Study On Key Factors Of Dopamine-mediated Silver Nanoparticles Surface Construction And The Evaluation Of Antibacterial Property In Vivo And In Vitro

Objectives Titanium and its alloys are widely used in oral implant restoration and hard tissue repair due to their desirable mechanical and biocompatibility properties.Dental implant itself does not have any antibacterial properties.The key factor for the success of implant is to modify the surface of implant without affecting its biocompatibility.The in-situ formation of silver nanoparticles(Ag NPs)on the surface of implants via reduction of Ag~+by dopamine has been extensively studied to achieve the purpose of endowing implant surfaces with antibacterial properties.In previous studies,the preparation of Ag NPs has focused on controlling the concentration of silver ions in the solution,with little reference to the p H of the reaction solution.In this research we systematically determined the influence of various p H(4,7,10)and Ag~+concentrations(0.01,0.1 mg/m L)to the formation of Ag NPs,followed by the evaluation of the antibacterial property in vitro and in vivo,as well as the biocompatibility of the Ag NPs bearing titanium.Methods Porous titanium was formed by means of alkaline heat treatment,then porpus titanium depositing with polydopamine was formed via immersing in dopamine solution,and then immersed in silver nitrate solution with different concentrations and p H values,antibacterial porous titanium loaded with silver nanoparticles was finally obtained.SEM,XPS,Uv-vis,QCM-D and water contact angle test were used to detect the successful construction silver nanoparticle coating.The antibacterial experiments in vitro including bacterial proliferation test,zone of inhibition test,spread plate method and Live/Dead staining.The in vivo antibacterial experiment process was to implant a titanium rod into the bone marrow cavity of SD rat femoral osteomyelitis constructed with S.aureus suspension.After 1 and 7 days of implantation,the samples were removed for further analysis.With the help of spread plate method to compare adhered bacteria on the surface of different titanium rods bacteria.These two experiments comprehensively evaluated the antibacterial ability in vitro and in vivo.The cytocompatibility of the coating to MC3T3-E1 line was evaluated by fluorescence staining.The co-culture of bacteria with cells was performed to evaluate if the antibacterial implant can function well in the complex environment and its importance for cell growth.Various samples pretreated with S.aureus were implanted subcutaneous into SD rats and collected for hematoxylin-eosin staining one month later to analyze the effect of the antibacterial properties of the materials on the inflammatory response of surrounding tissues.Results The results of material characterization showed that the alkaline environment was conducive to the formation of silver nanoparticle and resulted in more silver nanoparticles and more stable size.In addition,compared with the concentration of silver ions,the p H was easier to control the surface silver nanoparticles loading.The results of antibacterial experiments in vitro and in vivo showed that the silver nanoparticle loaded coatings had good antibacterial properties.When the concentration of silver nitrate solution was 0.1 mg/m L and the p H value was 10,the antibacterial activity of the coating was stronger than that of other groups,and the antibacterial activity could be maintained in the simulated body fluid for at least 7 days.The results of cell fluorescence staining showed that,although the adhesion of osteoblasts was affected to some extent by the coating,the cytocompatibility was significantly improved after 7 days of mineralization in simulated body fluid.The results of co-culture of bacteria and cells showed that the antibacterial ability of the coating surface created favorable conditions for cell adhesion and growth.The results of HE staining showed that the coating could inhibit the inflammatory response to some extent in the surrounding tissues.Conclusion The p H value and concentration of the silver nitrate solution in the reaction system occupied an important position in the amount and size of silver nanoparticles generation,and the p H value has a greater impact.This provides an opportunity to develop implant surfaces that meet different antibacterial requirements and avoid waste of silver ions.It is of great significance in clinical application.