Aspirin/PLGA coated 3D-printed Ti-6Al-4V alloy modulate macrophage polarization to enhance osteoblast differentiation and osseointegration.

Although titanium (Ti) and Ti-based alloy have been widely used as dental and orthopedic implant materials, its bioinertness hindered the rapid osseointegration. Therefore, it is recommended to acquire ideal topographic and chemical characteristics through surface modification methods. 3D printing is a delicate manufacture technique which possesses superior controllability and reproducibility. While aspirin serve as a well-established non-steroidal anti-inflammatory agent. Recently, the importance of immune system in regulating bone dynamics has attracted increasing attention. We herein superimposed the aspirin/poly (lactic-co-glycolic acid) (ASP/PLGA) coating on the 3D-printed Ti-6Al-4V surface with uniform micro-structure to establish the Ti64-M-ASP/PLGA substrate. Scanning electron microscopy (SEM), x-ray photoelectron spectroscopy (XPS) and contact angle test confirmed the successful fabrication of the Ti64-M-ASP/PLGA substrate, with increased wettability and sustained release pattern of ASP. Compared with the Ti64 base material, the Ti64-M-ASP/PLGA substrate showed enhanced M2 and depressed M1 genes and proteins expressions in macrophages. The novel Ti64-M-ASP/PLGA substrate also displayed enhanced osteoblast proliferation, adhesion, extracellular mineralization ability and osteogenic gene expressions when cultured with macrophage conditioned medium in vitro. Furthermore, rat femora implantation model was used for in vivo evaluation. After 4 weeks of implantation, push out test, micro-computed tomography (micro-CT) and histological analyses all confirmed the superior osseointegration capabilities of the Ti64-M-ASP/PLGA implant than the other groups. Our study revealed the synergistic role played by 3D-printed micro topography and immunoregulatory drug aspirin in promoting osteogenesis in vitro and accelerating osseointegration in vivo, thus providing a promising method for better modifying the implant surface. Graphical abstract.

Zn-Incorporated TiO2 Nanotube Surface Improves Osteogenesis Ability Through Influencing Immunomodulatory Function of Macrophages.

PURPOSE: Zinc (Zn), an essential trace element in the body, has stable chemical properties, excellent osteogenic ability and moderate immunomodulatory property. In the present study, a Zn-incorporated TiO2 nanotube (TNT) was fabricated on titanium (Ti) implant material. We aimed to evaluate the influence of nano-scale topography and Zn on behaviors of murine RAW 264.7 macrophages. Moreover, the effects of Zn-incorporated TNT surface-regulated macrophages on the behaviors and osteogenic differentiation of murine MC3T3-E1 osteoblasts were also investigated. METHODS: TNT coatings were firstly fabricated on a pure Ti surface using anodic oxidation, and then nano-scale Zn particles were incorporated onto TNTs by the hydrothermal method. Surface topography, chemical composition, roughness, hydrophilicity, Zn release pattern and protein adsorption ability of the Zn-incorporated TiO2 nanotube surface were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) and X-ray photoelectron spectroscopy (XPS), surface profiler, contact angle test, Zn release test and protein adsorption test. The cell behaviors and both pro-inflammatory (M1) and pro-regenerative (M2) marker gene and protein levels in macrophages cultured on Zn-incorporated TNTs surfaces with different TNT diameters were detected. The supernatants of macrophages were extracted and preserved as conditioned medium (CM). Furthermore, the behaviors and osteogenic properties of osteoblasts cultured in CM on various surfaces were evaluated. RESULTS: The release profile of Zn on Zn-incorporated TNT surfaces revealed a controlled release pattern. Macrophages cultured on Zn-incorporated TNT surfaces displayed enhanced gene and protein expression of M2 markers, and M1 markers were moderately inhibited, compared with the LPS group (the inflammation model). When cultured in CM, osteoblasts cultured on Zn-incorporated TNTs showed strengthened cell proliferation, adhesion, osteogenesis-related gene expression, alkaline phosphatase activity and extracellular mineralization, compared with their TNT counterparts and the Ti group. CONCLUSION: This study suggests that the application of Zn-incorporated TNT surfaces may establish an osteogenic microenvironment and accelerate bone formation. It provided a promising strategy of Ti surface modification for a better applicable prospect.