R. Miralami, F. Namavar, G.M. Thiele, J.G. Sharp, K.L. Garvin
University of Nebraska Medical Center,
Keywords: nanostructure, TiO2, tatanium implant, orthopaedic, SAOS-2, microscopy, biocompatibility
Summary:Introduction: Annually, more than one million Americans undergo arthroplastic surgeries, and by 2030 the projected number will exceed more than four million . Thus, there is a serious need to accommodate such overload. It has been shown that morphology and structure of the surface influence cellular behavior and interactions with bone which may increase implant longevity and decrease the necessity of revision surgery . Thus, there is a need to modify orthopaedic implants to improve their biocompatibility, and enhance osteoblast activation to induce more/stronger bone. One way to manipulate surface modifications on the implants is using ion beam assisted deposition (IBAD) technique. Therefore, it was the hypothesis of this study that applying IBAD technique to produce newly engineered nano-crystal films of titanium oxide (TiO2) will change the surface topography and increase initial cell adhesion, survival, growth and proliferation. Material and Methods: The IBAD combines an electron beam evaporation system with a simultaneous ion beam bombardment in a high vacuum environment. IBAD processes employ energetic ions to “stitch” TiO2 thin films with hydrophilic properties to the substrate . Atomic force microscopy of IBAD TiO2 with oxygen bombardment showed a roughness of 9.8 nm for a 5 micron scan size. A human osteosarcoma cell line (SAOS-2) was used to compare the biocompatibility of these nano-engineered surfaces with orthopaedic-grade Ti. To measure cell attachment and growth, DAPI stained nuclei were scanned and counted using Metamorph software after short term adhesion (2 hours) and long term adhesion (24 hours) and adherent cell proliferation (24 and 48 hours). For determination of calcium deposition, as an indication of successful in vitro bone formation, Alizarin red quantification was applied. Finally, actin stress fiber patterns were examined to compare actin remodeling at focal adhesion sites on the various surfaces. Data were derived from five independent experiments (n=5). Statistically significant differences between various substrates were evaluated using ANOVA with post-hoc Bonferroni’s multiple comparison test. Results and Discussion: Our results, showed that TiO2 support more adhesion and growth of SAOS-2 cells. Also, the attached cells labeled with actin showed an increase in focal adhesion patterns, indicating there were a greater number of adhesion sites on the nanomaterial. Calcium deposition on different substrates, using the Alizarin Red Assay, showed that more calcium was deposited on IBAD nanocrystalline TiO2 as compared to biomedical Ti by the 7th and 14th days of culture. Conclusion: These experiments clearly indicate that nanocrystalline TiO2 is superior to microcrystalline Ti in supporting growth, adhesion, and proliferation of bone forming cells. Improving the quality of surface oxide, i.e. fabricating stoichiometric oxides as well as nano-engineering the surface topology is crucial for increasing the biocompatibility of Ti implant materials.