pathway.