C. Luculescu

Structural, compositional, mechanical characterization and biological assessment of bovine-derived hydroxyapatite coatings reinforced with MgF2 or MgO for implants functionalization.

Hydroxyapatite (HA) is a consecrated biomaterial for bone reconstruction. In the form of thin films deposited by pulsed laser technologies, it can be used to cover metallic implants aiming to increase biocompatibility and osseointegration rate. HA of animal origin (bovine, BHA) reinforced with MgF2 (2wt.%) or MgO (5wt.%) were used for deposition of thin coatings with improved adherence, biocompatibility and antimicrobial activity. For pulsed laser deposition experiments, a KrF* (λ=248nm, τFWHM≤25ns) excimer laser source was used. The deposited structures were characterized from a physical-chemical point of view by X-Ray Diffraction, Fourier Transform Infra-Red Spectroscopy, Scanning Electron Microscopy in top- and cross-view modes, Energy Dispersive X-Ray Spectroscopy and Pull-out adherence tests. The microbiological assay using the HEp-2 cell line revealed that all target materials and deposited thin films are non-cytotoxic. We conducted tests on three strains isolated from patients with dental implants failure, i.e. Microccocus sp., Enterobacter sp. and Candida albicans sp. The most significant anti-biofilm effect against Microcococcus sp. strain, at 72h, was obtained in the presence of BHA:MgO thin films. For Enterobacter sp. strain a superior antimicrobial activity at 72h was noticed, in respect with simple BHA or Ti control. The enhanced antimicrobial performances, correlated with good cytocompatibility and mechanical properties recommend these biomaterials as an alternative to synthetic HA for the fabrication of reliable implant coatings for dentistry and other applications.

Laser micro-patterning of biodegradable polymer blends for tissue engineering

We propose a multistep all laser, maskless, and solvent free synthesis of micro-patterned substrates of biodegradable polymer blends, with applicability for guided cell adhesion and localized hyaluronic acid (HA) immobilization. The polymer blends comprised polyurethane (PU), poly(lactic-co-glycolic acid) (PLGA), and polylactide-polyethylene glycol-polylactide (PPP) in 1:1:1 blending ratios. Polymer patterning was performed by laser processing in two steps. First, the polymers were patterned with periodic micro-channels by direct femtosecond laser ablation, which provided flexibility in design and spatial accuracy for the patterns. As a second step, the micro-patterned polymers were coated with thin layers of polymer blends using matrix assisted pulsed laser evaporation (MAPLE). The resulted sandwich substrates were composed of a bottom, micro-patterned layer and thin, top layer which conserved the patterns from the underlying layer and preserved the polymers chemical composition. Depending on the bottom/top layers, the substrates were denominated PU/PU:PLGA:PPP and PU:PLGA:PPP/PU:PLGA:PPP, respectively. The laser generated micro-patterns were used for selective attachment of oral keratinocyte stem cells and for HA immobilization. The highest cellular density was found on the PU:PLGA:PPP/PU:PLGA:PPP substrate, where the spongy-like micro-channels provided multiple anchoring points for the cells. For both substrates, the micro-channels enabled localized immobilization of HA. The effectiveness of HA immobilization was tested against cell adhesion and protein adsorption.

One-step preparation of nitrogen doped titanium oxide/Au/reduced graphene oxide composite thin films for photocatalytic applications

Titanium dioxide (TiO2) and TiO2/Au/reduced graphene oxide (rGO) nanocomposite thin films were grown by ultraviolet matrix assisted pulsed laser evaporation (UV-MAPLE) in controlled O2 or N2 atmospheres. An UV KrF* excimer laser (λ = 248 nm, τFWHM ∼ 25 ns, ν = 10 Hz) was used for the irradiation of the MAPLE targets consisting of TiO2 nanoparticles (NPs) or mixtures of TiO2 NPs, Au NPs, and graphene oxide (GO) platelets in aqueous solutions. The effect of Au and GO addition as well as nitrogen doping on the photocatalytic activity of the TiO2 thin films was investigated. The evaluation of the photocatalytic activity was performed by photodegradation of the organic methylene blue model dye pollutant under UV-visible light, “simulated sun” irradiation conditions. Our results show that the photocatalytic properties of TiO2 were significantly improved by the addition of Au NPs and rGO platelets. Nitrogen inclusion into the rGO structure further contributes to the enhancement of the TiO2/Au/rGO nanocomposites photocatalytic activity.

Laser-assisted fabrication and non-invasive imaging of 3D cell-seeding constructs for bone tissue engineering

We report on laser-assisted fabrication and non-invasive imaging of porous 3D cell-seeding constructs (3D-CSCs) for bone tissue engineering. The 3D structures were built by two-photon polymerization-direct writing (2PP_DW) of IP-L780 photopolymer and consist in arrays of vertical microtubes arranged in triangular lattices. The microtubes were tightly, medium, and rarely packed, according to the constants of the triangular lattices of 8, 12, and 24 μm, respectively. The efficiency of the laser-generated 3D-CSCs for new bone formation was assessed in MG63 osteoblast-like cells cultures. High spatial resolution 3D images of the cell-seeded 3D-CSCs were obtained by digital holographic microscopy (DHM). The recorded holograms allowed the simultaneous evaluation of the 3D-CSCs and of the seeded cells, in terms of 3D shapes and dimensions, without intruding into the cells natural environment. The seeded cells, in particular the cells nuclei, conformed to the micro-architectures of the 3D-CSCs. Furthermore, the osteogenic potential of the 3D-CSCs was assessed in terms of cell morphology, viability, and level of mineralization. The microtubes packing density that allowed the seeded osteoblasts to reach the highest level of mineralization was established.

Combinatorial MAPLE deposition of antimicrobial orthopedic maps fabricated from chitosan and biomimetic apatite powders

Chitosan/biomimetic apatite thin films were grown in mild conditions of temperature and pressure by Combinatorial Matrix-Assisted Pulsed Laser Evaporation on Ti, Si or glass substrates. Compositional gradients were obtained by simultaneous laser vaporization of the two distinct material targets. A KrF* excimer (λ=248nm, τFWHM=25ns) laser source was used in all experiments. The nature and surface composition of deposited materials and the spatial distribution of constituents were studied by SEM, EDS, AFM, GIXRD, FTIR, micro-Raman, and XPS. The antimicrobial efficiency of the chitosan/biomimetic apatite layers against Staphylococcus aureus and Escherichia coli strains was interrogated by viable cell count assay. The obtained thin films were XRD amorphous and exhibited a morphology characteristic to the laser deposited structures composed of nanometric round shaped grains. The surface roughness has progressively increased with chitosan concentration. FTIR, EDS and XPS analyses indicated that the composition of the BmAp-CHT C-MAPLE composite films gradually modified from pure apatite to chitosan. The bioevaluation tests indicated that S. aureus biofilm is more susceptible to the action of chitosan-rich areas of the films, whilst the E. coli biofilm proved more sensible to areas containing less chitosan. The best compromise should therefore go, in our opinion, to zones with intermediate-to-high chitosan concentration which can assure a large spectrum of antimicrobial protection concomitantly with a significant enhancement of osseointegration, favored by the presence of biomimetic hydroxyapatite.

Electrically stimulated osteogenesis on Ti-PPy/PLGA constructs prepared by laser-assisted processes.

This work describes a versatile laser-based protocol for fabricating micro-patterned, electrically conductive titanium-polypyrrole/poly(lactic-co-glycolic)acid (Ti-PPy/PLGA) constructs for electrically stimulated (ES) osteogenesis. Ti supports were patterned using fs laser ablation in order to create high spatial resolution microstructures meant to provide mechanical resistance and physical cues for cell growth. Matrix Assisted Pulsed Laser Evaporation (MAPLE) was used to coat the patterned Ti supports with PPy/PLGA layers acting as biocompatible surfaces having chemical and electrical properties suitable for cell differentiation and mineralization. In vitro biological assays on osteoblast-like MG63 cells showed that the constructs maintained cell viability without cytotoxicity. At 24 h after cell seeding, electrical stimulation with currents of 200 μA was applied for 4 h. This treatment was shown to promote earlier onset of osteogenesis. More specifically, the alkaline phosphatase activity of the stimulated cultures reached the maximum before that of the non-stimulated ones, i.e. controls, indicating faster cell differentiation. Moreover, mineralization was found to occur at an earlier stage in the stimulated cultures, as compared to the controls, starting with Day 6 of cell culture. At later stages, calcium levels in the stimulated cultures were higher than those in control samples by about 70%, with Ca/P ratios similar to those of natural bone. In all, the laser-based protocol emerges as an efficient alternative to existing fabrication technologies.

3D Biomimetic Magnetic Structures for Static Magnetic Field Stimulation of Osteogenesis

We designed, fabricated and optimized 3D biomimetic magnetic structures that stimulate the osteogenesis in static magnetic fields. The structures were fabricated by direct laser writing via two-photon polymerization of IP-L780 photopolymer and were based on ellipsoidal, hexagonal units organized in a multilayered architecture. The magnetic activity of the structures was assured by coating with a thin layer of collagen-chitosan-hydroxyapatite-magnetic nanoparticles composite. In vitro experiments using MG-63 osteoblast-like cells for 3D structures with gradients of pore size helped us to find an optimum pore size between 20–40 µm. Starting from optimized 3D structures, we evaluated both qualitatively and quantitatively the effects of static magnetic fields of up to 250 mT on cell proliferation and differentiation, by ALP (alkaline phosphatase) production, Alizarin Red and osteocalcin secretion measurements. We demonstrated that the synergic effect of 3D structure optimization and static magnetic stimulation enhances the bone regeneration by a factor greater than 2 as compared with the same structure in the absence of a magnetic field.

Effect of the manufacturing parameters on the structure of nitrogen-doped carbon nanotubes produced by catalytic laser-induced chemical vapor deposition

Nitrogen-containing carbon nanotubes (CNx-NTs), with a relatively high level of nitrogen doping were prepared by the catalytic laser-induced CVD method. The nanotubes were catalytically grown directly on a silicon substrate from C2H2/NH3 gaseous precursors. X-ray photoelectron spectroscopy (XPS) and transmission electron microscopy (TEM) give firm evidence for the nitrogen doping. As determined by XPS, the N concentration for the prepared CNx-NTs increases from 3.6 to 30.6 at.% with increasing ammonia concentration and pressure. TEM images indicate that the nanotubes are bamboo like. As the nitrogen content increases, there is a transition from the bamboo shape with few defects and little distortion to a corrugated structure with a much larger number of defects. Raman spectroscopy revealed that with increasing nitrogen concentration, there is more disorder and defects, together with an increase in ID/IG ratio. By energy-filtering TEM, a higher N concentration was found on the outer amorphous nanolayer than in the compartment core of the nanotubes.

Development Of Magnetic Fe-C Nanocomposites Obtained Via The Laser Pyrolysis: Structural And Disaggregation Properties

Fe@C) nanoparticles have been successfully synthesized using the laser pyrolysis method and variable nozzle geometries. At large nozzle diameters, XRD and SAED analysis clearly identified distinct α‐Fe and Fe3C phases. TEM and HRTEM indicated that these Fe‐based nanoparticles have an average grain size of 3.5–10.2 nm. Temperature dependent Mossbauer spectra further confirm their distinct nanophases. By using a multi‐step reduction procedure, Fe@C powders can be disaggregated into stable, water soluble nanoparticles.