氟对软骨细胞与成骨细胞凋亡及H_2O_2、SOD、NO的影响

Effects of fluoride on level of H_2O_2, SOD, NO and apoptosis of the chondroblasts and osteoblasts in vitro

目的研究氟对体外培养的软骨细胞、成骨细胞凋亡及H2O2、SOD、NO的影响。方法采用流式细胞仪对细胞周期进行分析及测定凋亡细胞的百分比;化学方法检测细胞培养液中的H2O2、SOD、NO;透射电镜观察软骨细胞超微结构。结果软骨细胞:(1)不同剂量的氟均诱导细胞凋亡,培养基中的H2O2、NO均水平高于对照组,加SOD可使H2O2、细胞凋亡百分率明显下降,NaF20、160μmol/L可使G2/M期细胞升高。(2)NaF320μmol/L可使细胞超微结构损伤,加SOD可拮抗氟的损伤作用。成骨细胞:与对照组相比(1)加入不同剂量NaF,培养基中的SOD活性降低。但仅在加入160、240μmol/L,培养基中H2O2升高,NO亦同时升高。(2)NaF400μmol/L可诱导成骨细胞凋亡,同时使G0/G1期细胞增多,240、400μmol/L可使G2/M期细胞减少。结论(1)低、中剂量的氟可促进软骨细胞、成骨细胞凋亡,损害细胞超微结构。与此同时培养物中H2O2升高,SOD活性降低,表明细胞凋亡及损伤与氧化应激有关。此外,NO也升高,可能也起一定作用。氟对细胞周期的影响:对软骨细胞主要作用于G2/M期,对成骨细胞可同时作用于G0/G1及G2/M期;(2)一定量的SOD可明显拮抗氟促进的H2O2释放及细胞凋亡,对细胞有保护作用。(3)一定剂量的氟促使软骨细胞、成骨细胞凋亡,同时也促进成骨细胞增殖。

Objective To investigate the effects of fluoride on the content of H2O2, SOD and NO as well as apoptosis of cultured chondroblasts and osteoblast. Methods For condroblasts, the cell cycle and apoptosis were inspected by flow cytometry, the contents of H2O2 and NO in the culture media were assayed by chemical methods, the ultrastructure was observed by the transmission electron microscope. For osteoblasts, only the cell cycle, cell apoptosis and the H2O2 and NO contents were studied by the same methods. Results Chondroblasts:(1) Apoptosis was obviously induced by fluoride at various concentrations, meanwhile the contents of H2O2 increased. These changes were not found when SOD was added. Increase of NO was also observed under the influence of fluoride. (2) The cells in G2 / M stage increased at 20 μmol/L and 160 μmol/L NaF. (3) Ultrastructural lesions were observed at 320 μmol/L NaF, but no long found while SOD was added. Osteoblasts: (1)H2O2 remarkably increased at 160 umol/L, 240 μmol/L NaF while SOD lowered and NO raised in every concentration. (2) At 400 μmol/L, apoptosis and the cells in the G0 / G1 stage increased while at 240 μmol/L and 400 μmol/L, the cells in G2 / M stage decreased. Conclusions (1) Fluoride in low and medial concentrations causes increase of H2O2 and decrease of SOD, inducing apoptosis and damage of the cell ultrastructures. (2) Fluoride affectes mainly the chondroblasts in G2 / M stage and the osteoblasts in G0 / G1 stage and G2 / M stage. (3) SOD can antagonize the fluoride in the aspect of release of H2O2 and induction of apoptosis thus protecting the cells from damage. (4) It seems that excessive products of apoptosis induce cell proliferation and sclerotic changes during fluorosis. Increased NO might play a role too.