Exendin-4抑制缺血再灌注损伤大鼠皮质神经元凋亡及可能机制  

Protective effect and possible mechanism of Exendin-4 on apoptosis of rat cortical neuron induced by in vitro ischemia/reperfusion

作　　者：王梦嵽[1] 李俊敏[1] 方瑗[1] 梅元武[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院神经内科,武汉430022

出　　处：《中华神经科杂志》2011年第4期242-246,共5页Chinese Journal of Neurology

摘　　要：目的 探讨Exendin-4对大鼠脑缺血再灌注损伤的保护作用及机制.方法 体外培养SD乳鼠皮质神经元并采用随机数字表法将其分为对照组（正常培养）、模型组（体外模拟缺血再灌注）、治疗组（建模前、建模中均给予Exendin-4）.免疫荧光染色鉴定神经元纯度,RT-PCR检测胰高血糖素样肽-1受体（GLP-1R）mRNA表达,ELISA法检测神经元细胞内cAMP水平,MTT法测定神经元存活率,荧光染料Hoechest 33258检测神经元凋亡率,实时定量PCR检测干预后4～12 h C/EBP同源蛋白（CHOP）、生长抑制和DNA损伤基因34（GADD34）基因表达.结果 大鼠皮质神经元可体外纯化培养.皮质神经元存在活性GLP-1R.皮质神经元体外模拟缺血6 h再灌注12 h后,治疗组神经元凋亡率明显下降（分别为77.0%±5.3%和27.0%±3.5%,t=19.462,P＜0.01）.体外模拟缺血再灌注损伤可诱导神经元CHOP mRNA、GADD34 mRNA表达增加.Exendin-4干预后CHOP mRNA表达明显增加,峰值提前,后期降至模型组水平（t4h=3.023,P4h=0.039;t8h=6.988,P8 h=0.002）;GADD34 mRNA表达显著增加,峰值提前,干预4、8、12 h表达均明显高于对照组（t=11.036、26.098、11.709,P＜0.01）.结论 Exendin-4对大鼠皮质神经元体外模拟缺血再灌注损伤具有保护作用,其机制可能为通过干预未折叠蛋白反应抑制损伤诱导的内质网相关性细胞凋亡.Objective To investigate the effect of Exendin-4 （Ex-4） on ischemia/reperfusion （I/R） injury-induced apoptosis in primary rat cortical neurons and the possible underlying mechanisms.Methods Rat cortical neurons were cultured in vitro,identified by NES-immunohistological staining and immunofluorescence staining,and randomly divided into the following groups： control group,I/R group and Ex-4 group.RT-PCR was performed to establish the existence of active glucagon-like peptide-1 receptor （GLP-1R）.ELISA was used to measure the neuronal cytoplasmic cAMP level. MTT was used to detect viability. Fluorescent DNA binding dye Hoechest 33258 was used to reveal apoptosis. C/EBP-homologous protein （CHOP） and growth arrest and DNA-damage-inducible gene 34 （GADD34） mRNAs were detected by real-time PCR. Results The apoptosis rate induced by ischemia 6 h/reperfusion 12 h was 77.0% ±5.3% and was decreased to 27.0% ± 3.5% after Ex-4 （ 0. 4 μg/ml ） treating （ t = 19. 462,P ＜ 0. 01 ）.Levels of CHOP and GADD34 mRNA in cortical neurons increased since 4 h and peaked at 12 h after reperfusion. Ex-4 group showed a sharp elevation of levels of CHOP and GADD34 mRNA,peaking at 8 h reperfusion,and then tended to decrease.Conclusions Ex-4 has protective effect against rat cortical neurons injury induced by ischemia/reperfusion. The protective effect may be related to inhibition of ESR-related neuron apoptosis via regulation of unfolded protein response.

关 键 词：再灌注损伤 大脑皮质 神经元 细胞凋亡 肽类 毒液类 大鼠 

分 类 号：R285.5[医药卫生—中药学]