低氧条件下铁调节蛋白1在维持SH-SY5Y细胞铁稳态中的作用

Involvement of iron regulatory protein 1 in iron homeostasis under hypoxia in human neuroblastoma( SH-SY5Y) cells

目的：研究低氧条件下铁调节蛋白1（iron regulatory protein 1,IRP1）对人神经母细胞瘤SH-SY5Y细胞铁稳态的影响。方法：1%O2处理SH-SY5Y细胞0,1,2,4,6 h,蛋白质印迹法检测铁蛋白轻链（ferritin light chain,Ft-L）以及膜铁输出蛋白（ferroportin,Fpn）的表达;Calcein-AM法检测SH-SY5Y细胞对Fe2＋的吸收以及细胞可变铁池（labile iron pool,LIP）;采用硫氰酸盐分光光度法测定细胞总含铁量;特异性siRNA下调IRP1后检测SH-SY5Y细胞低氧2 h铁稳态的变化。结果：低氧0~6 h,SH-SY5Y细胞Ft-L和Fpn表达增加,Fe2＋摄入和LIP增加,但细胞总铁水平不变。siRNA下调IRP1表达后再低氧2 h,Ft-L蛋白表达和LIP明显增加,Fpn表达和Fe2＋摄入没有明显变化,细胞总铁水平增加。结论：低氧条件下,IRP1是SH-SY5Y细胞铁稳态的重要调节者。

Objective ： To investigate the effects of iron regulatmy protein 1 （ IRPI ） on iron homeostasis in human neuroblastoma SH-SY5Y cells under hypoxia. Methods： The SH-SY5Y cells were exposed to hy- poxia for 0, 1 , 2, 4, 6 h （ 1% 02） , and then the total proteins were extracted to detect the expression of ferritin light chain （Ft-L） and ferroportin（Fpn） by Western blotting. Calcein-AM was used to measure ferrous iron uptake and labile iron pool（LIP） in SH-SY5Y cells. SpectrophotometIy was used to determine the total cellular iron level with thiocyanate. The effects of IRPI siRNA on iron homeostasis in SH-SY5Y cells under hypoxia were detected. Results： Iron homeostasis remained unchanged when cells were exposed to 1% hypoxia for 0 - 6 h although the expression of Ft-L and Fpn increased, and the level of ferrous iron uptake and LIP elevated. When IRP1 was knocked down using IRPl-specific siRNA, the expression of Ft-L and LIP increased, while Fpn and ferrous iron uptake did not altered obviously. The iron homeostasis in SH-SY5Y cells altered under 2 h hypoxia in terms of the cellular total iron content increased. Conclusion： IRPI regulated iron homeostasis in SH-SY5Y cells under hypoxia.