A Water-Based Biocoating to Increase the Infection Resistance and Osteoconductivity of Titanium Surfaces

Abstract

As the population ages, the number of patients undergoing total hip arthroplasty (THA) and total knee arthroplasty (TKA) continues to increase. Infections after primary arthroplasty are rare but have high rates of morbidity and mortality, as well as enormous financial implications for healthcare systems. Numerous methods including the use of superhydrophobic coatings, the incorporation of antibacterial agents, and the application of topographical treatments have been developed to reduce bacterial attachment to medical devices. However, most of these methods require complex manufacturing processes. Thus, the main purpose of this study was to apply biocoatings to titanium (Ti) surfaces to increase their infection resistance and osteoconductivity via simple processes, without organic reagents. We modified titanium surfaces with a combination of aminomalononitrile (AMN) and an antibiotic-loaded mesoporous bioactive glass (MBG) and evaluated both the antibacterial effects of the coating layer and its effect on osteoblast proliferation and differentiation. The properties of the modified surface, such as the hydrophilicity, roughness, and surface morphology, were characterized via contact angle measurements, atomic force microscopy, and scanning electron microscopy. The cell proliferation reagent WST-1 assay and the alkaline phosphatase (ALP) assay were used to determine the degrees of adhesion and differentiation, respectively, of the MG-63 osteoblast-like cells on the surface. Antimicrobial activity was evaluated by examining the survival rate and inhibition zone of Escherichia coli (E. coli). The AMN coating layer reduced the water contact angle (WCA) of the titanium surface from 87° ± 2.5° to 53° ± 2.3° and this change was retained even after immersion in deionized water for five weeks, demonstrating the stability of the AMN coating. Compared with nontreated titanium and polydopamine (PDA) coating layers, the AMN surface coating increased MG-63 cell attachment, spreading, and early ALP expression; reduced E. coli adhesion; and increased the percentage of dead bacteria. In addition, the AMN coating served as an adhesion layer for the subsequent deposition of MBG-containing antibiotic nanoparticles. The synergistic effects of the AMN layer and antibiotics released from the MBG resulted in an obvious E. coli inhibition zone that was not observed in the nontreated titanium group.

Keywords: antibacterial; biocoating; osteoconductivity.

Figure 1

X-ray photoelectron spectroscopy analysis of the surfaces of (a) Ti, (b) aminomalononitrile-titanium (AT), and (c) polydopamine-titanium (PT).

Figure 2

Surface roughness using atomic force microscopy (AFM): (a) uncoated titanium, (b) AMN coating on titanium, and (c) PDA coating on titanium images of the surface.

Figure 3

Cell activity assay. The adhesion and proliferation of MG-63 cells on Ti coated with AMN and Ti coated with PDA for 6 and 24 h were compared. The cell seeding density was 1 × 10⁵ cells/cm². (*) denotes a significant difference p < 0.05.

Figure 4

Morphology of the MG-63 cells on various surfaces after 3 days of culturing: (a1,a2) PDA coating on titanium and (b1,b2) AMN coating on titanium. (a1,b1) are presented at 500× magnification; (a2,b2) are presented at 1000× magnification.

Figure 5

Alkaline phosphatase activity of MG-63 cells cultured on various surfaces for up to 14 days compared with that of cells cultured on Ti.

Figure 6

(a) Fluorescence microscopy images of the live/dead staining of E. coli inoculated on various surfaces after 24 h of culturing. The green and red spots represent live and dead bacteria, respectively. (A) Ti, (B) AT, and (C) DT are presented at 20× magnification. (b) Relative antibacterial efficacy of live/dead cells compared to that of untreated Ti, quantified using ImageJ (ImageJ software 1.42, National Institutes of Health, Bethesda, MD, USA). (*) denotes a significant difference p < 0.05.

Figure 7

SEM images of the Ti surface after various treatments. (a) Nontreated Ti and (b) Ti with the AMN coating plus MBG containing vancomycin. Scale bar: 100 μm.

Figure 8

Zones of E. coli inhibition in various samples. (a) Nontreated Ti, (b) vancomycin soaked in a filter the same size as Ti (control), and (c) Ti with AMN coating plus MBG-containing vancomycin.