| Part one:Construction of TiNF structure on titanium surfaceObjective:To explore the methods of construction of different diameters TiNF on titanium surface. Then, we study the effect of different experimental parameters on TiNF shape to prepare for the next phase of research.Methods:First, titanium sheets were divided and put into the anodization apparatus under different anodization conditions. The control conditions including time, electrolyte and voltage, while, the voltage were the main controlling factor. The samples were divided into five groups according to the different electrolysis voltage; each set of three specimens, namely 5V,25V,40V,60V and control groups (Ctrl group).5V and 25V group were inorganic mixed acid solution,40V,60V electrolyte group were organic mixed acid solution, control group were not treated. Finally, the surface-shape was morphology by scanning electron microscopy (SEM) and atomic force microscopy (AFM) observes the titanium sheet and calculate the surface roughness.Results:(1). The diameter of TiNF obtained by the anodic oxidation treatment was positive relative with the voltage;(2). When the voltage was 5V,25V, with the inorganic acid mixture as electrolyte, the TiNF diameter were about 40nm,80nm; when the voltage was 40V,60V, with organic acid mixture as electrolyte, the TiNF diameter were about 100nm, and 120nm.(3). the boundary of TiNF structure formed with organic electrolyte in each group was relatively clear, but TiNF formed with inorganic electrolyte was unclear.Conclusion:(1). we can change the diameter of the oxide film TiNF by adjusting the electrolysis voltage size, and the voltage were proportional to the diameter of TiNF(2). the kinds of electrolyte could influence the boundary of TiNF;(3). TiNF morphological structure was more stable, when the electrolysis voltage was 5V and 40V.Part two:Investigate the adhesion of human gingival fibroblasts on TiNF surfaceObjective:using the stable group of TiNF material (5V,40V group) to explore the impact of adhesion of human gingival fibroblasts, to provide a theoretical basis for dealing with the implant neck.Method:Specimens were divided into three groups, namely 5V,40V titanium group and Ctrl group. The titanium plate were divided into 3 groups and were cleaned with double distilled water for 3 min, to remove impurities on the surface, then routine autoclaving, dry and set aside. The 4th generation of human gingival fibroblasts were co-cultured in different groups of titanium, each set three vice holes were cultured 2 hours,8 hours,1 day,2 days. Respectively by means of detection, such as live/dead cell staining, MTT; the major extracellular matrix adhesion proteins:FN-1 expression level was detected by ELISA; the extracellular matrix VCL (Vinculin) and COL1-al (Collagen type I-alphal) expression were explored by RT-PCR technology to detect.Results:(1) Live/Dead stain and MTT results can be seen:in the 40V group, human gingival fibroblasts are fully extended, cell adhesion and proliferation were best, and the cells gradually depend on the culture time.(2) ELISA and RT-PCR to detect the extracellular matrix visible FN-1 adhesion proteins, COL1-al and VCL were better expression in titanium surface of 40V group compared with 5V group and Ctrl group (P<0.05), and the presentation was time-dependent expression.Conclusion:The anodizing voltage of 40V, the titanium surface is recessed nanometer diameter of about 100 nm, the gingival fibroblasts adhesion performance was best, and this performance impact structure for cell adhesion also was in a time-dependent manner. |
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