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目的 :使用微弧氧化(micro-arc oxidation,MAO)方法对纯钛圆片进行表面处理,观察其对MC3T3-E1成骨前体细胞生物学行为的影响。方法:使用分级电压增强过程对纯钛试件进行MAO处理,场发射扫描电镜观察试样表面形态,测量不同表面的接触角。MC3T3-E1细胞接种于试样表面并进行体外培养,MTT法检测60、120 min不同表面的细胞黏附量;培养4 h后,激光共聚焦扫描显微镜观察附着细胞的骨架蛋白;24、72、120和168 h后,检测细胞增殖情况;培养第16天,实时荧光定量PCR(q PCR)检测细胞成骨相关基因的表达。采用SPSS16.0软件包对数据进行统计学分析。结果:MAO处理后,可在纯钛表面形成一层多孔氧化层,MAO钛表面水和甘油的接触角显著小于抛光钛表面。在2个时间点,细胞在MAO钛表面的黏附均高于抛光钛表面。肌动蛋白染色提示,在MAO钛表面,细胞伸展良好。从培养第72 h开始,细胞在MAO钛表面的增殖略高于抛光钛表面,但差异无显著性。q PCR结果表明,RUNX2和ALP基因的m RNA水平在MAO钛表面和抛光钛表面之间并无显著差异。结论:MAO处理可促进MC3T3-E1细胞在纯钛表面的早期黏附,但RUNX2和ALP基因的m RNA表达水平在各组间无显著差异。
OBJECTIVE: To investigate the effect of micro-arc oxidation (MAO) on the purity of MC3T3-E1 osteoblast precursors. Methods: The pure titanium samples were treated with MAO by using the step-up voltage enhancement process. The surface morphology of the samples was observed by field emission scanning electron microscope and the contact angles of different surfaces were measured. MC3T3-E1 cells were inoculated on the surface of the samples and cultured in vitro. The cell adhesion amount on different surfaces at 60 and 120 min was measured by MTT method. After culturing for 4 h, the scaffold proteins attached to the cells were observed by confocal laser scanning microscopy. 24,72,120 And 168 h later, cell proliferation was detected. On the 16th day, the expression of osteogenesis-related genes was detected by real-time quantitative PCR (qPCR). SPSS16.0 software package for statistical analysis of the data. Results: After MAO treatment, a porous oxide layer was formed on the surface of pure titanium. The contact angles of water and glycerol on the surface of MAO titanium were significantly smaller than those of polished titanium surface. At 2 time points, the adhesion of cells to the MAO titanium surface was higher than that of the polished titanium surface. Actin staining suggested that on the MAO titanium surface, the cells stretched well. From 72 h after culture, the proliferation of cells on the surface of MAO titanium slightly higher than polished titanium surface, but the difference was not significant. q PCR results showed that m RNA levels of RUNX2 and ALP genes did not differ significantly between MAO titanium surfaces and polished titanium surfaces. Conclusion: MAO treatment can promote the early adhesion of MC3T3-E1 cells on the surface of pure titanium. However, there is no significant difference between the two groups in mRNA expression of RUNX2 and ALP.