Study On Antibacterial Properties And Cytocompatibility Of Copper-containing Titanium-zirconium Alloys

Objective: This research is based on the clinical application of titanium-zirconium(Ti-Zr)implants.By adding different concentrations of Cupper(Cu),a new type of Ti-Zr-xCu alloy was developed.It is expected that trace Cu ion can be released continuously and stably in body fluids and inhibit the growth and proliferation of bacteria.The purpose of this study is to explore the antibacterial property and biocompatibility of Ti-Zr-xCu biomaterials,and provide evidences for the future application of antibacterial titanium-zirconium narrow-diameter implants.Materials and Methods: Staphylococcus aureus and Escherichia coli were cultured on the surface of Ti-Zr-xCu alloy and determined by plate co-culture method for 24 h.The antibacterial properties of Ti-Zr-xCu alloy were determined by staining the proteins and polysaccharides in bacterial biofilms.The morphology of cytoskeleton was observed by phalloidine staining after co-culturing of 4 and 24h;Murine preosteoblast MC3T3-E1 cells were co-cultured with the biomaterials for 1,4 and 7days.CCK8 assay kit was used to evaluate cell proliferation of Ti-Zr-xCu alloy.The alloy was co-cultured with MC3T3-E1 cells for 72 h and the cell apoptosis was measured by co-staining of propidium iodide and fluorescein isothiocyanate using flow cytometry.Cell differentiation of Ti-Zr-xCu alloy was determined by ALP/BCA assay.The biocompatibility of Ti-Zr-xCu alloy was explored by in vitro studies.Results: The results showed that high concentration of the extracellular proteins and polysaccharides and dense density of bacteria were found on the surface of blank plate and Ti-Zr alloy samples,whilst the concentration and bacteria density of Ti-Zr-xCu alloy samples were lower.The antibacterial rate of Ti-Zr-xCu alloy against Staphylococcus aureus and Escherichia coli was more than 97%,which indicated that Ti-Zr-xCu alloy had excellent antibacterial property.The results of cytoskeleton staining showed that cells adhered and stretched well on the surface of Ti-Zr-xCu alloy,which further benefited the proliferation of adherent cells on the surface of Ti-Zr-xCu alloy.CCK8 assay showed that the relative proliferation rate of MC3T3-E1 on Ti-Zr-xCu alloy surface at 1,4 and 7 days were all higher than 99%,indicating that this alloy had low cytotoxicity.The results of flow cytometry showed that Ti-Zr-xCu alloy had no significant effect on cell apoptosis.ALP/BCA protein assay results showed that Ti-Zr-xCu alloy had no adverse effect on cell differentiation,which further proved that Ti-Zr-xCu had good cytocompatibility.Conclusion: Ti-Zr-xCu biomaterial has good antibacterial capacity and cell compatibility,and has no detrimental effects on cell adhesion,proliferation,apoptosis and differentiation.It has great potential in the clinic application of antibacterial narrow diameter implants.