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目的:建立MPP+(1-甲基-4-苯基吡啶离子)干预SAMP8(快速老化小鼠P8)小鼠中脑神经元的体外细胞模型。方法:SAMP8新生1 d小鼠的中脑神经元原代混合培养6 d,加入100μmol/L浓度的MPP+,再培养6、9、12 h和24 h后分别对各时间点的中脑原代培养神经元免疫荧光染色或提取蛋白测定TH水平。结果:MPP+导致原代培养的SAMP8小鼠中脑神经元形态学改变。正常对照组神经元形态完整,免疫反应性强,胞体大呈椭圆形,突起多而长且粗壮;MPP+组神经元随时间逐渐出现形态变化,MPP+后6 h即可见突起不完整断续,9 h突起数目减少,缩短;12 h神经元胞体明显变小,24 h突起多消失,神经元胞体小且免疫反应性弱。MPP+导致原代培养的SAMP8小鼠中脑神经元数量在MPP+后9 h开始有显著减少,同时TH蛋白的表达也开始有显著减少,24 h最低。结论:MPP+对原代培养的SAMP8小鼠中脑神经元具有毒性作用。MPP+导致SAMP8小鼠中脑神经元原代混合培养体系的神经元数量下降,同时TH蛋白表达降低,提示MPP+导致其中脑DA能神经元损伤死亡。
OBJECTIVE: To establish an in vitro cell model of MPP + (1-methyl-4-phenylpyridinium) intervening neurons in SAMP8 (fast-aging mouse P8) mice. Methods: Primary cultured neurons from SAMP8 neonatal mice were primarily cultured for 6 days. Six days after adding MPP + at a concentration of 100 μmol / L, Neuronal immunofluorescence staining or protein extraction was used to determine TH levels. Results: MPP + resulted in morphological changes of midbrain neurons in primary cultured SAMP8 mice. Neurons in the normal control group showed complete morphology, strong immunoreactivity, large oval cell body, long and thick protuberances, morphological changes of neurons in MPP + group over time, incomplete projection of neurons in 6 h after MPP +, 9 h neurites decreased, shortened; 12 h neuronal soma significantly reduced 24 h suddenly disappeared, neuronal cell body is small and immune reactivity weak. MPP + induced a significant decrease in the number of neurons in primary cultured SAMP8 mice at 9 h after MPP +, and at the same time, the expression of TH protein began to decrease significantly, reaching the lowest level at 24 h. CONCLUSION: MPP + is toxic to primary cultured midbrain neurons in SAMP8 mice. MPP + led to a decrease in the number of neurons in the primary mixed culture system of SAMP8 mice and a decrease in TH protein expression, suggesting that MPP + causes death of neuronal damage in midbrain DA.