No abstract

Polyetheretherketone (PEEK) is a high-performance thermoplastic polymer which has found increasing application in orthopaedics and has shown a lot of promise for ‘made-to-measure’ implants via additive manufacturing approaches. However, PEEK is bioinert and needs to undergo surface modification to make it at least osteoconductive to ensure a more rapid, improved, and stable fixation that will last longer in vivo. One approach to solving this issue is to modify PEEK with bioactive agents such as hydroxyapatite (HA). The work reported in this study demonstrates the direct 3D printing of PEEK/HA composites of up to 30 weight percent (wt%) HA using a Fused Filament Fabrication (FFF) approach. The surface characteristics and in vitro properties of the composite materials were investigated. X-ray diffraction revealed the samples to be semi-crystalline in nature, with X-ray Photoelectron Spectroscopy and Time-of-Flight Secondary Ion Mass Spectrometry revealing HA materials were available in the uppermost surface of all the 3D printed samples. In vitro testing of the samples at 7 days demonstrated that the PEEK/HA composite surfaces supported the adherence and growth of viable U-2 OS osteoblast like cells. These results demonstrate that FFF can deliver bioactive HA on the surface of PEEK bio-composites in a one-step 3D printing process.

Tissue-engineered (TE) scaffolds provide an ‘off-the-shelf’ alternative to autograft procedures and can potentially address their associated complications and limitations. The properties of TE scaffolds do not always match the surrounding bone, often sacrificing porosity for improved compressive strength. Previously, the layer-by-layer (LbL) assembly technique was used to deposit nanoclay containing multilayers capable of improving the mechanical properties of open-cell structures without greatly affecting the porosity. However, the previous coatings studied contained poly(ethylenimine) (PEI), which is known to be cytotoxic due to the presence of amine groups, rendering it unsuitable for use in biomedical applications. In this work, poly(diallydimethylammonium chloride) (PDDA)- and chitosan (CHI)-based polyelectrolyte systems were investigated for the purpose of nanoclay addition as an alternative to PEI-based polyelectrolyte systems. Nanocomposite coatings comprising of PEI, poly(acrylic acid) (PAA), Na+ montmorillonite (NC), PDDA, CHI and sodium alginate (ALG) were fabricated. The coatings were deposited in the following manner: (PEI/PAA/PEI/NC), PEI-(PDDA/PAA/PDDA/NC) and (CHI/ALG/CHI/ALG). Results from scanning electron microscopy (SEM) and energy-dispersive X-ray (EDX) analyses demonstrated that the nanoclay was successfully incorporated into each polymer bilayer system, creating a nanocomposite coating. Each coating was successful at tailoring the elastic modulus of the open-cell structures, with polyurethane foams exhibiting an increase from 0.15 ± 0.10 MPa when uncoated to 5.51 ± 0.40 MPa, 6.01 ± 0.36 MPa and 2.61 ± 0.41 MPa when coated with (PEI/PAA/PEI/NC), PEI-(PDDA/PAA/PDDA/NC) and (CHI/ALG/CHI/ALG), respectively. Several biological studies were conducted to determine the cytotoxicity of the coatings, including a resazurin reduction assay, scanning electron microscopy and fluorescent staining of the cell-seeded substrates. In this work, the PDDA-based system exhibited equivalent physical and mechanical properties to the PEI-based system and was significantly more biocompatible, making it a much more suitable alternative for biomaterial applications.

There is continued focus on the development of new biomaterials and associated biological testing methods needed to reduce the time taken for their entry to clinical use. The application of Raman spectroscopy to the study of individual cells that have been in contact with biomaterials offers enhanced in vitro information in a potentially non-destructive testing regime. The work presented here reports the Raman spectral analysis of discreet U-2 OS bone cells after exposure to hydroxyapatite (HA) coated titanium (Ti) substrates in both the as-deposited and thermally annealed states. These data show that cells that were in contact with the bioactive HA surface for 7 days had spectral markers similar to those cultured on the Ti substrate control for the same period. However, the spectral features for those cells that were in contact with the annealed HA surface had indicators of significant differentiation at day 21 while cells on the as-deposited surface did not show these Raman changes until day 28. The cells adhered to pristine Ti control surface showed no spectral changes at any of the timepoints studied. The validity of these spectroscopic results has been confirmed using data from standard in vitro cell viability, adhesion, and proliferation assays over the same 28-day culture period. In this case, cell maturation was evidenced by the formation of natural bone apatite, which precipitated intracellularly for cells exposed to both types of HA-coated Ti at 21 and 28 days, respectively. The properties of the intracellular apatite were markedly different from that of the synthetic HA used to coat the Ti substrate with an average particle size of 230 nm, a crystalline-like shape and Ca/P ratio of 1.63 ± 0.5 as determined by SEM-EDX analysis. By comparison, the synthetic HA particles used as a control had an average size of 372 nm and were more-rounded in shape with a Ca/P ratio of 0.8 by XPS analysis and 1.28 by SEM-EDX analysis. This study shows that Raman spectroscopy can be employed to monitor single U-2 OS cell response to biomaterials that promote cell maturation towards de novo bone thereby offering a label-free in vitro testing method that allows for non-destructive analyses.

Polycaprolactone (PCL) is a well-established biomaterial, offering extensive mechanical attributes along with low cost, biocompatibility, and biodegradability; however, it lacks hydrophilicity, bioactivity, and electrical conductivity. Advances in 3D fabrication technologies allow for these sought-after attributes to be incorporated into the scaffolds during fabrication. In this study, solvent-free Fused Deposition Modelling was employed to fabricate 3D scaffolds from PCL with increasing amounts of graphene (G), in the concentrations of 0.75, 1.5, 3, and 6% (w/w). The PCL+G scaffolds created were characterised physico-chemically, electrically, and biologically. Raman spectroscopy demonstrated that the scaffold outer surface contained both PCL and G, with the G component relatively uniformly distributed. Water contact angle measurement demonstrated that as the amount of G in the scaffold increases (0.75–6% w/w), hydrophobicity decreases; mean contact angle for pure PCL was recorded as 107.22 ± 9.39°, and that with 6% G (PCL+6G) as 77.56 ± 6.75°. Electrochemical Impedance Spectroscopy demonstrated a marked increase in electroactivity potential with increasing G concentration. Cell viability results indicated that even the smallest addition of G (0.75%) resulted in a significant improvement in electroactivity potential and bioactivity compared with that for pure PCL, with 1.5 and 3% exhibiting the highest statistically significant increases in cell proliferation.