Study Of Biological Behaviors Of Human Gingival Fibroblasts With Assessment Of Antimicrobial Ability On UVC-activated Titanium

Background:Soft tissue integration of titanium(Ti)dental implants plays a critical role in the maintenance of the long-term stability of implant-supported prostheses and the prevention of peri-implant diseases.Various surface designs have been applied in the commercial titanium implant abutments to improve soft tissue integration in clinical practice,including smooth,micro rough,and nanostructured surfaces.These methods improve the surface affinity for cells by the introduction of special structures on micro or nano scales.However,they are limited to inhibiting bacterial accumulation because either cells or microbes could be affected by the surface structures of the substrates.Thus,it still has some weaknesses in the biological properties of the currently used Ti implant abutments.Recently,Ultraviolet-C(UVC)activation has been reported to improve cell adhesion and reduce bacterial colonization on titanium,which may be expected to overcome this problem.Hence,this study hypothesized that UVC activation promoted the biological properties of titanium by exerting a synergistic effect for enhancing gingival healing and reducing the accumulation of oral bacteria so that improved the soft tissue integration of Ti implants.Objective:This study aimed to investigate the effects of UVC activation on the biological responses of human gingival fibroblasts(HGFs)and Porphyromonas gingivalis(P.g.)on Ti surfaces.Methods:1.The smooth surface design for Ti samples was adopted and fabricated by the process of machining and polishing,then triggered by UVC irradiation.Samples without UVC pretreatment were set as control.The physical properties were analyzed by scanning electron microscopy and atomic force microscopy;the wettability was measured by static water contact angles on the sessile-drop method measuring device;the chemical properties were investigated by energydispersive X-ray spectroscopy and X-ray photoelectron spectroscopy.2.Studies of the biological behaviors of HGFs were as followed:the initial cell adhesion was evaluated by nucleus counting;the morphology of cell spreading with the distribution of focal adhesions was visualized by immunofluorescence staining;the CCK-8 assay was used to assess the proliferating activity of cells;the reverse transcription-quantitative polymerase chain reaction assay was performed to investigate the adhesion-related gene expression levels of HGFs.3.Assessments of the response of P.g.were as followed:the viability of P.g.was evaluated by live/dead bacterial staining;biofilm formation was measured by crystal violet staining.4.The adhesive behavior of HGFs under the pre-accumulation of P.g.was explored by an in vitro co-culture model.Results:1.The smooth Ti surfaces exhibited scratch-like patterns with parallel polishing lines under the observation of scanning electron microscopy.After UVC activation,no significant changes in topography or roughness were observed while a superhydrophilic feature with declination of WCA to 0° was acquired.Meanwhile,the UVC-pretreated surfaces exhibited a higher proportion of hydroxyls and a lower percentage of carbohydrate contaminants compared with the control groups.2.The UVC-activated Ti surfaces increased the number of initially adhered cells and the area of cell spreading,and enhanced the activity of cell proliferation.UVCactivated Ti surfaces displayed a higher level of adhesion-related gene expression level at the initial stage compared to the control groups.3.The UVC-activated Ti surfaces showed a lower live/dead ratio and reduced biomass of P.g.compared with the control groups.4.The UVC-activated Ti surfaces enhanced the initial adhesion and the spread of HGFs under the HGFs-P.g.co-culture model compared with the control groups.Conclusion:UVC activation improved the cleanness and hydrophilicity of Ti surfaces by reducing the carbon contaminants and increasing the hydroxyls without alteration of surface structures.UVC activation enhanced the initial cellular adhesion with expression of focal adhesions and improved the proliferation of HGFs on Ti surfaces.UVC activation inhibited bacterial viability and biofilm formation on Ti surfaces.UVC activation improved the initial adhesion and spread of HGFs on Ti surfaces under the condition of co-culturing with bacteria.