JNK在牙囊细胞破骨细胞向分化的实验研究

An experimental study on JNK differentiation of osteoclast cells in dental foam cells

目的通过特异性抑制剂SP600125抑制JNK信号通路,探究大鼠牙齿萌出中JNK信号在成熟破骨细胞形成过程中的相关调控机制。方法分离培养SD大鼠牙囊细胞。用不同浓度的SP600125对细胞预处理,实时定量PCR检测DFCs中核因子κB受体活化剂配体、骨保护素基因表达量。DFCs和BMMs建立共培养体系,TRAP染色观察OC的数量。结果SP600125在20μmol/L浓度范围内对DFCs的增殖无显著影响(P>0.05),RT-PCR结果示:SP600125干预72 h后,RANKL基因表达量下降,OPG上升,与对照组比较差异均有统计学意义(P<0.05)。TRAP染色阳性计数结果:各组均有OC生成,9 d较7 d产生的OC数显著增加,加入不同浓度的抑制剂组破骨细胞数均有减少,与对照组相比差异有统计学意义(P<0.05)。结论本研究阐明了在大鼠牙齿萌出中,SP600125通过抑制JNK信号通路,对OC形成具有一定的抑制作用,可为牙齿萌出、牙槽骨改建等破骨细胞相关性口腔疾病提供研究基础。

Objective To investigate the mechanism of JNK signaling in the formation of mature osteoclasts during tooth eruption in rats by inhibiting the JNK signaling pathway with the specific inhibitor SP600125. Methods SD rat dental follicle cells were isolated and cultured. Different concentrations of SP600125 were used to pretreat cells. Real-time quantitative PCR was used to detect the expressions of nuclear factor κB receptor activator ligand and osteoprotegerin gene in DFCs. DFCs and BMMs established a co-culture system and TRAP staining was used to observe the amount of OC. Results SP600125 had no significant effect on the proliferation of DFCs under the concentration range of 20 μmol/L (P>0.05). RT-PCR results showed that after 72 hours of SP600125 intervention, the expression of RANKL gene decreased while that of OPG increased. There were statistical differences compared with the control group (P<0.05). Positive TRAP staining results showed that OC was generated in each group, and the number of OC generated on the 9th day significantly increased compared to the 7th day. The number of osteoclasts decreased in the inhibitor group under different concentrations, and the difference was statistically significant compared with the control group (P<0.05). Conclusion This study clarifies that in rat tooth eruption, SP600125 in some degree inhibits the formation of OC by inhibiting the JNK signaling pathway, which can provide a researh basis for the study of tooth eruption, alveolar bone remodeling and other osteoclast-related oral diseases.