目的：探讨Notch信号抑制对核因子κB受体活化因子配体（RANKL）诱导的小鼠RAW264.7细胞向破骨细胞分化的影响。方法建立RANKL诱导的小鼠RAW264.7细胞体外分化模型，实时定量聚合酶链反应（real-time PCR）检测Notch信号成分（Notch1、Notch2、Delta1、Jagged1）、下游靶基因Hes1以及破骨细胞标志基因抗酒石酸酸性磷酸酶（TRAP）和Cathepsin K在诱导前后mRNA的表达。在诱导体系中加入不同浓度的γ分泌酶抑制剂（GSI），抑制Notch受体的表达，TRAP染色检测破骨细胞分化的变化情况。结果50 ng·mL-1 RANKL诱导小鼠RAW264.7细胞3 d，Notch1、Notch2、Delta1、Jagged1及Hes1的mRNA表达均有不同程度的提高，其中以Notch2、Jagged1增高最明显；破骨细胞标志基因表达显著增高。在RANKL诱导的同时加入不同浓度GSI，抑制Notch的表达，可致Notch下游靶基因Hes1表达下降，同时TRAP阳性细胞计数显著减少，且呈剂量依赖性。结论 Notch信号可促进RANKL诱导的RAW264.7细胞向破骨细胞分化。%Objective This study aims to explore the effect of Notch signaling depression on the receptor activator of nuclear factor kappa B ligand (RANKL)-induced osteoclastogenesis of RAW264.7 cells. Methods Mice RAW264.7 cells were cultured and differentiated into osteoclasts with the induction of RANKL. The expressions of Notch1, Notch2, Delta1, Jagged1, Hes1, tartrate-resistant acid phosphatase (TRAP), and Cathepsin K genes during osteoclastogenesis were analyzed using real-time polymerase chain reaction. Osteoclast formation was analyzed using TRAP assay with suppression of Notch receptors by a selective γ-secretase inhibitor (GSI). Results Notch1, Notch2, Delta1, Jagged1, and Hes1 expressions in RAW264.7 cells were upregulated following 50 ng·mL-1 RANKL stimulation for 3 d, concomitant with the expression of the osteoclast differentiation markers TRAP and Cathepsin K. Notch2 and Jagged1 had the most remarkable increase in the Notch family members. GSI inhibited RANKL-induced osteoclastogenesis of RAW264.7 cells and Hes1 expression dose-dependently. Conclusion Notch signaling activation may promote RANKL-induced osteoclastogenesis of RAW264.7 cells.
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