Fe3+和Fe2+对原代培养的成骨细胞增殖、分化和矿化功能的影响

采用噻唑蓝(MTT)法、碱性磷酸酶(ALP)比活性测定、油红O染色和茜素红染色及定量分析,研究了不同浓度的Fe3+和Fe2+对原代培养的成骨细胞增殖、分化及矿化功能的影响.结果表明:浓度为1×10-9～1×10-4 mol·L-1的Fe3+和Fe2+促进成骨细胞增殖,但是在较高浓度1×10-3 mol·L-1时,它们则抑制成骨细胞增殖.与成骨细胞作用48 h,浓度为1×10-8～1×10-4 mol·L-1的Fe3+和Fe2+抑制其分化,但在较低的浓度1×10-9 mol·L-1时则对其分化没有影响:进一步延长作用时间为72 h,Fe3+对成骨细胞分化没有影响,除1×10-6mol·L-1浓度的Fe2+促进成骨细胞分化外,其他浓度的Fe2+则抑制其分化;测试浓度下的Fe3+对成骨细胞向脂肪细胞的横向分化表现为抑制或没有影响,而Fe2+的影响则依赖于浓度和作用时间.在1×10-8～1×10-5mol·L-1浓度范围内,Fe3+和Fe2+对矿化结节的影响表现出相反的效应.在较高浓度(1×10-4mol·L-1)下,它们促进矿化节结的形成,而在较低浓度(1×10-9mol·L-1)下,Fe3+抑制矿化节结的形成,Fe2+则没有影响.结果提示:浓度.作用时间和铁离子的价态都是影响Fe3+和Fe2+生物效应(从毒性到活性,从损伤到保护,从上调到下调)转变的关键因素.

Effects of Fe3+ and Fe2+ on Proliferation, Differentiation and Mineralization Function of Primary Osteoblasts in vitro

The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromid(MTT), alkaline phosphatase(ALP) activity, oil red O assays and alizarin red-S(ARS) stain were used to evaluate the effects of Fe³⁺ and Fe²⁺ on proliferation, differentiation and mineralization function of primary osteoblasts(OBs) in vitro. The results indicate that both Fe³⁺ and Fe²⁺(1×10^-9~1×10^-4 mol·L^-1) promote the proliferation of OBs, but turn to inhibit at a higher concentration of 1×10^-3 mol·L^-1. Both Fe³⁺ and Fe²⁺(1×10^-8~1×10^-4 mol·L^-1) inhibit differentiation of OBs, but have no effect on differentiation at a lower concentration of 1×10^-9 mol·L^-1 for 48 h. Whereas, Fe³⁺ shows no effect on differentiation of OBs, Fe²⁺ begins to promote differentiation of OBs at concentration of 1×10^-6 mol·L^-1, but Fe²⁺ inhibits differentiation of OBs at other concentrations by prolonging the incubation time. Fe³⁺ inhibits adipocytic trans-differentiation of OBs or has no effects on adipocytic trans-differentiation of OBs at tested concentrations. The promotion or inhibition effect of Fe²⁺ on adipocytic trans-differentiation of OBs depends on concentration and incubation time. Fe³⁺ and Fe²⁺(1×10^-8~1×10^-5 mol·L^-1) inhibit and promote the formation of mineralized matrix nodules of OBs, respectively. Fe³⁺ and Fe²⁺ promote the formation of mineralized matrix nodules at a higher concentration of 1×10^-4 mol·L^-1, but Fe³⁺ inhibits the formation of mineralized matrix nodules, Fe²⁺ shows no effect at a lower concentration of 1×10^-9 mol·L^-1. The results suggest that concentration, culture time and valence state of iron ion are key factors for switching the biological effects of Fe³⁺ and Fe²⁺ from toxicity to activity, from damage to protection, or from down-regulation to up-regulation.