α-突触核蛋白寡聚体经氧化应激途径致帕金森病小鼠模型多巴胺能神经元损伤

Alpha-Synuclein Oligomers Induce Loss of Dopaminergic Neurons via Oxidative Stress in Parkinson's Disease Mouse Model

目的:通过鱼藤酮持续灌胃构建的帕金森病小鼠模型研究α-突触核蛋白寡聚体(α-Syn)的毒性作用机制。方法:48只老年雄性C57小鼠随机分为鱼藤酮组和对照组,每组24只,鱼藤酮组灌注0.01 mL/g鱼藤酮氯仿溶液,对照组灌注0.01 m L/g氯仿溶液,均连续灌胃12周。于灌胃后取脑,行蛋白质印迹实验检测α-Syn表达水平、透射电镜观察中脑黑质神经元超微结构;于灌胃前、后取脑,免疫组化染色观察黑质部位酪氨酸羟化酶(TH)阳性细胞密度,并检测脑组织中超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)和丙二醛(MDA)水平。结果:灌胃12周后,Western blot显示鱼藤酮组中脑α-Syn的表达较对照组明显增多(P<0.05)。免疫组化染色显示鱼藤酮组中TH阳性细胞数量较对照组明显减少(P<0.05)。透射电镜显示鱼藤酮组小鼠黑质部可见线粒体、高尔基体不规则肿胀。鱼藤酮组SOD、GSH-PX水平较对照组明显下降(P<0.05),MDA水平较对照组显著增多(P<0.05)。结论:鱼藤酮小剂量持续灌胃可在脑内诱发形成α-Syn寡聚体,α-Syn寡聚体可能经氧化应激途径导致多巴胺神经元损伤。

Objective: To explore the mechanisms of the impairment of dopaminergic neurons by alpha-synuclein (α-Syn) oligomers in the Parkinson’s disease (PD) mouse model established by gavage with constant low-dose rotenone. Methods: A total of 48 elderly male C57 mice were randomly divided into the rotenone group and control group with 24 mice per group. Mice in the rotenone group were administered 0.01 mL/g rotenone by gavage while mice in the control group were administered 0.01 mL/g chloroform. Mice were sacrificed after 12 weeks of continuous treatment. In mice brains harvested after treatment, western blotting and immunohistochemical staining were carried out to detect α-Syn expression levels;electron microscopy was used to observe the ultrastructure of substantia nigra neurons in the midbrain. In mice brains harvested before and after treatment, immunohistochemical staining of tyrosine hydroxylase (TH) was performed to examine the density of TH-positive neurons in the substantia nigra, and levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and malondialdehyde (MDA) in the brain tissue were determined. Results: After the 12-week treatment, western blotting demonstrated that α-Syn expression in the midbrain of the rotenone group mice was significantly higher than that of the control group mice (P<0.05). Immunohistochemistry showed that there was a significant decrease in TH-positive neurons in the midbrain of the rotenone group mice compared with that of the control group mice (P<0.05). Electron microscopy showed swollen mitochondria and Golgi in the substantia nigra of the rotenone group mice. Compared to the control group, the rotenone group showed significantly decreased levels of SOD and GSH-PX (P<0.05) and significantly increased levels of MDA (P< 0.05). Conclusion:α-Syn oligomers can form in the brain after continuous intragastric administration of low-dose rotenone.α-Syn oligomers in the midbrain might impair dopaminergic neurons via oxidative stress.