Implant related infections result from persistent bacteria adhering to the biomaterial surface before, during, or after surgery, enabling colonization and biofilm formation on the implant . On average, 4% to 10% of implant surfaces are estimated to be contaminated with bacteria, however, the infection rate can be as high as 30% in intensive care units in developed countries, and as high as 45% in developing countries .
Antimicrobial peptides (AMPs) have emerged as a promising alternative to combat a broad spectrum of multidrug-resistant and persistent bacteria as they have shown to be able to successfully prevent bacteria adhesion to biomaterials, to kill bacteria residing within biofilms and to rupture the biofilm structure. Notably, recent studies have shown that covalent immobilization of AMPs onto different biomaterial surfaces increase their long-term stability in vivo and offer a proper orientation of the peptide that may result in enhanced antimicrobial activity . In this study, melimine, a chimeric cationic peptide that has been tested in Phase I and II/III human clinical trials, is covalently immobilized onto the surface of 3D printed medical-grade polycaprolactone (mPCL) scaffolds. The ability of melimine-tethered surfaces to inhibit S. aureus and P. aeruginosa bacteria adhesion is assessed, as well as their ability to prevent biofilm formation in vitro.
Macroporous 3D printed mPCL scaffolds were surface treated using a vacuum plasma cleaner (PDC-002-HP Harrick Plasma, USA) under O2/Ar2 for 6 min at high (45W) power. Plasma-treated scaffolds were then incubated in 2mg/ml 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) in acetate buffer for 30 minutes at room temperature. Scaffolds were washed twice with PBS pH 7.4, and subsequently incubated with a solution of 2 mg/ml melimine for 5 hours to allow covalent binding. Treated surfaces were characterized by X-ray photoelectron spectroscopy (XPS), quartz crystal microbalance (QCM) and time of flight secondary ion mass spectrometry (ToF-SIMS). Furthermore, 3D in vitro assays were used in order to investigate the antibacterial effectiveness of treated scaffolds against S. aureus and P. aeruginosa.
XPS and ToF-SIMS spectra of melimine-treated surfaces confirmed the covalent immobilization of the peptide, as well as its homogeneous distribution throughout the scaffold surface. Further surface characterization using amino acid analysis showed that 82.2 ± 12.4ng of melimine were immobilized onto each scaffold. In addition, the presence of melimine on the surface resulted in a reduction of Staphylococcus aureus and Pseudomonas aeruginosa colonization by 78.4% and 74.1%, respectively, in comparison to the non-modified control specimens. Surfaces maintained their antibacterial properties for 3 days, evidencing inhibition of biofilm formation in vitro.
The results of this study showed the in vitro efficacy of the melimine-treated mPCL surfaces against bacterial colonization.
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