Implant Titanium Surface Modification Using Nano-zinc Ion Deposition

Background and PurposeImplant surface microtopography is an important factor influencing implant osseointegration and its long-term survival. The titanium itself does not have anti-bacterial properties; peri-implant inflammation is one of the major factors leading to implant failure. Almost all implanting materials show no or little effectiveness of bone induction. The available treatment methods of implant surface have their own weakness and shortcoming. The implanting material with good biocompatibility and antimicrobial properties has not yet been reported. The ideal implant surface topography remains undefined. The purpose of the present study is to modify the implant surface using nano-zinc ion deposition for increasing of the adaptability of implant denture and improving the long-term implant success rate.Materials and MethodsThe fourth-generation plasma immersion ion implantation deposition device were used to generated the ion implantation and deposition. The device was designed by the national Laboratory of Advanced Welding Production Technology,a famous institute of the Harbin Institute of Technology. Experiments were performed at the Beijing Institute of Stomatology, School of Stomatology, Capital Medical University.Evaluation of the titanium surface propertiesPlasma immersion ion implantation and deposition technologies, electron microscopy (SEM), laser scanning confocal microscope (CO-FOCAL), x-ray diffraction instrument (XRD), X ray photoelectron spectroscopy (XPS), and transmission electron microscopy (TEM) were used to analyze of the surface properties and composition.Antimicrobial testThe morphological observation of colonies and assessment of the results, colony count, the influence of colony adhesion and colony morphology were used to evaluate the antimicrobial effect of modified implant surface.Biological safetyOsteoblast recovery and cultivation, cytotoxicity test (MTT), cell adhesion and proliferation experiments, cell adhesion and proliferation experiments, acridine orange staining, PI flow cytometry, formation of osteoblast adhesion plaque, formation and expression of extracellular matrix of osteoblast cells were used to assess the biological safety. Immunofluorescent assay of the formation of collagen type I, real time-PCR detection of mRNA expression of collagen type I, and Western blotting were performed to detect the expression of collagen type I.In vivo studyMiniature pigs were used for this in vivo study as a large animal model. Titanium Plate (Zn-Ti; experimental group) and cp-Ti (the control group) were implanted into miniature pig jaws; samples were taken at different time points (30d; 60d; 90d) for histological and scanning electron microscopy observation.Results1. Evaluation of the titanium surface properties(1) X ray photoelectron spectroscopyThe full range of X ray photoelectron spectroscopy indicated that the main difference of zinc PIIID (plasma immersion ion implantation and deposition) on the disc was the appearance of the signals of zinc and oxygen. At the same time, the relative atomic concentration of zinc increased gradually with the ion- implanting time, highest concentration of zinc was found in Zn-Ti-80 min (d) group compared with that of other groups.(2) Scanning electron microscope (SEM)Tiny minor fissures were existed in the surfaces cp-Ti, Zn-Ti-20 min and Zn-Ti-40 min, which presented a smooth shape. However, A good many of homogeneously sporadic distributed.micropores on Zn-Ti-60 min and Zn-Ti-80 min surfaces. Especially, surfaces of Zn-Ti-80 min appeared the nano- scale space, which indicated impacting into the basal material on its surface.Diameter about 60-100nm, as the depth of implantation and deposition , diameter about 200nm; the smooth surface about grade 9, roughness 0.4um.2. Antimicrobial testThe zinc implanted titanium could affect adherence and the form of S.mutans group. The adherence amount of bacteria in Zn-Ti-60 min and -80 min group were significantly less than those in cp-Ti group, Zn-Ti-20 min and -40 min group. The similar results were found in the volume of bacterium on modified surface during the ion-implanted and deposition procedure, e.g. few bacterial colony in Zn-Ti-80min.3. Biological safety(1) MG-63 cells were of irregular polygon shapesSEM analysis revealed that Zn-Ti In vitro the morphologies of MG-63 cells were influenced ,which were revealed through the analysid of SEM.An increased cell-to-substrate contact ratio was found in the spreading and flattening of cells on the Zn-Ti discs,which was faster than the ratio of those grew on the cp-Ti discs.The density of zinc ion improved the cells activities, such as proliferation, differentiation and externalization. Furthermore, the implanting time was contributed with the density of cells, which on the cp-Ti had far less and shorter fibrillar extensions.(2) The number of cells which were observed in different times.No statistical differences were found after 6 hours.However, the number of cells attached on Zn-Ti surfaces increased rapidly compared with the cp-Ti surfaces after 24 hours. The trend was more significantly with the time extended. But no significant difference were found between the Zn-Ti-60 group and the Zn-Ti-80 group.4. In vivo studyThe grinded bone-implant slices were obtained after histological pretreatment. Bone integration and osteogenesis with bone regenerative materials were clearly detected. Much better bone regeneration was found in experimental groups (Zn-Ti) compared with the control groups (cp-Ti).ConclusionOur data demonstrated that implant modification using the nano-zinc implantation and deposition technologies can improve biocompatibility and antimicrobial properties of implants, which may have potential wide clinical application for dental implantation.