钛表面掺钴-钙磷涂层成血管作用的体外研究

Angiogenic property of cobalt and calcium-phosphorus doped coating on titanium surfaces in vitro

目的探讨钛掺钴(cobalt,Co)-钙磷(Ca/P)涂层的成血管作用。方法通过微弧氧化(micro-arc oxidation,MAO)方法在钛表面制备掺钴涂层组(Co组)和掺钴-钙磷涂层组(Co-Ca/P组),以未经MAO处理的纯钛试件作为对照组(Ti组)。通过扫描电镜(scanning electron microscope,SEM)观察涂层表面微观形貌,并通过能谱分析仪检测涂层表面元素成分与含量。应用内皮细胞培养基(endothelial cell medium,ECM)制备各组试件标准浸提液,用于成血管实验。人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)接种于基质胶表面并加入不同浸提液体外培养;于第4、8 h观察新生血管形态与数目,并用Image J软件对新生血管环数和血管节点数进行定量分析。结果SEM观察显示,Co组、Co-Ca/P组涂层表面均出现不规则晶体沉积,直径<2μm,Co-Ca/P组晶体更多且均匀;而Ti组只观察到打磨留下的划痕。能谱分析仪分析显示,Co、Ca、P三种元素被成功掺入到Co-Ca/P组涂层中,Co组只有Co元素掺入,而Ti组未检测到上述三种元素。Co-Ca/P组浸提液中HUVECs形成血管环数和节点数显著高于Co组(P<0.05),而这两组成血管效果显著优于Ti组(P<0.05)。结论掺Co-Ca/P微弧氧化涂层具有较好的体外促血管生成能力,对新型高生物活性种植体涂层研发具有一定价值。

Objective To investigate the effect of cobalt(Co)and calcium-phosphate(Ca/P)doped coating on titanium surfaces and their angiogenic effect.Methods Microarc oxidation(MAO)was used to prepare Co-Ca/P-doped and Co-doped coatings.Titanium(Ti)sheet without MAO treatment was used as control.Scanning electron microscopy(SEM)was used to observe the surface micromorphology of the coatings.Energy dispersive spectrometry(EDS)was also applied to detect the doped chemicals and their contents.Standard soaking solutions of these coatings were prepared using an endothelial cell medium(ECM)solution for subsequent angiogenesis experiments.Human umbilical vein endothelial cells(HUVECs)were cultured on Matrigel with ECM soaking solutions for 4 h and 8 h.The microvessels were observed under a microscope,and the number of microtubules and their interconnecting nodes were analyzed with Image J software.Results Co doped and Co-Ca/P-doped coatings were successfully prepared by MAO,which was demonstrated by both SEM observation and EDS analysis.SEM observation showed that irregular crystals of the above chemicals were present on both Co and Co-Ca/P-doped coatings,commonly with a diameter<2μm.However,more crystals were observed on the Co-Ca/P coatings than on the Co coating,and the distribution of the crystals was more homogenous on the Co-Ca/P coatings.However,only polishing scratches were observed on the Ti sample surface.EDS analysis indicated that in contrast to only Co in the Co coating,Co,Ca and P were doped within the Co-Ca/P coating,and none of the three elements were observed on the Ti plate surface.The number of vascular rings and nodes formed by HUVECs in the extract of the Co-Ca/P group was significantly higher than that of the Co group(P<0.05),and the angiogenic effect of these two components was significantly better than that of the Ti group(P<0.05).Conclusion The Co-Ca/P coating exhibits good angiogenic properties in vitro and is valuable for the development of new titanium implants with high surface bioactivity.