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Zhao, Liya (2016): The morphology and growth behavior of human gingival fibroblasts on three different titanium alloy surfaces. Dissertation, LMU München: Medizinische Fakultät
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Abstract

Research on dental implants over the last decades mainly concentrated on the osseointegration between bone-to-implant interface; however, studies on the assessment of soft tissue incorporation around dental implant are still limited. As innovative materials were developed, each new alternative demands the assessment of its biocompatibility and performance on both bone and soft tissue integration onto dental implant surfaces. The understanding of cell-substrate interactions is of high importance for the development of biocompatible implants, it paves the way for in vivo studies into device functionality, it is important to study how Ti surfaces with different microstructures affect the behaviour of spreading and attached cells. However, the future of the cells on the materials cannot be presumed with the evaluation after short-time seeding, the method that allows long-term study of a cell-to-material interface is closer to the in vivo situation. The goal of this project is to evaluate the influence of different topographies and roughness of titanium specimens on human gingival fibroblast's morphology, adhesion, cellular proliferation with SEM and CLSM and to evaluate by FACS the expression of proteins involved in cell/surfaces adhesion. In this study, the initial attachment and subsequent growth behaviour up to 30 days of human fibroblasts on three different commercial Ti substrates were investigated and compared with confocal microscopic imaging; it could be shown that the extent of fibroblast’s spreading at various points of time differed on the implant surfaces tested, the cells responded to Osseotite surfaces in a manner similar to or even better than their behavior on Nanotite surfaces. The cells cultured on Osseotite and Nanotite surfaces are cuboidal in shape and have the dendritic branching pattern characteristic. In contrast, the cells on the Machined surfaces appear more flattened. Alamar Blue assay demonstrated the gingival fibroblasts grown on Osseotite and Nanotite surfaces showed a notable higher proliferation compared to the Machined surfaces after two weeks of incubation; however, there is no considerable difference on cell proliferation between these two groups. The flow cytometry data analysis suggested that the cells grown on the Osseotite implant material produce a better initial attachment with a higher α5 and β1 integrin expression localized mainly within central areas of the cell grown on the Machined and the Osseotite surfaces; distinct focal contacts localizations were evident at the cell edges on the Nanotite surfaces. At day 30, high vinculin expression and a dense network of actin stress fibers on the human gingivals were observed on all tested substrates; a similar F-actin distribution was found on Osseotite and Nanotite surfaces. The present in vitro study for long-term cellular responses on three different titanium surfaces demonstrated that topographic structures can influence the morphology, proliferation and adhesion of human gingival fibroblasts. With the limitation of our study, there is not enough evidence to show that the Nanotite implants is more beneficial to the growth behavior of human gingival fibroblasts, compared with Osseotite surfaces; especially at the early stage of incubation. Our findings may contribute to a better understanding of the processes involved in the soft tissue integration surrounding dental implants and hopefully give information for the development of innovative implant materials.