铁离子对人成骨细胞RANKL/OPG基因及蛋白表达的影响

Effects of ferric ammonium citrate on the RANKL/OPG gene and protein expression ofhuman osteoblasts

目的研究人成骨细胞(hFOB1.19)在不同铁离子状态下RANKL/OPG基因及蛋白的表达。方法成骨细胞株(hFOB1.19)在DMEM/F-12培养基培养传代至第3代后,用不同终浓度枸橼酸铁铵(50、100、200μmol/L)加入细胞培养基中干预24h,用RT-PCR方法和免疫印迹法(WesternBlot)检测干预后成骨细胞的RANKL、OPGmRNA和蛋白表达并计算RANKL/OPG比率。结果RT-PCR检测结果显示,对照组、50、100、200μmol/L组RANKL/OPGmRNA表达比分别为0.56±0.13、0.58±0.01、0.69±0.01、1.84±0.92;Westernblot结果显示,对照组、50、100、200μmol/L组RANKL/OPG蛋白表达比分别为0.82±0.66、0.82±0.64、1.09±0.11、1.25±0.14。统计学分析显示,在mRNA和蛋白水平,100和200μmol/L浓度的枸橼酸铁铵干预后RANKL/OPG比值明显高于对照组(P<0.05),50μmol/L枸橼酸铁干预后与对照组差异无统计学意义。结论不同浓度枸橼酸铁铵可以影响人成骨细胞RANKL/OPG基因及蛋白的表达,进而可能影响骨形成和骨吸收。

Objective To investigate the effects of Ferric Ammonium Citrate （FAC） on gene and protein ex- pression of RANKL and OPG of human osteoblasts. Methods After human osteoblasts were cultured in the medium, cells were supplemented with FAC at the final concentration of O, 50, 100, and 200 μmol/L. After treatment with FAC for 24h, the mRNA and protein expression of RANKL and OPG both secreted by osteobiasts were detected by reverse transcription polymerase chain reaction （RT-PCR） and Western blot respectively, and calculated the RANKL/OPG ratio afterwards. Results The mRNA expression of RANKL/OPG ratio in human osteoblast hFOB1.19 cells of control, 50, 100, 200 Ixmol/L groups were （0.56 ±0. 13）, （0.58 ±0.01）, （0. 69 ±0.01）, （1.84±0.92） respectively; the protein expression of RANKL/OPG ratio in human osteoblast hFOB1.19 cells of control, 50, 100, 200 μmol/L groups were （0. 82 ± 0. 66）, （0. 82 ± 0. 64）, （ 1.09± 0. 11 ）, （ 1.25± 0. 14） respectively. Statistical analysis revealed that FAC treatment at 100 μmol/L and 200 μmol/L significantly increased the expression of RANKL/OPG compared to control group; whereas FAC treatment at 50 Iμmol/L did not significantly alter the mRNA and protein expression of RANKE/ OPG. Conclusion These results indicated that different concentration of FAC affected the expression of RANKL/OPG in human osteoblasts, which might further change bone formation and bone resorption.