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Tissue engineering aims at developing complex composite scaffolds for articular cartilage repair. These scaffolds must exhibit a mechanical behavior similar to the whole osteochondral unit. In situ spherical indentation allows us to map the mechanical behavior of articular cartilage, avoiding removal of the underlying bone tissue. Little is known about the impact of grid spacing, indenter diameter, and induced deformation on the cartilage response to indentation. We investigated the impact of grid spacing (range: a to 3a, where a is the radius of the contact area between cartilage and indenter), indenter diameter (range: 1 to 8 mm), and deformation induced by indentation (constant indentation depth versus constant nominal deformation) on cartilage response. The bias induced by indentations performed in adjacent grid points was minimized with a 3a grid spacing. The cartilage response was indenter-dependent for diameters ranging between 1 and 6 mm with a nominal deformation of 15%. No significant differences were found using 6 mm and 8 mm indenters. Six mm and 8 mm indenters were used to map human articular cartilage with a grid spacing equal to 3a. Instantaneous elastic modulus E0 was calculated for constant indentation depth and constant nominal deformation. E0 value distribution did not change significantly by switching the two indenters, while dispersion decreased by 5–6% when a constant nominal deformation was applied. Such an approach was able to discriminate changes in tissue response due to doubling the indentation rate. The proposed procedure seems to reduce data dispersion and properly determine cartilage mechanical properties to be compared with those of complex composite scaffolds.

In this study, we observe that the poromechanical parameters in human meniscus vary spatially throughout the tissue. The response is anisotropic and the porosity is functionally graded. To draw these conclusions, we measured the anisotropic permeability and the “aggregate modulus” of the tissue, i.e., the stiffness of the material at equilibrium, after the interstitial fluid has ceased flowing. We estimated those parameters within the central portion of the meniscus in three directions (i.e., vertical, radial and circumferential) by fitting an enhanced model on stress relation confined compression tests. We noticed that a classical biphasic model was not sufficient to reproduce the observed experimental behaviour. We propose a poroelastic model based on the assumption that the fluid flow inside the human meniscus is described by a fractional porous medium equation analogous to Darcy’s law, which involves fractional operators. The fluid flux is then time-dependent for a constant applied pressure gradient (in contrast with the classical Darcy’s law, which describes a time independent fluid flux relation). We show that a fractional poroelastic model is well-suited to describe the flow within the meniscus and to identify the associated parameters (i.e., the order of the time derivative and the permeability). The results indicate that mean values of λβ,β in the central body are λβ=5.5443×10−10m4Ns1−β, β=0.0434, while, in the posterior and anterior regions, are λβ=2.851×10−10m4Ns1−β, β=0.0326 and λβ=1.2636×10−10m4Ns1−β, β=0.0232, respectively. Furthermore, numerical simulations show that the fluid flux diffusion is facilitated in the central part of the meniscus and hindered in the posterior and anterior regions.

Simple Summary Polycaprolactone (PCL) is a bioresorbable and biocompatible polymer that has been widely used in long-term implants. However, when it comes to regenerative medicine, PCL suffers from some shortcomings such as a slow degradation rate, poor mechanical properties, and low cell adhesion. The incorporation of ceramics such as bioactive glasses into the PCL matrix has yielded a class of hybrid biomaterials with remarkably improved mechanical properties, controllable degradation rates, and enhanced bioactivity, which are suitable for bone tissue engineering. The use of conventional approaches (such as solvent casting and particulate leaching, phase separation, electrospinning, freeze drying, etc.) in realizing these composite scaffolds strongly affects the control of both the internal and the external architecture of scaffolds, including pore size, pore morphology, and overall structure porosity. Accordingly, 3D printing was used in this study because of the benefits offered over conventional methods, such as high flexibility in shape and size, high reproducibility, capabilities of precise control over internal architecture down to the microscale level, and a customized design that can be tailored to specific patient needs. The optimization of the scaffold structure was previously investigated in terms of architecture through the combination of the Taguchi method and CAD drawing, and, in this study, it was investigated by varying the composition of the composite material. Abstract Polycaprolactone (PCL) is widely used in additive manufacturing for the construction of scaffolds for tissue engineering because of its good bioresorbability, biocompatibility, and processability. Nevertheless, its use is limited by its inadequate mechanical support, slow degradation rate and the lack of bioactivity and ability to induce cell adhesion and, thus, bone tissue regeneration. In this study, we fabricated 3D PCL scaffolds reinforced with a novel Mg-doped bioactive glass (Mg-BG) characterized by good mechanical properties and biological reactivity. An optimization of the printing parameters and scaffold fabrication was performed; furthermore, an extensive microtopography characterization by scanning electron microscopy and atomic force microscopy was carried out. Nano-indentation tests accounted for the mechanical properties of the scaffolds, whereas SBF tests and cytotoxicity tests using human bone-marrow-derived mesenchymal stem cells (BM-MSCs) were performed to evaluate the bioactivity and in vitro viability. Our results showed that a 50/50 wt% of the polymer-to-glass ratio provides scaffolds with a dense and homogeneous distribution of Mg-BG particles at the surface and roughness twice that of pure PCL scaffolds. Compared to pure PCL (hardness H = 35 ± 2 MPa and Young’s elastic modulus E = 0.80 ± 0.05 GPa), the 50/50 wt% formulation showed H = 52 ± 11 MPa and E = 2.0 ± 0.2 GPa, hence, it was close to those of trabecular bone. The high level of biocompatibility, bioactivity, and cell adhesion encourages the use of the composite PCL/Mg-BG scaffolds in promoting cell viability and supporting mechanical loading in the host trabecular bone.

In this work, an interdisciplinary approach was employed to investigate the impact on thermoplastic catheters from the deposition of a thin (180 nm), metallic silver film by a pulsed ablation technique. Our characterization firstly involved tensile and bending tests, each one accompanied by finite element modeling aiming to elucidate the contributions resulting from bulk and coating to the device&rsquo, s mechanical behavior. The morphological assessment of the surface before and after the deposition was performed by atomic force microscopy, specifically implemented to visualize the nanostructured character of the film surface and the extent to which the polymer was modified by the deposition process, focusing on coating delamination due to tensile stress. Finally, thermogravimetric&ndash, differential thermal analysis was carried out to evaluate whether silver deposition has affected the physiochemical structure of the polymer matrix. Our results establish that the deposition does not significantly alter the physical and chemical properties of the device. The presented characterization sets a useful precedent for elucidating how structural properties of polymeric materials may change after coating by electronic ablation techniques, highlighting the importance of employing a comprehensive approach for clarifying the effects of additive manufacturing on medical devices.

Biomimetic bone apatite coatings were realized for the first time by the novel Ionized Jet Deposition technique. Bone coatings were deposited on titanium alloy substrates by pulsed electron ablation of deproteinized bovine bone shafts in order to resemble bone apatite as closely as possible. The composition, morphology and mechanical properties of the coatings were characterized by GI-XRD, FT-IR, SEM-EDS, AFM, contact angle measurements, micro-scratch and screw-insertion tests. Different post-treatment annealing conditions (from 350 °C to 425 °C) were investigated. Bone apatite coatings exhibited a nanostructured surface morphology and a composition closely resembling that of the deposition target (i.e. natural bone apatite), also regarding the presence of magnesium and sodium ions. Crystallinity and composition of the coatings were strongly influenced by annealing temperature and duration; in particular, upon annealing at 400 °C and above, a crystallinity similar to that of bone was achieved. Finally, adhesion to the titanium substrate and hydrophilicity were significantly enhanced upon annealing, all characteristics being known to have a strong positive impact on promoting host cells attachment, proliferation and differentiation.

Due to poor mechanical properties and brittleness of bioactive glasses, the deposition of bioactive glass coatings on bioinert metallic implants for bone regeneration is a promising route to combine the high bioactivity of the glassy phase with the mechanical strength of metallic substrate. The Pulsed Electron Deposition (PED) technique has been recently demonstrated to be an effective method to fabricate highly-adherent and nanostructured bioactive thin films and coatings, with fine control over film composition. In this paper, we investigated the deposition by PED of 45S5 Bioglass ® and of a novel CaO-rich bioactive glass, also containing potassium oxide. Composition, microstructure, surface morphology, wettability and adhesion to the titanium substrate were assessed for both as-deposited and annealed coatings. All samples exhibited a nanostructured surface morphology and high hydrophilicity, both positive features for biological applications. In particular, annealed samples exhibited increased roughness and adhesion degree to the titanium substrate compared to the as-deposited ones. The results showed in this paper suggest that bioactive glass coatings deposited by PED are promising for being further investigated as bioactive coatings for bone implants.

In orthopedics and dentistry, novel approaches for fabricating biomimetic and mechanically robust bioactive coatings are highly desirable in order to truly improve the clinical results of coated implants compared to uncoated ones. In this paper, a biological-like apatite coating is deposited for the first time by plasma-assisted deposition of a natural apatite source. Specifically, we deposited bone apatite-like (BAL) thin films from bone apatite targets by pulsed electron deposition (PED). Morphology, composition, structure and mechanical properties of as-deposited and annealed BAL and stoichiometric hydroxyapatite (HA) films were investigated. While as-deposited BAL and HA films were poorly crystalline at room temperature, they crystallized to an extent very close to that of natural apatite when annealed at 400 °C. In addition, FTIR analysis pointed out that BAL films closely resembled the composition of the starting natural apatite target. Finally, nanoindentation tests indicated that BAL films with high mechanical properties could be deposited by PED.

Nanoindentation has been emerging as a powerful tool for the investigation of the quality of bone tissue, as it allows to relate compositional/structural heterogeneity to mechanical variations at the micro- and nano-scale. To date, the investigation of nanomechanical properties of bone has been almost exclusively limited to healthy or pathologic tissues, while very few studies attempted to evaluate the mechanical properties of the tissue during the regeneration process. Here, we report the analysis by nanoindentation of the mechanical properties of healing tissue considering different animal models (rabbit and sheep), anatomical regions (condylar osteochondral interface and femoral trabecular bone), comparing pre-existent (i.e. native) and newly-formed (i.e. engineered) tissue according to different engineering approaches. In particular, we focused on bone tissue regenerated by means of nanostructured multi-layered biomimetic scaffolds or macroporous hydroxyapatite scaffolds implanted in critical-size bone defects. Our data confirm that nanoindentation represents an extremely useful tool to follow the degree of maturation of bone tissue during the healing process, for instance allowing the comparison of the efficacy of different regenerative approaches. In addition, the knowledge of the level of reconstruction of the osteochondral interface by revealing the (nano-)mechanical gradient can provide clinically relevant information to predict the correct load transfer from the cartilage to the underlying bone tissue

Nanostructured thin films of silver (Ag) are receiving attention in many production fields requiring adaptation to different substrate materials and fine control over surface texture, such as hi-tech industry and biomedicine. Here we report the deposition at room temperature of Ag thin films by pulsed electron ablation, the films thickness ranging from tens to hundreds of nanometers. Films micro- and nanostructure were investigated by means of AFM, STM and XRD, and chemical composition assessed by XPS. The data were elaborated and discussed in the context of depositions under non-local growth effects, and compared with pulsed electron ablation of different target materials. Our films resulted nanometrically smooth and homogeneous, with high degree of purity and characterized by a narrow distribution of grains of nanometric size at all the thicknesses considered. We suggest that a balance between the efficiency of the ablation process and the unique properties of Ag as a metal accounts for the overall reduction of the system size, and is at the origin of the observed nanostructuration. Our results are interesting in the perspective of metals ablation and provide an optimal platform for the study of functional surfaces with high surface-to-volume ratio.

One of the most important issues leading to the failure of total joint arthroplasty is related to the wear of the plastic components, which are generally made of ultra high molecular weight polyethylene (UHMWPE). Therefore, the reduction of joint wear represents one of the main challenges the research in orthopedics is called to address nowadays. Surface treatments and coatings have been recognized as innovative methods to improve tribological properties, also in the orthopedic field. This work investigated the possibility to realize hard ceramic coatings on the metal component of a prosthesis, by means of Pulsed Plasma Deposition, in order to reduce friction and wear in the standard coupling against UHMWPE. Ti6Al4V substrates were coated with a 2 μm thick yttria-stabilized zirconia (YSZ) layer. The mechanical properties of the YSZ coatings were assessed by nanoindentation tests performed on flat Ti6Al4V substrates. Tribological performance was evaluated using a ball-on-disk tribometer in dry and lubricated (i.e. with fetal bovine serum) highly-stressing conditions, up to an overall distance of 10 km. Tribology was characterized in terms of coefficient of friction (CoF) and wear rate of the UHMWPE disk. After testing, specimens were analyzed through optical microscopy and SEM images, in order to check the wear degradation mechanisms. Progressive loading scratch tests were also performed in dry and wet conditions to determine the effects of the environment on the adhesion of the coating. Our results supported the beneficial effect of YSZ coating on metal components. In particular, the proposed solution significantly reduced UHMWPE wear rate and friction. At 10 km of sliding distance, a wear rate reduction of about 18% in dry configuration and of 4% in presence of serum, was obtained by the coated group compared to the uncoated group. As far as friction in dry condition is concerned, the coating allowed to maintain low CoF values until the end of the tests, with an overall difference of about 40% compared to the uncoated balls. In wet conditions, the friction values were found to be comparable between coated and uncoated materials, mainly due to a premature delamination of the coating. Scratch tests in wet showed in fact a reduction of the critical load required to a complete delamination due to a formation of blister, although no change or damage occurred at the coating during the soaking period. Although conditions of high values of contact pressure were considered, further analyses are however required to fully understand the behavior of YSZ coatings in wet environment and additional research on the deposition process will be mandatory in order to improve the coating tribological performance at long distances addressing orthopedic applications.

Realizing hard ceramic coatings on the plastic component of a joint prosthesis can be strategic for the mechanical preservation of the whole implant and to extend its lifetime. Recently, thanks to the Plasma Pulsed Deposition (PPD) method, zirconia coatings on ultra-high molecular weight polyethylene (UHMWPE) substrates resulted in a feasible outcome. Focusing on both the highly specific requirements defined by the biomedical application and the effective possibilities given by the deposition method in the perspectives of technological transfer, it is mandatory to optimize the coating in terms of load bearing capacity. The main goal of this study was to identify through Finite Element Analysis (FEA) the optimal coating thickness that would be able to minimize UHMWPE strain, possible insurgence of cracks within the coating and stresses at coating-substrate interface. Simulations of nanoindentation and microindentation tests were specifically carried out. FEA findings demonstrated that, in general, thickening the zirconia coating strongly reduced the strains in the UHMWPE substrate, although the 1 μm thickness value was identified as critical for the presence of high stresses within the coating and at the interface with the substrate. Therefore, the optimal thickness resulted to be highly dependent on the specific loading condition and final applications.

Osteochondral scaffolds are emerging as a promising alternative for articular cartilage regeneration, although with still controversial results. In particular, the restoration of the osteochondral interface remains an open challenge. The current available investigative procedures are not optimal to quantify the properties of this region, neither to evaluate the quality of the regenerated tissue with respect to the physiological one. This study investigates an advanced procedure able to quantitatively evaluate the mechanical gradient between stiff and compliant tissues, such as in the osteochondral region where the interface between hyaline and calcified cartilage (tidemark) plays an integral role in transferring articular loads from the compliant articular surface to the stiffer underlying bone. A series of nanoindentation line scans was performed along the tidemark - starting from hyaline and expanding across calcified cartilage - on histological sections derived from sheep osteochondral tissue regenerated by a three-layered biomimetic scaffold, as well as to the adjacent healthy tissue for comparative purposes. After an accurate assessment of the indentation parameters, a sigmoid curve-fit function was applied on the reduced modulus profiles to extract gap, width and regularity of the mechanical transition. The designed procedure succeeded in quantitatively assessing the transition between compliant and stiff regions, limiting experimental issues that generally affect the reliability of the indentation mechanical data, such as apex-blunt indenter tip effect, surface roughness, and influence of the substrate. Among the evaluated parameters, the mechanical gap highlighted the main difference between native and regenerated tissues. Thanks to the information retrievable through this procedure, this load transmission area can be further investigated, providing data to tailor osteochondral engineered tissues in the future.

The use of ceramic coatings to improve mechanical, chemical, and biological properties of a large variety of materials including polymers and metals has often produced technologically attractive as well as problematic surfaces to study, because of their complicate morphology compared to smooth surfaces obtained, for example, by atomistic processes. In this work we deposited thin films of zirconia by a new generation pulsed electron deposition system named Ionized Jet Deposition onto four materials, different from each other by structure and surface texture, and applied methods of fractal geometry to investigate their microstructure and roughening mechanism at different thickness values. Our findings show that the film growth does not follow any known class of universality, but is strongly influenced by non-local effects inherent to deposition technique. In this context, we show that deposition onto rough materials is dominated by a strong memory effect that leads to uniform surface coverages that microscopically retain the shape of the substrate. This circumstance is potentially useful for deposition of conformal coatings in view of applications of such plasma-based deposition technique to cases of technological interest.

A previous study reported the structural characterization of biogenic apatite (BAp) thin films realized by a pulsed electron deposition system by ablation of deproteinized bovine bone. Thin films annealed at 400°C exhibited composition and crystallinity degree very close to those of biogenic apatite; this affinity is crucial for obtaining faster osseointegration compared to conventional, thick hydroxyapatite (HA) coatings, for both orthopedics and dentistry. Here, we investigated the adhesion, proliferation, and osteogenic differentiation of human dental pulp stem cells (hDPCS) on as-deposited and heat-treated BAp and stoichiometric HA. First, we showed that heat-treated BAp films can significantly promote hDPSC adhesion and proliferation. Moreover, hDPSCs, while initially maintaining the typical fibroblast-like morphology and stemness surface markers, later started expressing osteogenic markers such as Runx-2 and OSX. Noteworthy, when cultured in an osteogenic medium on annealed BAp films, hDPSCs were also able to reach a more mature and terminal commitment, with respect to HA and as-deposited films. Our findings suggest that annealed BAp films not only preserve the typical biological properties of stemness of, hDPSCs but also improve their ability of osteogenic commitment.

In joint arthroplasty one of the main issues related to the failure of prosthetic implants is due to the wear of the ultra-high molecular weight polyethylene (UHMWPE) component. Surface treatments and coatings have been recognized as enhancing methods, able to improve the tribological properties of the implants. Therefore, the main objective of this work was to investigate the possibility to fabricate yttria-stabilized zirconia (YSZ) coatings on a metal (AISI 316-L) substrate by means of Pulsed Electron Deposition, in order to improve the tribological behavior of the polymer-metal coupling, by reducing the initial wear of the UHMWPE component. In order to optimize the coating characteristics, the effects of working gas pressure on both its morphological and tribological properties were analyzed. Morphological characterization of the films was evaluated by Atomic Force Microscopy (AFM). Coating wettability was also estimated by contact angle (CA) measurement. Tribological performance (coupling friction and wear of UHMWPE) was evaluated by using a ball-on-disc tribometer during highly-stressing tests in dry and lubricated (i.e. NaCl and serum) conditions; friction and wear were specifically evaluated at the initial sliding distances - to highlight the main effect of coating morphology - and after 100m - where the influence of the intrinsic materials properties prevails. AFM analysis highlighted that the working pressure heavily affected the morphological characteristics of the realized films. The wettability of the coating at the highest and lowest deposition pressures (CA ~ 60°, closed to substrate value) decreased for intermediate pressures, reaching a maximum CA of ~ 90°. Regarding tribological tests, a strong correlation was found in the initial steps between friction coefficient and wettability, which decreased as the distance increased. Concerning UHMWPE wear associated to coated counterpart, at 100m a reduction rate of about 7% in dry, 12% in NaCl and 5% in presence of serum was obtained compared to the uncoated counterpart. Differently from what highlighted for friction, no correlation was found between wear rate and morphological parameters. These findings, in agreement with literature, underlined the effect of the deposition pressure on the morphological properties, but suggested that physical characteristics are influenced too. Further research on the deposition process will be required in order to improve the tribological performance of the coating at long distances, addressing - above all - orthopedic applications.

Herein the preparation of Sr2+-doped calcium phosphate (CaP) coatings on poly(etheretherketone) (PEEK) substrates by pulsed electron deposition (PED) was investigated. Hydroxyapatites (HA) substituted with tailored amounts of Sr2+ up to 9 wt% were synthesized to generate the deposition targets. Dense and uniform amorphous CaP films having Sr/Ca molar ratios close to that of targets were achieved. The roughness and the wettability of the bare PEEK were significantly enhanced after coating deposition. The presence of Sr2+ led to a slight increase of the particles size while it did not mainly affect the RMS of the coatings that consisted of nanostructured aggregates of globular shape. After a mild annealing treatment at 130 degrees C for 6 h, the amorphous coatings transformed into nanocrystalline HA films incorporating sr(2+) having Sr/Ca molar ratios close to those of the as-deposited films. The annealing did not affect the topography and the roughness of the coatings, while improved the hardness of the films un-doped and doped at low-extent. This study shows that PED is feasible technique to coat PEEK implants with CaP films with control over the Sr/Ca ratio, which could improve bone fixation and in vivo stability especially under osteoporotic conditions thanks to the anabolic effect of Sr2+. (C) 2017 Elsevier B.V. All rights reserved.

Deposition of nanostructured and low-wear zirconia (ZrO2) thin films on the metallic component of a total joint implant is envisaged to reduce wear of the soft ultra-high molecular weight polyethylene (UHMWPE) counterpart. In this work, morphological surface features, wear resistance and in vitro-biocompatibility of zirconia thin films deposited by the novel Pulsed Plasma Deposition (PPD) method have been investigated. Film thickness, roughness and wettability were found to be strongly dependent on deposition gas pressure. Interestingly, wear rate of UHMWPE disks coupled to zirconia-coated titanium spheres was only poorly correlated to the contact angle values, while film roughness and thickness seemed not to affect it. Furthermore, wear of UHMWPE, when coupled with zirconia coated-titanium spheres, significantly decreased with respect to uncoated spheres under dry or NaCl-lubricated conditions; besides, when using bovine serum, similar results were obtained for coated and uncoated spheres. Finally, suitable mesenchymal stem and osteoblast cells adhesion, proliferation and viability were observed, suggesting good biocompatibility of the nanostructured zirconia films. Taken together, the results shown in this work indicate that zirconia thin films deposited by the PPD method deserve further investigations as low-wear materials for biomedical applications such as total joint replacement.

Joint prosthesis are usually subjected to several failing mechanisms, including wear of the ultra-high molecular weight polyethylene (UHMWPE) insert. The main goal of this study was to assess the possibility to improve the tribological properties of titanium component by depositing zirconia thin films on its surface by pulsed plasma deposition (PPD) method. Zirconia-coated titanium spheres were tested against UHMWPE disks, both in dry and wet conditions. Zirconia films exhibited a homogenous sub-micrometric grain size distribution and low roughness. Interestingly, zirconia-coated titanium spheres showed lower wear rate of the UHMWPE component, compared to uncoated titanium spheres, supporting the feasibility of the proposed approach.