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目的观察低氧和(或)高糖对小鼠神经母细胞瘤细胞Neuro-2a活性的影响,并探讨其自噬相关机制。方法 Neuro-2a细胞分为4组:正常对照组、高糖组、低氧组和高糖低氧组。使用MTS方法检测细胞活性;透射电镜、免疫印记、免疫荧光染色检测各组细胞自噬水平。结果相较于正常对照组,低氧组、高糖低氧组的Neuro-2a细胞活性明显下降(P<0.01),胞质内LC3B蛋白明显聚集;细胞内自噬囊泡样物质增加;自噬相关蛋白LC3B-Ⅱ、P62蛋白水平均升高(P<0.05)。自噬抑制剂(bafilomycin A1)处理后,正常对照组、高糖组Neuro-2a细胞LC3B-Ⅱ、P62蛋白水平升高(P<0.05),活性显著下降(P<0.01)。结论低氧可以降低Neuro-2a细胞活性,可能是通过抑制自噬降解途径发挥作用。
Objective To investigate the effect of hypoxia and / or high glucose on Neuro-2a activity of neuroblastoma cells in mice and to explore the mechanism of autophagy. Methods Neuro-2a cells were divided into 4 groups: normal control group, high glucose group, hypoxia group and high glucose hypoxia group. The cell viability was detected by MTS method. The autophagy of each group was detected by transmission electron microscopy, immunoblot and immunofluorescence staining. Results Compared with the normal control group, the activity of Neuro-2a cells in hypoxia group and high glucose hypoxia group was significantly decreased (P <0.01), and the LC3B protein in the cytoplasm was significantly aggregated; the content of autophagic vesicle-like substance increased The protein levels of LC3B-Ⅱand P62 were all increased (P <0.05). After bafilomycin A1 treatment, the levels of LC3B-Ⅱ and P62 protein in Neuro-2a cells in normal control group and high glucose group were significantly increased (P <0.05), and the activity was significantly decreased (P <0.01). Conclusion Hypoxia can reduce the activity of Neuro-2a cells, which may be through the inhibition of autophagy degradation pathway.