Experimental Study Of Zinc-modified CaSiO₃ Bioceramics Coating On Biological Response Of MC3T3-E1 Cells In Vitro

ObjectiveThe research of using the method of plasma spray to modificate the surface of titanium and titanium alloy implants,which can enhance the ability to integrate with the bone tissue,is one of hot spot in orthopedics and traumatology. Hydroxyapatite coating is used widely in clinic. However, high dissolution rate of hydroxyapatite and low bonding strength of titanium may result in bad clinical long-term effects.Therefore, the important of the study is to explore the ideal biological coating material. In this experiment,we prepare a series of plasma sprayed zinc-modified CaSiO3 bioceramics coating.After that, the biological safty and biocompatibility with osteoblasts are evaluated,and the biological response of MC3T3-E1 cells in vitro are studied, in order to explore an ideal coating materials with superior biological activity and higher bonding strength to meet clinical need.MethodZinc-modified CaSiO3 bioceramics powders were synthesized by the sol–gel method using zinc nitrate hexahydrate (Zn(NO3)2·6H2O),calcium nitrate tetrahydrate (Ca(NO3)2·4H2O;) and tetraethyl orthosilicate ((C2H5O)4Si, TEOS) as raw materials. And then, the obtained powders were sieved to 80 meshes and then were sprayed onto pure Ti disks.Atmosphere plasma spray(APS)system was applied to fabricate all coatings under modified spray parameters. The component of prepared raw materials powder and coating materials are analyzed by X-ray diffraction (XRD),and surface morphology and characteristics of the coatings was observed by scanning electron microscopy (SEM). Evaluation of biological safety of materials is evaluated by cytotoxicity.test. Then, the attachment difference of MC3T3-E1 cells were studied on zinc-modified CaSiO3 bioceramics coating, CaSiO3 bioceramics coating,and pure titanium sheet without coating using MTT method when cells are cultured 4,8 and 12 hours.We also use scanning electron microscopy (SEM) to observe the adhesive and spread ability of MC3T3-E1 cells.The proliferation of cells on all the surfaces are evaluated by MTT experiments and observation of confocal microscopy.The materials may improve the differentiation ability of MC3T3-E1 cells.we use enzyme-linked immunosorbent assay to detect the expression of ALP,COL-I and BGP.Meanwile,the ability of osteoblast mineralization was observed by alizarin red staining.Results1,XRD shows that the main of ingredients in. zinc-modified CaSiO3 bioceramics powders is Ca2ZnSi2O7 ,and other rare ingredients are Ca2SiO4与CaSiO3.There are no significant difference between the original zinc-modified CaSiO3 bioceramics powders and coating materials in component.SEM observe that the coating has a rough and uneven surface,and the particle size is about 60-120um. ICP results show that the material can release calcium ions(96.60±6.37ppm),silicon ions (39.03±6.29ppm) and zinc ions (9.05±0.79ppm),and the material has a better chemical stability than CaSiO3 bioceramics coating. Cytotoxicity test showes that zinc-modified CaSiO3 bioceramics coatings are nontoxic and have good biocompatibility.2,MC3T3-E1 cells culture in zinc-modified CaSiO3 bioceramics coatings, CaSiO3 bioceramics coatings and uncoated titanium.The cells attach more quickly on zinc-modified CaSiO3 bioceramics coatings(P<0.05),and the early and late proliferation of cells also increases significantly on the coating(P<0.05). The rate of adhesion and proliferation is higher than CaSiO3 bioceramics coatings,which indicate that the presence of zinc ions may benefit to adhesion and proliferation of cells and the coatings has a better cell compatibility.3,The outcome of enzyme-linked immunosorbent assay and experiment of mineralization shows that the zinc-modified CaSiO3 bioceramics coatings can promote the secretion of alkaline phosphatase in the early stage,and can improve the synthesizing of type I collagen(P <0.05) and osteocalcin (P <0.05) in the early and late stage of MC3T3-E1cells differentiation.The coating can promote early osteoblast calcification,and have a better ability of differentiation and mineralization than CaSiO3 bioceramics.ConclusionPlasma Sprayed zinc-modified CaSiO3 Bioceramics Coating has excellent biocompatibility for MC3T3-E1 cells.It can better promote the MC3T3-E1 cells attachment,proliferation in the early stage and improve differentiation and mineralization in the early and late stage.Therefore, zinc-modified CaSiO3 Bioceramics Coating as a new bone implant biomedical material would have a broad application prospects in orthopedics.