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目的探讨粒细胞-巨噬细胞集落刺激因子(GM-CSF)对体外培养大鼠海马神经元氧糖剥夺(OGD)后凋亡的影响。方法体外培养乳鼠海马神经元并分为正常对照组、OGD组、GM-CSF1ng/ml、10ng/ml、20ng/ml和100ng/ml组;制备OGD模型,并给与相应剂量的GM-CSF干预。应用流式细胞仪及Annexin V/PI双染色法检测神经元凋亡率,测定上清液乳酸脱氢酶(LDH)活性了解神经元细胞膜损伤程度,RT-PCR法检测凋亡相关基因Bcl-2、Bax mRNA表达。结果与正常对照组相比,OGD组细胞凋亡率及LDH活性明显升高(均P<0.01);与OGD组相比,除GM-CSF1ng/ml组外,GM-CSF各浓度组神经元凋亡率及LDH活性明显降低(均P<0.01),其中GM-CSF20ng/ml组对神经元凋亡影响最显著。与正常对照组比较,OGD组Bcl-2 mRNA表达明显下降,Bax mRNA表达显著升高,Bcl-2/Bax降低(均P<0.01);与OGD组比较,GM-CSF20ng/ml组Bcl-2 mRNA表达升高,Bax mRNA表达降低,Bcl-2/Bax增加(均P<0.01)。结论 GM-CSF能够有效地抑制OGD后海马神经元的凋亡,20ng/ml抗凋亡的效果最佳。其神经保护作用机制可能与其调节凋亡蛋白Bcl-2、Bax表达有关。
Objective To investigate the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) on the apoptosis of cultured rat hippocampal neurons after OGD. Methods The hippocampal neurons were cultured in vitro and divided into normal control group, OGD group, GM-CSF 1ng / ml, 10ng / ml, 20ng / ml and 100ng / ml group; OGD model was prepared and the corresponding dose of GM-CSF Intervention. The apoptotic rate of neurons was detected by flow cytometry and Annexin V / PI double staining. The activity of lactate dehydrogenase (LDH) in the supernatant was assayed for the degree of neuronal cell membrane damage. The apoptosis-related gene Bcl- 2, Bax mRNA expression. Results Compared with the normal control group, the apoptotic rate and LDH activity in OGD group were significantly increased (all P <0.01). Compared with OGD group, in addition to the GM-CSF 1ng / ml group, The apoptotic rate and LDH activity were significantly decreased (all P <0.01), of which the GM-CSF 20ng / ml group had the most significant effect on neuronal apoptosis. Compared with the normal control group, the expression of Bcl-2 mRNA in OGD group was significantly decreased, the expression of Bax mRNA and Bcl-2 / Bax in OGD group were significantly decreased (all P <0.01) mRNA expression increased, Bax mRNA expression decreased, Bcl-2 / Bax increased (all P <0.01). Conclusion GM-CSF can effectively inhibit the apoptosis of hippocampal neurons after OGD. The anti-apoptotic effect of 20ng / ml is the best. Its neuroprotective mechanism may be related to its regulation of apoptosis protein Bcl-2, Bax expression.