Sanchez-Ramos J R, Song S, Facca A, Basit A, Epstein C J
Miami V.A. GRECC, Florida, USA.
J Neurochem. 1997 Jan;68(1):58-67. doi: 10.1046/j.1471-4159.1997.68010058.x.
Primary dopaminergic neuronal cultures with increased superoxide dismutase (SOD) activity were established for studying the role of superoxide anion (O2-) in 1-methyl-4-phenylpyridinium (MPP+)-induced degeneration of dopamine (DA) neurons. Mean SOD activity in cultures prepared from transgenic (human) Cu/Zn SOD (hSOD1) mice was 2.46-2.60 times greater than in cultures prepared from nontransgenic control mice. After 1 and 2 weeks in culture, the mean density of DA neurons [number of tyrosine hydroxylase-immunoreactive (TH-ir) cells per visual field] was significantly higher in cultures prepared from transgenic mice compared with those prepared from nontransgenic control mice (4.55-5.63 TH-ir neurons per field in hSOD1 cultures vs. 2.66-2.8 TH-ir neurons per field in control cultures). However, uptake of [3H]DA relative to uptake of [3H]GABA was only slightly greater in hSOD1 cultures than in normal cultures (14.1 nmol of DA/100 nmol of GABA vs. 12.1 nmol of DA/100 nmol of GABA). Resistance to MPP+ toxicity was not significantly different from that in normal cultures when based on density of surviving TH-ir cell bodies (EC50 = 0.54 microM in hSOD1 and EC50 = 0.37 microM in normal cultures). A more sensitive measure of DA neuron integrity and function ([3H]DA uptake) also failed to demonstrate increased resistance of hSOD1 cultures to the toxin (EC50 = 73.7 nM in hSOD1 and EC50 = 86.2 nM in controls). These results do not support the hypothesis that neurotoxicity of the active metabolite of MPTP, MPP+, is mediated by generation of O2- in the cytoplasm. Nevertheless, mesencephalic cultures with increased hSOD1 activity appear to survive better than normal control cultures in the oxidatively stressful environment of cell culture incubators, and such mesencephalic cells may be useful for cell grafting studies in animal models of Parkinson's disease.
为了研究超氧阴离子（O2-）在1-甲基-4-苯基吡啶离子（MPP+）诱导的多巴胺（DA）神经元变性中的作用，建立了超氧化物歧化酶（SOD）活性增加的原代多巴胺能神经元培养物。从转基因（人）铜/锌超氧化物歧化酶（hSOD1）小鼠制备的培养物中的平均SOD活性比从非转基因对照小鼠制备的培养物中的平均SOD活性高2.46 - 2.60倍。培养1周和2周后，与从非转基因对照小鼠制备的培养物相比，从转基因小鼠制备的培养物中DA神经元的平均密度[每个视野中酪氨酸羟化酶免疫反应性（TH-ir）细胞的数量]显著更高（hSOD1培养物中每个视野有4.55 - 5.63个TH-ir神经元，而对照培养物中每个视野有2.66 - 2.8个TH-ir神经元）。然而，相对于[3H]GABA的摄取，[3H]DA在hSOD1培养物中的摄取仅比正常培养物略高（14.1 nmol DA/100 nmol GABA对12.1 nmol DA/100 nmol GABA）。基于存活的TH-ir细胞体密度，对MPP+毒性的抗性与正常培养物中的抗性没有显著差异（hSOD1中的半数有效浓度（EC50）= 0.54 microM，正常培养物中的EC50 = 0.37 microM）。对DA神经元完整性和功能的更敏感测量（[3H]DA摄取）也未能证明hSOD1培养物对毒素的抗性增加（hSOD1中的EC50 = 73.7 nM，对照中的EC50 = 86.2 nM）。这些结果不支持MPTP的活性代谢产物MPP+的神经毒性是由细胞质中O2-的产生介导的这一假设。然而，hSOD1活性增加的中脑培养物在细胞培养箱的氧化应激环境中似乎比正常对照培养物存活得更好，并且这种中脑细胞可能对帕金森病动物模型的细胞移植研究有用。
