抑制p38 MAPK对小鼠釉质发育相关基因表达的影响

The effect of inhibiting p38 MAPK on the expression of genes related to enamel development in mice

目的研究p38丝裂原活化蛋白激酶(p38 mitogen activated protein kinase,p38 MAPK)对成釉上皮中釉质发育相关基因表达的影响,为釉质发育分子机制的研究提供基础。方法在体外小鼠下颌磨牙牙胚的培养基中加入以DMSO溶解的p38 MAPK特异性抑制剂SB203580为实验组,以培养基中加入等量DMSO处理的牙胚为对照组,利用Western blot检测成釉上皮中磷酸化p38(pp38)的蛋白表达水平,实时定量PCR检测成釉上皮中Runt相关转录因子2(runtrelated transcription factor 2,Runx2)、成骨细胞特异性转录因子(osterix,Osx)以及成釉细胞标志物牙成釉细胞相关蛋白(odontogenic ameloblast associated protein,ODAM)、釉成熟蛋白(amelotin,AMTN)、基质金属蛋白酶20(matrix metalloproteinase 20,MMP20)和激肽释放酶4(kallikrein 4,KLK4)的mRNA表达水平。结果Western blot结果显示,在抑制剂SB203580的作用下,小鼠成釉上皮中p38 MAPK的磷酸化水平降低,差异有统计学意义(P<0.05)。实时定量PCR结果表明,SB203580组的转录因子Runx2和Osx以及成釉细胞标志物ODAM、AMTN、MMP20、KLK4的表达均低于对照组,差异有统计学意义(P<0.05)。结论p38 MAPK信号通路可能通过调控转录因子Runx2、Osx及成釉细胞标志物ODAM、AMTN、MMP20和KLK4的表达介导釉质发育。

Objective To study the effect of p38 mitogen activated protein kinase(p38 MAPK)on the expression of genes related to enamel development in the enamel epithelium and to provide a basis for the study of the molecular mechanism of enamel development.Methods The p38 MAPKspecific inhibitor SB203580 dissolved in DMSO was added to the culture medium of mouse mandibular molar tooth germs in vitro as experiment group,and mouse mandibular molar tooth germs treated with the same amount of DMSO were used as control group.Western blot was used to detect the protein expression level of phosphorylated p38(pp38)in the enamel epithelium.Realtime PCR was used to detect the mRNA expression levels of runtrelated transcription factor 2(Runx2),osteoblastspecific transcription factor(Osx),ameloblast markers odontogenic ameloblast associated protein(ODAM),amelotin(AMTN),matrix metalloproteinase 20(MMP20)and kallikrein 4(KLK4)in the enamel epithelium.Results Western blot results showed that under the action of the inhibitor SB203580,the phosphorylation level of p38 MAPK in mouse enamel epithelium decreased,and the difference was statistically significant(P<0.05).Realtime PCR results showed that the expression levels of the transcription factors Runx2 and Osx and the ameloblast markers ODAM,AMTN,MMP20,and KLK4 in the SB203580 group were lower than those in the control group,and the difference was statistically significant(P<0.05).Conclusion The p38 MAPK signaling pathway can mediate enamel development by regulating the expression of the transcription factors Runx2 and Osx and the ameloblast markers ODAM,AMTN,MMP20 and KLK4 in the mouse enamel epithelium.