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目的:以原代培养的大鼠海马神经细胞缺氧/缺糖再给氧为模型,观察三七总皂甙对神经细胞凋亡的抑制作用。方法:流式细胞术检测凋亡细胞百分率,激光共聚焦扫描显微镜检测细胞内Ca2+浓度,荧光显微镜观察细胞形态学变化和坏死细胞百分率,同时,测定细胞LDH的释放。结果:神经细胞缺氧/缺糖5h后再给氧3h时,细胞凋亡百分率明显增高,为19.06%(P<0.01),细胞坏死百分率为6.83%(P<0.01),细胞内Ca2+浓度为169.32nmol/LCa2+浓度(P<0.01),LDH的释放率为27.63%(P<0.01);神经细胞缺氧/缺糖5h后再给氧24h时,细胞凋亡百分率为49.85%(P<0.01),细胞坏死百分率为11.49%(P<0.01),细胞内Ca2+浓度为298.11nmol/L(P<0.01),LDH的释放率为60.35%(P<0.01)。三七总皂甙(25,50mg/L)能降低神经细胞凋亡及坏死的百分率,降低细胞内Ca2+浓度,减少LDH的释放,三七总皂甙的作用随剂量增加而作用增加。结论:三七总皂甙对缺血再灌注损伤后海马神经元的凋亡过程具有抑制作用,这种作用可能与其能降低细胞内Ca2+浓度有关。
OBJECTIVE: To observe the inhibitory effect of total saponins of Panax notoginseng on the apoptosis of neural cells in primary cultured rat hippocampal neurons under hypoxia/glucose reoxygenation. Methods: The percentage of apoptotic cells was detected by flow cytometry. The intracellular Ca2+ concentration was detected by confocal laser scanning microscopy. The morphological changes and percentage of necrotic cells were observed by fluorescence microscope. Meanwhile, the release of LDH was measured. Results: The percentage of apoptotic cells increased significantly after hypoxia/hypoglycemia for 5 h and 3 h after oxygenation (19.06%, P<0.01). The percentage of necrosis was 6.83% (P<0.01). The intracellular Ca2+ concentration was At 169.32nmol/L Ca2+ concentration (P<0.01), the release rate of LDH was 27.63% (P<0.01). The percentage of apoptosis was 49.85% when neurons were hypoxia/glucose deprived for 5 h and then given oxygen for 24 h (P<0.01). The percentage of cell necrosis was 11.49% (P<0.01), the intracellular Ca2+ concentration was 298.11 nmol/L (P<0.01), and the release rate of LDH was 60.35% (P<0.01). Panax notoginseng saponins (25,50 mg/L) can reduce the percentage of apoptosis and necrosis of nerve cells, decrease the concentration of intracellular Ca2+, and reduce the release of LDH. The effect of panax notoginseng saponins increases with increasing dose. Conclusion: Panax Notoginseng has an inhibitory effect on the apoptosis of hippocampal neurons after ischemia-reperfusion injury, which may be related to the decrease of intracellular Ca2+ concentration.