Antibacterial and osteogenic thin films on Ti-6Al-4V surface formed by passivation process in copper hydroxide solution

Abstract

Implant-associated infections are threatening and devastating complications that lead to bone destruction and loss. As a smooth surface is suitable for inhibiting bacterial adhesion, endowing antibacterial activity to the Ti surface without any structural changes in the surface topography is an effective strategy for preventing infection. The thin film on the Ti-6Al-4 V surface was functionalized to endow antibacterial activity by immersion in a Cu(OH)₂ solution. The resulting surface maintains the surface topography with a surface roughness of 0.03 μm even after the immersion in the Cu(OH)₂ solution. Moreover, Cu was detected at approximately 10 atom% from the surface and was present up to a depth of 30 nm of thin film. In vitro experiments revealed that the resulting surface exhibited antibacterial activity against methicillin-resistant Staphylococcus aureus and allowed the cellular proliferation, differentiation, and calcification of MC3T3-E1 cells. Furthermore, in vivo experiments determined that the presence of Cu in the thin film on the Ti-6Al-4 V surface led to no inflammatory reactions, including bone resorption. Thus, immersion in a Cu(OH)₂ solution incorporates and immobilizes Cu into the thin film on the Ti-6Al-4 V surface without any structural alternations in the surface topography, and the resulting smooth surface exhibits antibacterial activity and osteogenic cell compatibility without cytotoxicity or inflammatory reactions. Our findings provide fundamental insights into the surface design of Ti-based medical devices, to achieve bone reconstruction and infection prevention.

Keywords: Passivation; antibacterial activity; bone formation; copper; infection prevention; metallic biomaterials; smooth surface; titanium alloy.

Graphical abstract

Figure 1.

3D laser microscopy images and surface roughness (a), quantification and optical images of water contact angles (b), scanning electron microscopy images and Ti, Al, V, O, and Cu mappings (c) obtained from NT and Cu10.

Figure 2.

X-ray photoelectron spectroscopy (XPS) survey scan spectra obtained from NT and Cu10 (a), Cu 2p (b), Cu LMM (c) energy region spectra and the Wagner plot of Cu (D) obtained from Cu10.

Figure 3.

The proportion of Cu species calculated from Cu 2p energy region spectrum of Cu10 (a). Ti 2p (b) and Cu 2p (c) energy region spectra obtained from Cu10 before and after sputtering during 0 to 120 s.

Figure 4.

The amount of Cu ions released from the NT, Cu10 and pure Cu into physiological saline during the 24 h of immersion (a). The viability of MRSA (b) and MC3T3-E1 cells (c) on the NT, Cu10 and pure Cu during 24 h of incubation.

Figure 5.

Cellular proliferation of MC3T3-E1 cells on NT and Cu10 after 1, 3, and 7 days of incubation (a). Fluorescence images of the MC3T3-E1 cells attached to the NT and Cu10 after 7 days of incubation (b).

Figure 6.

Alkaline phosphatase (ALP) activity levels in MC3T3-E1 cells on the samples after 7 days of incubation (a). The area percentage of the alizarin red S (ARS) stained area on the samples after 21 days of incubation, and the optical microscope images of ARS-stained samples (b).

Figure 7.

Hematoxylin-eosin-stained sections 4 weeks after the implantation of NT (a, d), Cu10 (b, e), and pure Cu (c, f). (d – f) magnified views of the regions indicated by black squares in panels (a – c).