神经生长因子对兔牙髓干细胞体外增殖、成骨分化影响的实验研究

目的探讨神经生长因子（nerve growth factor，NGF）对兔牙髓干细胞（dental pulp stem cells，DPSCs）体外增殖及成骨分化的影响，为牙髓组织的损伤修复提供新线索。方法采用酶解组织块法分离培养健康新西兰兔的DPSCs，光镜下观察其形态学及生长变化。将培养的第3代DPSCs分为3组:空白组（只有DPSCs）、实验组（DPSCs混合100 μg/L NGF）和对照组（DPSCs混合矿化液）。观察实验7 d和14 d时3组的细胞形态及碱性磷酸酶活性结果，RT-qPCR检测各组Runx-2、COL-1以及OCN的基因表达水平。结果体外培养的DPSCs生长良好，贴壁生长，大多为多角形或长梭形，呈巢式或集落生长；3组的碱性磷酸酶活性在实验14 d时均高于实验7 d时的数值，且7 d和14 d时实验组与空白组、对照组差异均有统计学意义（P < 0.05）；RT-qPCR结果显示培养14 d时实验组中Runx-2、COL-1和OCN基因的表达量均高于空白组（P < 0.05）。结论NGF可以提高DPSCs的增殖和成骨分化能力，两者的联合应用修复效果优于DPSCs单独应用。

Effect of nerve growth factor on the proliferation and differentiation of rabbit dental pulp stem cells in vitro

ObjectiveTo investigate the effects of nerve growth factor(NGF) on the proliferation and differentiation of rabbit dental pulp stem cells(DPSCs) in vitro, and provide a new clue for the repair of dental pulp tissue.MethodsThe DPSCs from healthy New Zealand rabbits were isolated and cultured by tissue enzymedigestion method, and the morphology and growth changes of cells were observed under light microscope.The third passage DPSCs were divided into the blank group(only DPSCs), experimental group(DPSCs mixed with 100 g/L NGF) and control group(DPSCs mixed with mineralization solution).The cell morphology and alkaline phosphatase(ALP) activity in three groups were investigated after 7 days and 14 days of culture, and the mRNA expression levels of COL-1, Runx-2 and OCN were detected using RT-qPCR.ResultsThe DPSCs cultured in vitro grew well, adhered to the wall, and most of them were polygonal or spindle shaped with nest or colony growth.The ALP activity value in three groups after 14 d of culture were higher than that on the 7th day of culture, and the differences of which among three groups after 7 and 14 d of culture were significant(P < 0.05).The results of RT-qPCR showed that the expression levels of COL-1, Runx-2 and OCN mRNA in experimental group after 14 d of culture were higher than those in blank group(P < 0.05).ConclusionsThe NGF can promote the proliferation and osteogenic differentiation of rabbit DPSCs, and the combined application effect of the two is better than that of DPSCs alone.