细胞外基质磷酸化糖蛋白在人牙髓细胞成牙本质分化中的表达

Expression of matrix extracellular phosphoglycoprotein in human dental pulp cells undergoing odontoblastic differentiation

目的检测人牙髓细胞（human dental pulp cells，hDPC）诱导矿化过程中细胞外基质磷酸化糖蛋白（matrix extracellular phosphoglycoprotein，MEPE）的表达变化，探讨MEPE在人牙髓细胞向成牙本质样细胞分化过程中的可能作用。方法酶消化法分离培养hDPC，取诱导前及矿化诱导7、14、21d的hDPC，实时荧光定量反转录聚合酶链反应（RT-PCR）及蛋白质印迹法检测MEPE、牙本质涎磷蛋白（dentin sialophosphoprotein，DSPP）、牙本质涎蛋白（dentin sialoprotein，DSP）、骨涎蛋白（bone sialoprotein，BSP）和I型胶原的表达，蛋白质印迹法检测蛋白表达。结果hDPC经矿化诱导，MEPE、DSPP、BSP和I型胶原mRNA表达呈时间依赖性上调，MEPE和BSP mRNA在诱导各时间点与对照组间的差异均有统计学意义（P〈0.001），DSPP和I型胶原的mRNA相对系数在诱导第21天分别为（12943.33±3805.73）和（250.55±31.86），与对照组间的差异均具有统计学意义（P〈0.05）。蛋白质印迹法检测显示各矿化标记的蛋白表达均较对照组上调。结论在hDPC诱导成牙本质分化的过程中，MEPE与DSPP、DSP、BSP、I型胶原呈现相似的表达变化趋势，提示MEPE与hDPC的成牙本质分化有关，可作为hDPC向成牙本质样细胞分化的标志。

Objective To investigate the expression of matrix extracellular phosphorylated protein （MEPE） in human dental pulp cells （hDPC） undergoing odontogenic induction and explore the role of MEPE in odontoblast-like differentiation. Methods hDPC were isolated by enzymatic digestion and preceded to odontogenic induction for 7, 14 and 21days respectively, hDPC before induction served as controls. The expressions of MEPE, dentin sialophosphoprotein （DSPP）/dentin sialoprotein （DSP）, bone sialoprotein （BSP） and collagen type I were determined by quantitative real-time RT-PCR and Western blotting. Results The mRNA levels of MEPE, DSPP, BSP and type I collagen were increased in a time-dependent manner as hDPC were induced along odontoblastie lineage. Statistical differences were detected for MEPE and BSP mRNA expressions in induced hDPC compared with control group （P 〈 0. 001 ）. For DSPP and type I collagen, the mRNA levels in the induced groups were （12943.33±3805.73） and （250.55±31.86） respectively, which were significantly higher than those in control group on days 21（P〈0.05）. Western blotting also revealed the increased expressions of MEPE, DSP, BSP and type I collagen in the induced DPC. Conclusions hDPC showed analogously temporal expressions of MEPE, DSPP/DSP, BSP and collagen type I while differentiating along odontoblast lineage. MEPE may play an important role in the odontogenic differentiation of hDPC and may be a potential marker of odontoblast-like differentiation.