目的 研究白细胞介素-17(IL-17)联合干扰素-γ(IFN-γ)在小鼠破骨前体细胞系RAW264.7分化为成熟破骨细胞过程中,对细胞增殖和凋亡的影响.方法 将核因子κB受体活化因子配体(RANKL)和巨噬细胞集落刺激因子(M-CSF)诱导小鼠破骨前体细胞系RAW264.7建立破骨细胞体外诱导分化研究模型,被诱导的RAW264.7细胞培养24 h后,分为IL-17组、IFN-γ组、IL-17+IFN-γ联合组(IL-17+IFN-γ等量组、IL-17固定+IFN-γ梯度组)进行干预.抗酒石酸酸性磷酸酶(TRAP)染色对破骨细胞成熟进行鉴定,利用CCK-8细胞增殖试验检测各组细胞增殖情况,AnnexinⅤ细胞凋亡实验评价各组细胞凋亡的差异,实时荧光定量PCR检测凋亡相关基因mRNA表达差异.结果 50 ng/m L IL-17与不同浓度的IFN-γ联合,随着IFN-γ浓度升高,IL-17对RAW264.7细胞生长抑制呈浓度依赖性.IL-17和IFN-γ的联合作用比单独应用IL-17,凋亡率更高;与单独应用IFN-γ相比,凋亡相关基因Fas L表达明显增加.结论 试在RAW264.7细胞向破骨细胞的分化过程中,IL-17和IFN-γ联合能抑制破骨前体RAW264.7细胞向破骨细胞的增殖,并促进RAW264.7细胞的凋亡.%Objective To investigate the effects of IL-17 combined with IFN-γduring the process of inducing RAW264.7 of mouseosteoclast precursor cell line into mature osteoclasts on the proliferation and apoptosis. Methods The differentiation study model of osteoclastin in vitro was established with 50 ng/m L RANKL and 30 ng/m L M-CSF inducing RAW264.7 of mouse osteoclast precursor cell line.After RAW264.7 cells were cultured for 24 hours, they were divided into three groups: the first group of IL-17, the second group of IFN-γ, and the third group of IL-17 combined with IFN-γ. The cultured osteoclasts were identified by the staining of tartrate-resistance acid phosphatase (TRAP).Viability was determined using cell counting kit-8. Apoptosis was measured with Annexin V cell apoptosis test.Furthermore, RT-PCR was used to examine the expression difference of mRNA. Results With 50 ng/m L IL-17 combined with different concentrations of IFN-γ, the growth of RAW264.7 was inhibited by the increase of IFN-γ in a concentration dependent manner.IFN-γ inhibited the proliferation of pre-osteoclast in adose-dependent manner (P＜ 0.05).The apoptosis rate of combined effect of IL-17 and IFN-γ was higher than that of IL-17 alone, and the expression of apoptosis-related gene Fas L was significantly higher in IFN-γwith IL-17 than IFN-γ only. Conclusion During the differentiation of RAW264.7 into osteoclasts, IL-17 combined with IFN-γ can inhibit the proliferation of osteoclast precursor cell and promote the apoptosis of RAW264.7.
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