聚多巴胺辅助红景天苷改善微弧氧化纯钛的成骨性能

Polydopamine assisted salidroside to improve bone formation on micro-arc oxidation of pure titanium

背景:微弧氧化技术目前已被较多用于提高钛植入体骨整合性的研究中,在钛表面构建出多种含不同活性元素的多孔涂层。近年来研究发现,红景天苷具有较好的促成骨活性。聚多巴胺作为材料表面优良的黏合剂和二次吸附的载体具有较好负载活性分子的特性。目的:为进一步提高钛表面微弧氧化涂层的促成骨能力,在多孔形貌表面构建骨整合性优良的复合涂层。方法:在纯钛表面制作微弧氧化涂层,记为MAO组;进一步在微弧氧化涂层表面制作聚多巴胺涂层,记为PDA组;在聚多巴胺涂层表面加载1,2,4 g/L的红景天苷,依次记为Sal-1组、Sal-2组、Sal-4组,利用扫描电镜及X射线能谱仪分析各组涂层的微观形貌、表面元素含量及分布;通过CCK-8及荧光染色实验评估各组涂层表面的细胞增殖活性,通过碱性磷酸酶染色、茜素红染色实验观察各组涂层的促成骨分化及矿化能力;利用计算机分子对接技术预测红景天苷分子的潜在促成骨靶点。结果与结论:①扫描电镜显示,5组试件表面为多孔微米形貌结构,平均孔径相似;X射线能谱仪分析显示,通过聚多巴胺载体可将红景天苷加载到微弧氧化涂层表面;②CCK-8实验显示,Sal-2组、Sal-4组培养第3,5天的MC3T3-E1细胞存活率高于MAO组(P<0.05);荧光染色实验显示,5组试件表面MC3T3-E1间形态差异较明显,与MAO组相比,Sal-2组、Sal-4组细胞形态更好、铺展面积大,与相邻细胞连接较好;③Sal-2组、Sal-4组培养第4,7天的的碱性磷酸酶活性高于MAO组(P<0.05);培养21 d时的茜素红染色显示,各组均有一定程度的钙结节出现,其中Sal-2组、Sal-4组的钙结节数量更加明显;④计算机分子对接结果表明,细胞外调节蛋白激酶2、c-Jun氨基末端激酶3、雌激素受体β、孕激素受体可能是红景天苷发挥促成骨作用时活性较强的靶点;⑤结果表明,通过聚多巴胺将红景天苷负载到钛微弧氧化涂层表面,适当加载量的红景天苷较好地改善了微弧氧化涂层的促成骨活性。

BACKGROUND:Micro-arc oxidation technology has been used to improve the osseointegration of titanium implants.A variety of porous coatings containing different active elements are constructed on the surface of titanium.In recent years,studies have found that salidroside has good bone-promoting activity.Polydopamine,as an excellent binder on the surface of the material and a carrier for secondary adsorption,has the characteristics of loading active molecules.OBJECTIVE:To further improve the bone-promoting ability of the micro-arc oxidation coating on the titanium surface,a composite coating with excellent osseointegration was constructed on the porous surface.METHODS:The micro-arc oxidation coating was made on the surface of pure titanium,which was marked as MAO group,and the polydopamine coating was further prepared on the surface of micro-arc oxidation coating,which was marked as PDA group.Salidroside(1,2,4 g/L)was loaded on the surface of polydopamine coating,which was divided into Sal-1 group,Sal-2 group and Sal-4 group.The micro-morphology,surface element content and distribution of each coating were analyzed by scanning electron microscope and X-ray energy dispersive spectrometer.The proliferation activity of cells on the surface of each group was evaluated by CCK-8 assay and fluorescence staining,and the abilities of bone differentiation and mineralization were observed by alkaline phosphatase staining and alizarin red staining.The potential bone-promoting targets of salidroside were predicted by computer molecular docking technique.RESULTS AND CONCLUSION:(1)Scanning electron microscope showed that the surface of the five groups of specimens had porous micron morphology and the average pore diameter was similar,and X-ray energy dispersive spectrometer analysis showed that salidroside could be loaded on the surface of micro-arc oxidation coating by polydopamine carrier.(2)CCK-8 experiment showed that the survival rate of MC3T3-E1 cells in Sal-2 group and Sal-4 group was higher than that in MAO group on the 3rd and 5th days of culture(P<0.05).Fluorescence staining showed that there were significant differences in MC3T3-E1 morphology among the five groups,and compared with MAO group,Sal-2 group and Sal-4 group had better cell morphology,larger cell spreading area and better connection with neighboring cells.(3)The activity of alkaline phosphatase in Sal-2 group and Sal-4 group was higher than that in MAO group on the 4th and 7th days(P<0.05).Alizarin red staining showed that calcium nodules appeared in all groups on the 21st day,especially in Sal-2 group and Sal-4 group.(4)Computer molecular docking results show that extracellular regulated protein kinases 2,c-Jun N-terminal kinase 3,estrogen receptorβ,and progesterone receptor may be the more active targets for salidroside to promote bone formation.(5)The results showed that salidroside was loaded on the surface of titanium micro-arc oxidation coating by polydopamine,and the bone-promoting activity of micro-arc oxidation coating was improved by proper amount of salidroside.