APOE基因亚型及其短肽COG1410对神经细胞损伤后早期凋亡的影响

Effect of APOE alleles and apoE-mimetic peptide COG1410 on early neuron apoptosis after in vitro traumatic brain injury

目的探讨载脂蛋白E(APOE代表基因,apoE代表蛋白)亚型特异性及其短肽与早期凋亡的相关性,以期明确APOE亚型影响脑创伤病情转归及预后的病理机制。方法采用稳定表达人APOE各等位基因的APOE敲除鼠的神经干细胞,优化诱导分化条件,构建神经元/胶质细胞共培养体系及细胞划痕损伤模型。通过Annexin V/PI联合流式细胞技术检测损伤后各组细胞早期凋亡率,分析人APOE各等位基因及其短肽影响继发性神经细胞损伤的差异。结果成功构建携带人源性APOE各亚型的细胞划痕损伤模型;伤后24 h时间点各组细胞早期凋亡率较6、12 h明显增高(P<0.05),且人APOEε4组较其余亚型组早期凋亡率明显增高(P<0.05)。在伤后24 hapoE短肽COG1410可显著降低各亚型组早期凋亡率(P<0.01)。结论 APOEε4携带者可能通过早期凋亡导致脑创伤急性期病情加重。而apoE短肽COG1410可通过降低早期凋亡发挥神经保护作用。

Objective To elucidate the mechanism mediating the effects of apolipoprotein E （APOE for gene, apoE for protein） alleles and apoE-mimetic peptide COG1410 on early apoptosis in an in vitro model of experimental traumatic brain injury. Methods Eukaryotic expression vectors carrying individual APOE alleles （ε2, ε3, ε4） were transferred into neural stem cells （NSCs） derived from APOE knockout mice. An in vitro neuronal/glial co-cuhure model of mechanical injury was developed using a controlled scratch. Flow cytometry was performed to analyze the correlations among APOE genotypes, apoE-mimetic peptide COG1410 and early apoptosis. Results Early apoptosis after injury was observed in all groups, which was significantly higher at 24 h after injury than at 6 h and 12 h after injury （P 〈0. 05）. The group transfeeted with APOE84 showed more significant early apoptosis 24 h after injury as compared to the groups transfected with APOEε2 or APOE83 （P 〈 0. 05 ）. Furthermore, decreases of early apoptosis were detected in the groups treated with apoE- mimetic peptide COG1410 at 24 h after injury （P 〈0.01 ）. Conclusion APOEε4-trasfected NSCs showing a higher rate of early apoptosis indicates that the patients with APOEε4 may suffer aggravation of brain trauma in acute stage due to early apoptosis. Our results also identify the early neuroprotective effect of apoE-mimetic peptide COG1410.