Research On The Mechanism Of Osteogenesis Induced By Tantalum And Development And Application Of Tantalum Coated Pedicle Screw

Backgrounds and Objectives:Tantalum metal because of its good biocompatibility and osteoinductive properties has been widely used in oral and maxillofacial surgery,joint surgery and repair of bone defects.But because it is different from the traditional titanium alloy,it has high elastic modulus,far greater than human cancellous bone.If the direct application of the load-bearing parts,could cause secondary fractures because of stress shielding effect.We intend to design and fabricate titanium-based tantalum coated pedicle screws though chemical gas phase deposition(CVD)method.The screw can be well combined with the titanium alloy with good mechanical properties and tantalum metal excellent biological characteristics,and animal experiments confirmed that the coated screw could promote bone formation and improve anchoring.In virto experiment,we study the influence of tantalum on adhesion and proliferation of MC3T3-E1 cells and h BMSCs.Also,we'd like to confirm whether tantalum can improve the ability of h BMSCs in absorbing calcium inion in culture medium and mineralizing.Last but not least,we want to figure out the signaling pathways through which tantalum promote the process of osteogenic differentiation.Methods:1.By using chemical vapor deposition(CVD)preparation of tantalum coating in conventional titanium alloy screw surface,and to observe the coating surface morphology by scanning electron microscopy(SEM).2.By using the method of CCK-8 quantitative comparison between MC3T3-E1 cells and h BMSC cells in tantalum and titanium alloy surface adhesion,proliferation,SEM analysis of cell morphology on surface of two kinds metal.3.Using ALP staining and alizarin red staining,analyzed the tantalum and titanium alloy for the differentiation of h BMSC cells.4.The mechanism research of tantalum induced osteogenic differentiation by Real-time PCR and Western Blot method.5.Implant the tantalum coated screws and conventional titanium alloy screws in pigs' pedicle and measure immediate withdrawal force and 3 months after surgery and analysis of the content of trabecular bone around the screw by the method of hard tissue slicing and micro-CT scan and reconstruction.Result:1.SEM showed that tantalum coating prepared by CVD method showing uniform fish scale like structure,increases the surface roughness of the implant can be beneficial for cell attachment.2.The OD value in 450 nm of CCK-8 method showed that MC3T3-E1 cells on 8th hour and h BMSC cells on 2nd,4th,8th hour of the tantalum surface adhesion was higher than titanium alloy,and the difference was statistically significant.3.Both ALP staining was used to evaluate the early differentiation stage(on 7th and 14 th day)and alizarin red staining was used to evaluate the late differentiation stage(on 21 st and 28 th day)demonstrated h BMSC cells in the Tantalum surface showed stronger ability of differentiation,more red calcium nodules were stained.And h BMSC cells on tantalum surface absorbed more calcium ions in the medium also proved this point.4.Quantitative ?-Catenin protein for 7 days,p-Smad3 protein for 21 and 28 days were significantly higher in tantalum group by western blot method.Real-time PCR determination of downstream related-RNA level also had high expression.The use of Smad3 inhibitors blocked the TGF-?/smad pathway,and ALP staining showed that the differentiation of h BMSC cells decreased.5.Animal experiments showed that tantalum-coating screw has greater withdrawal force in 3 month and histological examination showed that the tantalum coated screws induced formation of more bone trabecular.Conclusion:1.CVD method of preparation of titanium-based tantalum coated pedicle screws is simple,and the coating is firmly combined.2.Tantalum can promote cell adhesion and proliferation compared with the traditional titanium alloy screws.3.Tantalum can activate Wnt/?-Catenin pathway and TGF-?/smad pathway to promote osteogenesis.