白细胞介素-7对巨噬细胞分化形成破骨细胞的影响

Effect of Interleukin-7 on the Differentiation of Macrophages into Osteoclasts

目的:白细胞介素-7(IL-7)在类风湿关节炎骨破坏过程中具有重要作用,然而IL-7的具体作用方式不明确,因此本研究分析了IL-7对破骨细胞分化形成的作用及相关分子机制。方法:用M-CSF,M-CSF+RANKL及IL-7分别诱导RAW264.7巨噬细胞分化形成破骨细胞,通过抗酒石酸酸性磷酸酶(TRAP)染色评价IL-7对破骨细胞分化形成的影响。然后使用JAK信号通路抑制剂(AG490),AKT信号通路抑制剂(LY294002)和JNK信号通路抑制剂(SP600125),采用RT-PCR和ELISA分别检测IL-7对破骨细胞骨溶性相关活性基因和蛋白分泌的影响。最后使用蛋白印迹检测探讨IL-7促进破骨细胞分化形成的相关分子机制。结果:IL-7在核因子κB配体(RANKL)不存在的情况下能够显著增加破骨细胞的数量。IL-7能够显著诱导破骨细胞溶骨相关活性成分包括基质金属蛋白酶-9(MMP9)、组织蛋白酶(CathK)、抗酒石酸酸性磷酸酶(TRAP)、降钙素受体(CTR)的基因表达和蛋白分泌,JAK(AG490)和JNK(SP600125)信号通路抑制剂能显著抑制IL-7诱导的上述基因表达和蛋白分泌。IL-7显著诱导JAK,AKT及JNK信号通路信号蛋白的活化。结论:IL-7能不依赖于RANKL诱导破骨细胞的分化形成,其作用可能是通过活化JAK和JNK信号通路实现的。

Objective:Interleukin-7(IL-7) plays an important role in the process of bone destruction in rheumatoid arthritis.However,the specific mode of action of IL-7 is not clear.Therefore,this study analyzed the effect of IL-7 on osteoclastogenesis and its related molecular mechanism.Methods:M-CSF,M-CSF+RANKL and IL-7 were used to induce RAW264.7 cells to differentiate into osteoclasts,respectively.The effect of IL-7 on osteoclastogenesis was evaluated by tartrate-resistant acid phosphatase(TRAP) staining.Then,JAK signaling pathway inhibitor(AG490),AKT signaling pathway inhibitor(LY294002) and JNK signaling pathway inhibitor(SP600125) were used to evaluate the effects of IL-7 on bone solubility related active genes and protein secretion of osteoclasts by RT-PCR and ELISA,respectively.Western blot was used to explore the molecular mechanism of IL-7 promoting osteoclastogenesis.Results: IL-7 could significantly increase the number of osteoclasts in the absence of nuclear factor κB ligand(RANKL).IL-7 could significantly induce the gene expression and protein secretion of osteoclast osteolytic related components,including matrix metalloproteinase-9(MMP9),cathepsin(CathK),tartrate-resistant acid phosphatase(TRAP) and calcitonin receptor(CTR).JAK(AG490) and JNK(SP600125) signaling pathway inhibitors could significantly inhibit IL-7-induced gene expression and protein secretion.IL-7 significantly induced the activation of JAK,AKT and JNK signaling proteins.Conclusion:IL-7 can induce osteoclastogenesis independent of RANKL,and its effect may be achieved by activating JAK and JNK signaling pathways.