Abstract
The aim of the work is to develop innovative antibacterial surface modification treatments for titanium capable to limit the bacterial adhesion and proliferation as weel as the biofilm formation while maintaining an high osteointegrative potential. The goal is to contrast the infections which represent a serius complication related to the use of implantable devices.
Introduction
Titanium and titanium alloy are considered the golden standard materials for the applications in contact with bone especially for dental and orthopaedic applications. To extend the implantable component lifetime and increase their clinical performance some surface modifications are required, to promote and speed up the osteointegration process increasing the rate of bone bonding. Unfortunately, among the different complications related to the use of titanium implantable devices the infections represent the most serious, often leading to implant failure and revision. The use of surface modification with specific metal ions represents a promising approach to fight implant-related infections. In particular gallium has recently shown efficacy in the treatment of infections: exploiting the chemical similarity of Ga3+ with Fe3+, it can interfere in the iron metabolism for a wide range of bacteria. The aim of this work is to develop and characterise new biocompatible biomimetic treatments with anodic spark deposition (ASD) technique on titanium characterised by antibacterial properties maintaining high osteointegrative potential.
Experimental Methods
Three surfaces were developed using titanium grade 2 samples (12 mm diam., 0.5 mm thick): i) SiB-Na: ASD treatment performed in an electrolytic solution containing Ca, P, Si and Na1 used as control; ii) GaOss: ASD treatment performed in the SiB-Na solution enriched with gallium nitrate and oxalic acid; iii) GaCis: ASD treatment performed in the SiB-Na solution enriched with with gallium nitrate and L-cysteine. The ASD was carried out in galvano-static condition with a current density of 10 mA/cm2 reaching 295V (for SiB-Na, GaCis) and 310V for GaOss. Untreated Ti was used as control. The surface morphology and chemistry were analysed using SEM, EDS and XPS. Ga release in D-PBS was studied up to 21 days using ICP/OES analysis. The structure of the titanium oxide was investigated using XRD while the surface wettability was studied using OCA measurements. The coating mechanical stability was evaluated using scratch test and three-point bending test. Human osteoblastic cells (Saos2) indirect citotoxicity was asessed using Alamar Blue assay. Saos2 morphology and adhesion to the treated surfaces were evaluated using SEM and actin staining. Saos2 viability was assessed up to 21 of cell cultured in direct contact with antibacterial surfaces while the Saos2 alkaline phosphatase activity (ALP) was evaluated up to 21 day as a marker of new bone formation. The antibacterial properties were assessed with S. mutans, S. epidermidis and E. coli bacterial strains even after 21 days of the antibacterial agents release to test the long lasting antibacterial activity. Also the effectiveness in limiting biofilm formation was evaluated against S. epidermidis and A. baumanni biofilm producers.
Results and discussion
The developed surfaces showed a microporous morphology without the presence of any delamination. The EDS showed the presence of Ga, Si, P and Ca for GaCis and GaOss. Ga-based treatments revealed a similar concentration of the antimicrobial agent although the Ga released from GaOss was extremely higher than on GaCis. XRD analysis revealed the presence of the anatase cristallographic form which is associated with an higher surface wettability than Ti. The coatings showed a good mechanical stability both after three point bending test and scratch test. The antibacterial surfaces did not show any indirect citotoxicity for Saos2. Also the cell morphology and viability were not negatively affected by the presence of the antibacterial agent: GaOss treatment displayed a stimulating effect on ALP activity of osteoblastic cells than controls. A strong reduction of bacterial adhesion and proliferation for both of the Ga-based treatments especially for GaOss (≈ 40% vs Ti) was observed as well as a long-lasting antibaterial activity. Finally, a significative reduction of S. epidermidis and A. baumanni biofilm production than Ti was observed for GaOss and GaCis treatments.
Conclusion
The treatments developed in the present study represent a promising class of antibacterial and osteointegrative coatings for titanium in particular for dental and orthopaedic applications.
