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目的:分析一个ALS家系突变铜、锌超氧化物歧化酶(Cu/Zn superoxide dismutase,SOD1)基因在原代培养的大鼠皮层神经元中的定位及作用。方法:将构建好的含有SOD1正常基因与突变基因在内的增强型绿色荧光蛋白真核表达载体(pEGFP-hSOD1和pEGFP-mSOD1)分别转染大鼠皮层神经元,转染后观察绿色荧光在细胞内的分布情况,同时设对照组,检测神经元的SOD1活性、MDA含量及细胞活性。结果:转染后荧光物质表达于胞浆内,与转染pEGFP-mSOD1质粒的神经元相比,转染pEGFP-hSOD1质粒神经元的SOD1活性、MTT值显著增高,而MDA、LDH水平显著降低。结论:突变SOD1蛋白的酶活性下降,使脂质过氧化物相对增多,导致神经元损伤,可能是该家系发病的主要原因之一。
OBJECTIVE: To analyze the localization and role of Cu / Zn superoxide dismutase (SOD1) gene in primary cultured rat cortical neurons from one ALS family. Methods: Eukaryotic expression vectors (pEGFP-hSOD1 and pEGFP-mSOD1) containing enhanced green fluorescent protein (SOD1) gene and mutant genes were transfected into rat cortical neurons. After transfection, green fluorescent protein Intracellular distribution of the same time set the control group, detection of neurons SOD1 activity, MDA content and cell activity. Results: Compared with the neurons transfected with pEGFP-mSOD1 plasmid, the transfection of pEGFP-hSOD1 plasmid neurons showed a significant increase in MTT and MDA, LDH levels . CONCLUSION: The decrease of SOD1 protein enzyme activity leads to the relative increase of lipid peroxides, resulting in neuronal damage, which may be one of the main causes of the onset of this pedigree.