抑制葡糖脑苷脂酶对多巴胺神经细胞内α-突触核蛋白寡聚体形成及细胞自噬功能的影响

Effect of glucocerebrosidase inhibition on intracellular α-synuclein oligomer formation and autophagic activity of dopaminergic cells

目的研究抑制葡糖脑苷脂酶(glucocerebrosidase,GBA)活性对多巴胺神经细胞内α-突触核蛋白(α-synuclein,α-Syn)寡聚体形成及自噬功能的影响。方法在培养MES23.5细胞外添加不同浓度的GBA活性抑制剂(conduritol-β-epoxide,CβE),观察GBA活性的变化,同时在细胞外添加α-Syn单体使其进入细胞内并孵育48 h。使用双抗体夹心ELISA方法测定细胞内α-Syn寡聚体含量,观察细胞自噬活性、氧化应激和凋亡。结果随着CβE质量浓度的增加,GBA活性呈质量浓度依赖性的下降,而细胞内α-Syn寡聚体的含量则呈质量浓度依赖性的增加;在CβE处理细胞,随着细胞外添加的α-Syn单体的质量浓度加大,自噬活性和氧化应激液增强,同时细胞凋亡数量增加。结论抑制GBA活性导致细胞内α-Syn寡聚体增多以及依赖于α-Syn浓度的自噬活性和氧化应激增强和细胞凋亡增加。

Objective To investigate the effect of glucocerebrosidase （GBA） inhibition on intracellular α-synuclein（α-Syn） oligomer formation and autophagic activity of dopaminergic ceils. Methods The activity of GBA was measured in MES 23.5 cells treated with different concentrations of CβE, the specific GBA inhibitor. After treatment with CβE, monomeric β-Syn was added into the medium and the amount of intracellular α-Syn oligomers were evaluated by ELISA. The autophagic activity, oxidative stress, and apoptotic cell counts were also examined. Results CβE dose-dependently decreased the activity of GBA and increased the levels of intracellular α-Syn oligomers. CβE also enhanced the autophagic activity, oxidative stress levels and apoptotic cell counts in an α-Syn concentration- dependent manner. Conclusion GBA inhibition causes intracellular accumulation of α-Syn oiigomers and α-Syn concentration-dependent enhancement of autophagy, oxidative stress and apoptosis. [ Key words] glucocerebrosidase; α-synuclein oligomer; MES 23.5 cells; autophagy