中国实用口腔科杂志

中国实用口腔科杂志

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不同铜含量的钛铜合金对成骨细胞黏附与迁移的影响研究

李悦1，张燕婷1，何晶1，马凯1，王敏雅1，李胜义2，邓春富1，张二林2，赵宝红1   

  1. 1. 中国医科大学口腔医学院·附属口腔医院种植中心，辽宁省口腔医学研究所口腔种植研究室，辽宁  沈阳  110002；2. 东北大学材料各向异性与织构教育部重点实验室，辽宁  沈阳  110004
  • 出版日期:2017-06-15 发布日期:2017-07-11
  • 基金资助:

    沈阳市科技计划项目（F11-264-1-25）；辽宁省科技计划项目（2013225049）；辽宁省自然科学基金（20170541059）

  • Online:2017-06-15 Published:2017-07-11

摘要：

目的    研究不同铜含量的钛铜合金对成骨细胞黏附与迁移的影响，找到有利于成骨细胞黏附与迁移的最优含铜质量分数。方法    制备含铜质量分数分别为2%、5%和10%的钛铜合金样片，分别命名为Ti-2wt%Cu、Ti-5wt%Cu和Ti-10wt%Cu 3组，以不含铜的纯钛材料样片作为对照，命名为Ti组。小鼠MC3T3-E1成骨细胞接种于置有各组样片的24孔培养板中进行培养。进行以下几个方面的观测与分析：（1）成骨细胞的形态学观察：表面形貌观察、细胞骨架观察。（2）MTT法检测成骨细胞的增殖。（3）细胞划痕实验法分析细胞的迁移情况。采用SPSS 17.0软件对观测数据进行统计学处理。 结果    （1）扫描电镜观察显示：细胞接种6 h与24 h后，Ti-5wt%Cu和Ti-10wt%Cu两组表现出更快速、更充分的细胞黏附与铺展，但Ti-5wt%Cu组表现更为明显；鬼笔环肽染色观察显示：细胞接种2 h后，仅Ti-10wt%Cu组可见细胞骨架肌动蛋白丝，4 h和24 h后Ti-5wt%Cu组细胞铺展面积最大。（2）MTT法结果显示：随着铜含量的增加，材料表面的成骨细胞增殖数量增多，即Ti-10wt%Cu组＞Ti-5wt%Cu组＞Ti-2wt%Cu组＞Ti组，且4组之间差异有统计学意义（P＜0.05），但第7 天时，Ti-10wt%Cu与Ti-5wt%Cu两组之间差异无统计学意义。（3）细胞划痕24及48 h后，划痕平均宽度为Ti组 > Ti-2wt%Cu组 > Ti-10wt%Cu组 > Ti-5wt%Cu组，且4组之间差异有统计学意义（P＜0.05）。结论    在本实验检测的铜含量范围内，含铜质量分数5%对成骨细胞的黏附、细胞骨架的形成与细胞迁移的促进作用最佳。

关键词: 钛铜合金, 成骨细胞, 细胞形态, 细胞增殖, 细胞迁移

Abstract:

Objective    To study the effect of different copper content of titanium-copper alloy on adhesion and migration of osteoblasts，and to find the optimal copper content. Methods    The titanium-copper alloy with copper content of 2%，5% and 10% was prepared and named Ti-2wt% Cu，Ti-5wt% Cu and Ti-10wt% Cu，respectively.The pure titanium material without copper was used as the control，Ti group. Mouse MC3T3-E1 osteoblasts were cultured on the surface of these materials.（1）Morphological observation of osteoblasts：the surface morphology of osteoblasts was observed；the cytoskeleton-actin staining was observed.（2）The proliferation of osteoblasts was detected by MTT assay.（3）Cell migration was analyzed by cell scratch test. SPSS 17.0 software was used for statistical analysis. Results    （1）Under scanning electron microscope：cells showed more rapid and fuller adhesion and expansion on the Ti-5wt%Cu，Ti-10wt%Cu when the cells were seeded for 6 and 24 hours，but Ti-5wt%Cu was more obvious.（2）By phalloidine staining：the cytoskeleton actin filaments were observed only in the Ti-10wt%Cu group when the cells were inoculated with phalloidin for 2h.The spreading area of Ti-5wt%Cu was the largest at 4h and 24h.（3）The MTT results showed：the number of osteoblasts on the surface of materials increased with the increase of copper content，that is，Ti-10wt%Cu＞Ti-5wt%Cu＞Ti-2wt%Cu＞Ti，the difference being statistically significant （P＜0.05）. However，there was no significant difference between Ti-10wt% Cu and Ti-5wt% Cu on the 7th day.（4）After 24 and 48 hours of scratches，the average width of scratches was Ti＞Ti-2wt%Cu＞Ti-10wt%Cu>Ti-5wt%Cu and there was significant difference among the four groups （P＜0.05）. Conclusion    In the range of copper content detected in this experiment，osteoblast adhesion，the formation of cytoskeleton and cell migration are the best in the range of 5% of copper content.

Key words: titanium-copper alloy, osteoblast, cell morphology, cell proliferation, cell migration