釉基质蛋白对猪骨髓基质细胞生物学特性的影响

Effects of Enamel Matrix Proteins on the Biological Activity of Porcine Bone Marrow Stromal Cells in Vitro

目的研究釉基质蛋白(enamel matrix proteins,EMPs)对体外培养的猪骨髓基质细胞(bone marrow stromal cells, BMSCs)生物学特性的影响。方法抽取猪髂骨骨髓,全血培养法获得骨髓基质细胞。培养液中EMPs的浓度分别为25、50、100、200ug／ml,以不加EMPs为对照。用比色法检测不同浓度EMPs对BMSCs粘附性的影响。通过计数预定视野中伸展的细胞数,计算。BMSCs在1h、3．5h、6．5h的伸展率。MTT法测定各组细胞的增殖活性。结果猪BMSCs在含有EMPs的培养液中生长良好。对照组以及不同浓度EMPs对细胞粘附性的影响无统计学差异。在1h、3．5h、6．5h,各组细胞的伸展率无显著不同。EMPs对BMSCs的促增殖作用呈浓度和时间依赖性,200ug／ml浓度的EMPs从实验的第三天开始显著促进猪BMSCs的增殖。结论EMPs对体外培养的猪BMSCs的粘附性和伸展率无影响,200ug／ml浓度的EMPs可显著促进猪BMSCs增殖,提示可尝试联合应用EMPs和BMSCs修复牙周组织缺损。

Purpose The purpose of this study was to investigate the influence of enamel matrix proteins (EMPs) on the attachment, spreading and proliferation of cultured porcine bone marrow stromal cells (BMSCs). Methods BMSCs were obtained from porcine bone marrow aspiration and cultured in DMEM medium with 10% FBS.The third passage cells were exposed to various concentrations of EMPs (25,50,100 and 200ug/ml). Controls were BMSCs cultured in DMEM medium without EMPs.The cell attachment was analyzed by a colorimetric assay and cell spreading rates were performed at culture time of 1h, 3.5h and 6.5h by analysis of micrographs taken at predetermined sites of each wells. BMSCs proliferation rates were carried out over a 9-day period and assessed by an MTT assay. Results It was shown that BMSCs well attached and spread on EMPs-coated substrata,however, there were no significant differences between the control group and various concentrations of EMPs groups. The proliferation of BMSCs was significantly stimulated by EMPs in a dose-and time-dependent manner,and EMPs at a concentration of 200ug/ml significantly enhanced BMSCs proliferation from day 3 over the experiment. Conclusions The results demonstrate that EMPs stimulate BMSCs proliferation ,whereas this factor does not affect the attachment and spreading of these cells. These findings lend theoretical support to the combined application of EMPs and BMSCs in repairing periodontal defects.