DNCP诱导面突外胚间充质细胞向成牙本质样细胞分化的浓度优化

Concentration optimization of dentin non-collagenous protein on differentiation of ectomesenchymal cells of mandibular process to odontoblast-like cells

目的探讨牙本质非胶原蛋白(DNCP)诱导大鼠面突外胚间充质细胞向成牙本质样细胞分化的最适浓度。方法取妊娠12.5d SD大鼠胎鼠第4代下颌突外胚间充质细胞,DNCP诱导浓度分别为10、50、100、200、500、1 000μg/mL,观察细胞形态变化,诱导6d后采用定量RT-PCR检测牙本质涎磷蛋白(DSPP)、牙本质基质蛋白1(DMP1)的表达。结果6d后除10μg/mL组和1000μg/mL组外,其他诱导组细胞出现极性改变,部分细胞出现平行排列趋势;细胞表达DSPP和DMP1,以200μg/mL组最强,500μg/mL组和200μg/mL组表达量相当,而1 000μg/mL组细胞死亡。结论在DNCP诱导面突外胚间充质细胞向成牙本质样细胞分化中过低浓度不利于成牙本质细胞的分化,而过高浓度则引起细胞死亡。200μg/mL是较为合适的有效诱导浓度。

Objective To investigate the suitable concentration of non-collagenous protein on differentiation of ectomesenchymal cells to odontoblast-like cells. Methods Different doses of non-collagenous protein （DNCP） were added to the 4th passage ectomesenchymal cells. The morphological change was observed by phase contrast microscopy. 6 days later,of mandibular process RT-PCR was used to value the quantity of DSPP and DMP1. Results. No obvious morphological change was observed in 10μg/mL group. Most cells supplemented with 1 000μg/mL DNCP died. Some cells in other differentiation doses were tall columnar with long processes. Some cells polarized. DSPP and DMP1 were expressed in 50,100,200,500μg/mL groups. Quantitative RT-PCR showed the quantity of DSPP and DMP1 in 200μg/mL group were more than that in 50μg/mL and 100μg/mL group. Expression of DSPP and DMP1 in 500μg/mL was not more than that in 200μg/mL group. Conclusion Ectomesenehymal cells can be induced to differentiate into odontoblast-like cells by dentin non-collagenous protein. 200μg/mL DNCP maybe a potential suitable dose in odontoblast-like cells differentiation.