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目的 探讨一氧化氮 (NO)在缺氧复氧诱导神经细胞凋亡中的作用及中药银杏叶提取物的保护机制。方法 实验使用胎龄 16～ 17日Wistar大鼠的大脑皮层神经细胞进行原代分离培养 ;采用Wright Giemsa染色 ,光镜、透射电子显微镜观察 ;原位末端标记法确立缺氧复氧神经细胞凋亡病理模型 ;应用NADPH d组织化学方法检测神经细胞一氧化氮合酶 (NOS)的表达并用计算机图像分析系统进行定量检测。 结果 缺氧复氧可以使大鼠大脑皮层神经细胞发生凋亡 ,随缺氧时间的延长 ,凋亡细胞数渐多 ,至缺氧 8h复氧 18h达高峰 ;在缺氧 2h(H2 R0 组 )和缺氧 8h复氧 18h(R8R1 8)组中神经细胞NOS表达均显著增高 ,与正常对照组比有显著性差异 (P <0 0 1;P <0 0 5 )。EGB能显著抑制此双时相NOS活性的增强 ,并明显降低神经细胞凋亡率。 结论 缺氧复氧损伤可诱导培养的大鼠大脑皮层神经细胞发生凋亡。NOS表达增强从而使NO产生增加可能是缺氧复氧诱导神经细胞凋亡的机制之一。银杏叶提取物 (EGB)经下调NOS表达活性 ,抑制NO的产生保护培养的大鼠大脑皮层神经细胞免于凋亡。
Objective To investigate the role of nitric oxide (NO) in the apoptosis induced by hypoxia-reoxygenation and the protective mechanism of Ginkgo biloba extract. Methods Wistar rats were cultured and subcultured in primary cortex of Wistar rats with gestational age of 16-17 days. Wright Giemsa staining, light microscopy and transmission electron microscopy were used to observe the apoptosis of hypoxic-reoxygenated nerve cells Pathological model; NADPH d histochemical method was used to detect the expression of nitric oxide synthase (NOS) in nerve cells and quantified by computer image analysis system. Results Hypoxia-reoxygenation could induce neuronal apoptosis in rat cerebral cortex. With the prolongation of hypoxia, the number of apoptotic cells gradually increased to the peak at 18 hours of hypoxia and reached the peak at 18 hours of hypoxia. At hypoxia 2 hours (H2R0 group) (P <0 01; P 0 05) .Conclusion: The expression of NOS in nerve cells of rats with hypoxia for 8 h and reoxygenation 18 h (R8R1 8) is significantly higher than that of normal control group (P <0.01). EGB could significantly inhibit the increase of NOS activity in this two-phase phase and significantly reduce the rate of neuronal apoptosis. Conclusion Hypoxia-reoxygenation injury induces apoptosis of cultured rat cerebral cortical neurons. One of the mechanisms of hypoxia-reoxygenation-induced neuronal apoptosis is the increase of NOS expression and NO production. Ginkgo biloba extract (EGB) down-regulated NOS expression and inhibited the production of NO in cultured rat cerebral cortical neurons from apoptosis.