Local Drug Release And Biological Evaluation Of Antimicrobial Peptide-loaded Coatings Using A Layer-by-layer Process On Titanium

The prevention and control of peri-implant diseases is a challenge in dental implant surgery. Implants with coatings which have sustained antimicrobial activity is an ideal way of preventing peri-implant diseases. This study reports a non-cytotoxic multilayered coating which had sustained antimicrobial activity and inhibited early biofilm formation. Since some bacteria have multi-drug resistance, taking traditional antibiotics has become a controversial choice. In this study, the broad spectrum cationic antimicrobial peptide, Tet213(KRWWKWWRRC), was grafted to collagen IVthrough sulfosuccinimidyl-4-(p-maleimidophenyl)butyrate (sulfo-SMPB). This compound is renamed as AMPCol. The multilayer AMPCol coatings were assembled on smooth titanium surfaces using a layer-by-layer technique. In general, the titanium plates were immersed in chitosan solution with positive charges(5mg/ml, pH=2.8) for20min, thus a precursor layer with a stable positive charge was formed, which initiated the layer-by-layer assembly process. Next, the titanium plates with a CS layer were dipped into the HA solution (0.5mg/ml, pH=7.2) and kept at room temperature for5 min, during which time HA was adsorbed electrostatically onto the surface. Then the plates with negative charges were dipped into the AMPCol solution(lmg/ml) for5min at room temperature in order to obtain positive charges. The last two steps were repeated. The assembly process and the physicochemical properties of the coating were characterized using XPS, AFM, contact angle, QCM. Through XPS analysis, Ti, O, C, N and S signals were detected on the CS-(HA-AMPCol)10surface. With an increase in layer number, the N signal was enhanced and the Ti signal was decreased. High resolution AFM images showed that granular-like structures and island-like structures could be obsered on the coated titanium surfaces. The value of contact angle fluctuated between34°and40°when the number of coating layers increased. It meaned that the wettability of the coating was much better than the uncoated pure titanium. The QCM results indicate that the thickness of HA-AMPCol multilayer films can be precisely controlled by changing the number of deposition cycles. The AMPCol coating continued to release AMPCol for almost one month.This coating with controlled release of AMP decreased the growth of both a Gram-positive aerob (Staphylococcus aureus) and a Gram-negative anaerobe (Porphyromonas gingivalis) up to one month. When examined over one month(29d), compared with the control group, the broth with coated titanium plates inhibited P. gingivalis and S.aureus by99.31%and99.56%, respectively. Early S. aureus biofilm formation was inhibited by the coating, especialy CS-(HA-AMPCol)7and CS-(HA-AMPCol)10. Noncytotoxicity to epithelial cells (HaCaT) or low erythrocyte hemolysis was observed. Compared with traditional systemic treatment, this local AMP delivery system has better local does and delivery speed. The excellent long-term sustained antimicrobial activity of the new multilayer coating is a potential method for preventing peri-implant diseases.