高铁与低铁环境对成骨细胞hFOB1.19膜铁转运蛋白1表达的影响

Expression of ferroportin 1 in cultured osteoblast hFOB1.19 under iron excess and deficiency

目的观察细胞培养液中加入铁离子或铁离子螯合剂后成骨细胞(hFOB1.19)膜铁转运蛋白1(FPN1)表达的变化,探讨铁离子浓度对成骨细胞FPN1表达的影响。方法将不同浓度的枸橼酸铁铵(FAC,50、100、200μmol/L)和去铁胺(DFO,5、10、20μmol/L)分别加入人hFOB1.19培养基,24h后用共聚焦显微镜(CLSM)测定细胞内铁离子浓度,用定量PCR、Westernblotting和免疫荧光法测定不同组别成骨细胞FPN1的表达并进行统计比较。结果用不同浓度的FAC和DFO干预20h后,成骨细胞内铁离子浓度随FAC浓度增加而增加,随DFO浓度增加而降低,组间差异有统计学意义(P<0.05);FPN1基因和蛋白表达随FAC浓度增加而增加,随DFO浓度增加而降低,组间差异有统计学意义(P<0.05)。结论高铁环境可上调FPN1表达,低铁环境可下调FPN1表达。FPN1表达量的改变可有效维持成骨细胞内铁离子浓度的平衡。

Objective To investigate the effects of iron excess and iron deficiency on the expression of ferroportin 1 （FPN1） in human osteoblast. Methods Human osteoblast cells （hFOB1. 19） were treated with ferric ammonium citrate （FAC） or deferoxamine （DFO） of different concentrations. The intracellular iron ion was measured by a confocal laser scanning microscope （ CLSM ）. The expression of FPN1 at mRNA and protein levels was detected with real-time quantitative polymerase chain reaction （PCR） , western blotting and immunofluoreseenee. Results The confocal microcopy measurements showed that the fluorescence intensity representing intracellular iron concentration was significantly weakened with increasing of FAC concentration and enhanced with increasing of DFO （ P 〈 0. 05 ）. The real-time PCR, western blotting and immunofluoreseence indicated that the mRNA and protein expression of FPN1 in osteoblast was increased with increasing of FAC in a concentration-dependent manner and decreased with increasing of DFO in a concentration-dependent manner （P 〈 0. 05 ）. Conclusion Increasing of intracellular iron ion by the FAC treatment increased the FPN1 expression. In contrast, decreasing intracellular iron ion by the DFO treatment decreased the FNP1 expression. The changes of FPN1 expression may help to maintain the balance of intracelullar iron in osteoblasts.