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目的探讨脑血管痉挛(CVS)后细胞外信号调节激酶(ERK)途径的作用机制,及重组载脂蛋白E(apoE)拟肽对脑血管痉挛损伤的保护作用。方法采用非开颅血管内线栓法制备小鼠蛛网膜下腔出血(SAH)并脑血管痉挛模型。将134只健康雄性ICR小鼠随机分为4组:假手术组、模型对照组、拮抗剂组,apoE治疗组。ApoE治疗组将拟apoE-1410以无菌磷酸盐缓冲液溶解后,经尾静脉注射0.6mg/kg,术前30min开始第1次,每12h1次。拮抗剂组于SAH后10 min侧脑室注射生理盐水或LY367385(500nmol/L)5μl,术后行神经功能评分。分别在SAH后6、24、48h 3个时相点取脑组织标本,在光镜和电镜下观察脑组织病理变化,采用反转录-聚合酶链式反应(RT-PCR)检测各组代谢型谷氨酸受体1(mG1uR1)mRNA的表达变化;免疫印迹法(Western blotting)检测mGluR1、磷酸化细胞外信号调节激酶(ERK1/2)蛋白的表达;用原位缺口末端标记法(TUNEL)检测神经细胞凋亡情况。结果与假手术组比较,模型对照组小鼠神经功能评分显著降低,随脑血管痉挛时间延长,各组小鼠mGluR1 mRNA、mGluR1和磷酸化ERK1/2蛋白均有不同程度增加,凋亡细胞增多,神经细胞出现变性坏死、细胞器数量减少。与模型对照组比较,拮抗剂组和apoE治疗组小鼠神经功能评分增加,mGluR1 mRNA、mGluR1、磷酸化ERK1/2蛋白表达均有不同程度下调,神经细胞凋亡数目减少,应用apoE1410拟肽减轻了脑组织形态学和超微结构损伤。结论脑血管痉挛后mGluR1的表达增强可通过激活ERK信号途径诱导神经细胞凋亡,apoE拟肽对脑血管痉挛损伤具有抗损坏作用。
Objective To investigate the mechanism of extracellular signal-regulated kinase (ERK) pathway after cerebral vasospasm (CVS) and the protective effect of apoE peptidomimetic against cerebral vasospasm injury. Methods A non-craniotomy intra-arterial instillation method was used to prepare mouse models of subarachnoid hemorrhage (SAH) and cerebral vasospasm. 134 healthy male ICR mice were randomly divided into 4 groups: sham operation group, model control group, antagonist group, apoE treatment group. ApoE treatment group will be apoE-1410 to be dissolved in sterile phosphate buffer, the tail vein injection of 0.6mg / kg, 30min before the start of the first, every 12h1 times. The antagonist group was intracerebroventricular injection of saline or LY367385 (500nmol / L) 5μl 10 min after SAH, and neurological function score was performed after operation. The brain tissue samples were taken at 6, 24, and 48h after SAH respectively. The pathological changes of brain tissue were observed under light microscope and electron microscope. The metabolism of each group was detected by reverse transcription-polymerase chain reaction (RT-PCR) The expression of mGluR1 and phosphorylated extracellular signal-regulated kinase (ERK1 / 2) was detected by Western blotting. The expression of mGluR1 mRNA was detected by TUNEL ) Detected neuronal apoptosis. Results Compared with the sham operation group, the neurological scores of the model control group were significantly decreased. The mGluR1 mRNA, mGluR1 and phosphorylated ERK1 / 2 protein of the mice in each group increased to different extents and apoptotic cells increased with the time of vasospasm , Degeneration and necrosis of nerve cells, reducing the number of organelles. Compared with the model control group, the neurological scores of the mice in the antagonist group and the apoE treatment group increased, the expressions of mGluR1 mRNA, mGluR1 and phosphorylated ERK1 / 2 decreased to different extents, and the number of apoptotic neurons decreased. The apoE1410 peptoid decreased Brain tissue morphology and ultrastructure damage. Conclusions The increased expression of mGluR1 after cerebral vasospasm can induce neuronal apoptosis through activation of ERK signaling pathway. ApoE-peptidomimetic has anti-damaging effects on cerebral vasospasm injury.