氟对人神经母细胞瘤SH-SY5Y细胞自噬水平的影响

Effects of fluoride on autophagy level in human neuroblastoma SH-SY5Y cells

目的探讨氟对人神经母细胞瘤SH-SY5Y细胞自噬水平的影响。方法用不同浓度(20、40、60 mg/L)的氟化钠(Na F)染毒SH-SY5Y细胞24 h,采用透射电子显微镜观察细胞内自噬体超微结构;吖啶橙染色观察细胞内自噬囊泡的生成;Western blotting技术检测自噬延伸相关蛋白Atg5和LC3-II与自噬降解相关蛋白P62的表达水平。结果透射电子显微镜下可观察到细胞内自噬体结构,且随NaF剂量的增加自噬体数量明显减少;吖啶橙染色可见,与对照组相比,40和60 mg/L NaF染毒组自噬囊泡数量逐渐减少,尤以60 mg/L NaF染毒组减少最为明显;Western blotting检测发现,与对照组相比,Atg5和LC3-II的表达水平呈剂量依赖性降低,而P62的表达水平呈剂量依赖性升高,且差异具有统计学意义(P<0.05)。结论氟可抑制SH-SY5Y细胞的自噬水平。

Objective To explore the effects of fluoride on autophagy level in human neuroblastoma SH-SY5Y cells. Methods SH-SY5Y cells were treated with different concentrations（ 20,40,60 mg/L） of fluoride for 24 h. Transmission electron microscope（ TEM） was used to detect the ultrastructure of autophagosomes in SH-SY5Y cells.Acridine orange staining was used to examine the form of autophagic vesicles and Western blotting was used to check the expression levels of autophagy relevant proteins of Atg5,LC3 and P62 in SH-SY5 Y cells. Results The TEM results showed the presence of autophagic ultrastructure in SH-SY5Y cells and fluoride could decrease the amount of autophagosomes. Acridine orange staining results showed that the amount of autophagic vesicles were obviously decreased in 40 and 60 mg/L Na F-treated groups compared with control,and 60 mg/L Na F-treated group was decreased the most. The expression level of Atg5 and LC3-II protein was dose-dependently decrease and P62 protein was dosedependently increased compared with control（ P〈0. 05）. Conclusion Fluoride maysuppress the level of autophagy in SH-SY5Y cells.