目的 探讨蜕皮甾酮(EDS)对蛛网膜下腔出血(SAH)后家兔脑组织的影响及其机制.方法 选择6～9月龄雄性日本大耳兔18只,体重2.2～3.4kg,随机分为假手术对照组(L组)、SAH组和ⅡS组,每组6只.采用结扎右侧颈总动脉后二次枕大池注血的方法制作SAH模型,EDS组静脉给予EDS(3mg/kg,4次/d,共3d).通过Strong神经学评分观察各组动物神经功能的变化.3d后处死动物,测定脑含水量,HE染色观察海马组织结构变化,免疫组化染色观察海马组织Bax及Bcl-2的表达变化,TUNEL染色观察海马神经元凋亡情况,并采用PCR和Westem blotting观察海马P13K mRNA和蛋白的表达情况.结果 建模后第1,2,3天SAH组神经功能评分分别为6.00±1.79、8.00±1.90、8.00±2.37,而EDS组分别为4.16±0.75、5.17±0.75、4.83±1.17,其中第2、第3天EDS组神经功能明显优于SAH组(P＜0.05,P＜0.01).3d时测定SAH组家兔脑含水量为78.6％±0.5％,明显高于L组(77.7％±0.4％,P＜0.01),而EDS组经治疗后脑含水量(78.1％±0.5％)明显下降,与SAH组比较差异有统计学意义(P＜0.05).HE染色观察见SAH组家兔海马CA1区神经无排列紊乱,细胞固缩、深染,而EDS组CA1区神经元结构基本正常.免疫组化染色显示,与L组比较,SAH组Bax表达增加、Bcl-2表达减弱,而EDS组Bax、Bcl-2表达水平与L组接近.TUNEL染色显示,SAH组凋亡细胞明显增多,而EDS组凋亡细胞数与L组接近.PCR和Western blotting结果显示,SAH组PI3K mRNA和蛋白表达均明显低于L组和EDS组,后两组比较差异无统计学意义.结论 EDS可减轻家兔SAH后的神经损害,其机制可能与通过PI3K途径抑制神经细胞凋亡有关.%Objective The present study aims to probe into the effect and mechanism of ecdysterone (EDS) on rabbit brains after suffering subarachnoid hemorrhage (SAH). Methods A total of 18 male Japanese rabbits, aged 6 months to 9 months and weighing 2. 2 kg to 3.4 kg, were randomly divided into three groups. Each group, namely, the Sham operation control (L), SAH, and ECS groups, had six rabbits. The SAH group was treated using the quadric cisterna magna blood injection method followed by the ligation of the right common carotid artery, whereas the EDS group was treated with EDS intravenously (3 mg/kg, 4 times/day, 12 times in all). The changes in the neurological function of the animals in each group were observed using the Strong neurological score. Then, the animals were killed after three days to measure their brain water contents. The changes in the hippocampal tissue structure and in the Bax and Bcl-2 expressions in the hippocampal tissue were observed using HE and immunohistochemistry staining, respectively. The apoptosis of the hippocampal neuron was also observed by TUNEL staining. Finally, PCR and Western blot analysis were adopted to observe the PI3K mKNA and protein expression in the hippocampus. Results The neurological scores of the SAH group for the first three days after modeling were 6. 00±l. 79, 8. 00±l. 90, and 8. 00±2. 37, respectively, whereas those for the EDS/SAH group were 4. 16±0. 75, 5. 17 ±0. 75, and 4. 83±1. 17. The neurological functions of the rabbits in the EDS group (P<0. 05) two to three days after treatment were evidently better than those in the SAH group ( P<0. 01). The brain water contents in the SAH group (78. 6%±0. 5%) were higher than those in the L group (77. 7 % ± 0. 4 %, P<0. 01), whereas the brain water contents in the EDS group after treatment (78. 1 % + 0. 5 %) were significantly lower than those in the SAH group (P<0. 05). HE staining revealed that the neurons in the hippocampal CA1 regions of the rabbits in the SAH group were disorganized due to cell pyenosis and hyperchromatism, whereas those of the rabbits in the EDS group were normal. Immunohistochemistry staining showed that the Bax and Bcl-2 expressions were higher and lower, respectively, in the SAH group, compared with those in the L group. Meanwhile, the Bax and Bcl-2 expressions in the EDS group were almost the same as those in the L group. TUNEL staining showed an evident increase in the apoptotic cells in the SAH group, whereas the number of apoptotic cells in the EDS group was close to that in the L group. Lastly, the PCR and Western blot analysis revealed that the PI3K mRNA and protein expressions in the SAH group were evidently lower than those in the L and EDS groups, whereas no statistical difference was observed between the results for the latter groups. Conclusion EDS can mitigate the cerebral injuries of rabbits after suffering SAH. Its mechanism may be related to the inhibition of nerve apoptosis through the PI3K pathway.
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