低氧模拟物去铁胺对大鼠纤维环细胞增殖和细胞周期的影响

Influence of desferoxamine-activated hypoxia inducible factor-1α on the proliferation and cell cycle of rat annulus fibrosus cell

[目的]研究不同浓度去铁胺对大鼠纤维环细胞增殖及细胞周期的影响。[方法]体外分离、培养大鼠纤维环细胞,应用CCK-8细胞活力检测试剂盒、流式细胞仪检测不同浓度的去铁胺（25μmol/L、50μmol/L、100μmol/L、200μmol/L）对细胞增殖及细胞周期的影响;RT-PCR和Western-blot检测25μmol/L DFO干预纤维环细胞后低氧诱导因子1α表达水平变化。[结果]CCK-8结果显示：与对照组相比,25μmol/L DFO干预纤维环细胞时,OD值明显升高,两组比较差异有统计学意义（P〈0.05）;100μmol/L、200μmol/L DFO干预时,OD值明显降低,两两比较差异有统计学意义（P〈0.05）;流式细胞仪结果显示：25μmol/L DFO干预时,处于S/G2期细胞所占比例增加,与对照组相比差异有统计学意义（P〈0.05）;而100μmol/L、200μmol/L DFO干预时,S/G2期细胞所占比例减少,与对照组相比差异具有统计学意义（P〈0.05）;低浓度DFO干预纤维环细胞8~24 h后,低氧诱导因子1αmRNA和蛋白质水平都有明显上升。[结论]低浓度DFO可以促进体外培养纤维环细胞的增殖,HIF-1α可能参与了这一过程,DFO可能作为一种治疗药物用于延缓椎间盘退变的过程。

[Objective] To observe the influence of different concentrations of desferoxamine（ DFO） on the proliferation and cell cycle of rat annulus fibrosus cells（ AFCs）. [Methods] Rat AFCs were harvested and cultured in vitro. Then,theAFCs were cultured with different concentrations of DFO（ 25 μM,50 μM,100 μM,and 200 μM） for 8 ~ 24 hours; the proliferation and cell cycle of the AFCs were examined using the cell counting kit-8（ CCK-8） assay and flow cytometry. The expression level of hypoxia inducible factor-1α（ HIF-1α） wastested using semi-quantitative real-time polymerase chain reaction（ RT-PCR） and western-blot. [Results] The CCK-8assay and flow cytometry indicated that the viability and percentage of S /G2 phase cells of AFCs cultured with 25 μM DFO increased when compared to those in the control group,while the viability and percentage decreased with 100 μM and 200 μM DFO. When AFCs were cultured with 25 μM DFO for 8 ~ 24 hours,the expression of HIF-1α increased at both mRNA and protein levels compared to that in the control group. [Conclusion] Moderate concentrations of DFO can promote the proliferation process of AFCs,and HIF-1α may play a role in this process. DFO may become a therapeutic drug for the restoration of cell numbers during degenerative disc disease.