First in vitro cell co-culture experiments using laser-induced high-energy electron FLASH irradiation for the development of anti-cancer therapeutic strategies.

Radiation delivery at ultrahigh dose rates (UHDRs) has potential for use as a new anticancer therapeutic strategy. The FLASH effect induced by UHDR irradiation has been shown to maintain antitumour efficacy with a reduction in normal tissue toxicity; however, the FLASH effect has been difficult to demonstrate in vitro. The objective to demonstrate the FLASH effect in vitro is challenging, aiming to reveal a differential response between cancer and normal cells to further identify cell molecular mechanisms. New high-intensity petawatt laser-driven accelerators can deliver very high-energy electrons (VHEEs) at dose rates as high as 1013 Gy/s in very short pulses (10-13 s). Here, we present the first in vitro experiments carried out on cancer cells and normal non-transformed cells concurrently exposed to laser-plasma accelerated (LPA) electrons. Specifically, melanoma cancer cells and normal melanocyte co-cultures grown on chamber slides were simultaneously irradiated with LPA electrons. A non-uniform dose distribution on the cell cultures was revealed by Gafchromic films placed behind the chamber slide supporting the cells. In parallel experiments, cell co-cultures were exposed to pulsed X-ray irradiation, which served as positive controls for radiation-induced nuclear DNA double-strand breaks. By measuring the impact on discrete areas of the cell monolayers, the greatest proportion of the damaged DNA-containing nuclei was attained by the LPA electrons at a cumulative dose one order of magnitude lower than the dose obtained by pulsed X-ray irradiation. Interestingly, in certain discrete areas, we observed that LPA electron exposure had a different effect on the DNA damage in healthy normal human epidermal melanocyte (NHEM) cells than in A375 melanoma cells; here, the normal cells were less affected by the LPA exposure than cancer cells. This result is the first in vitro demonstration of a differential response of tumour and normal cells exposed to FLASH irradiation and may contribute to the development of new cell culture strategies to explore fundamental understanding of FLASH-induced cell effect.

Fs Laser Patterning of Amorphous As2S3 Thin Films.

This study investigates the morphological changes induced by femtosecond (fs) laser pulses in arsenic trisulfide (As2S3) thin films and gold-arsenic trisulfide (Au\As2S3) heterostructures, grown by pulsed laser deposition (PLD). By means of a direct laser writing experimental setup, the films were systematically irradiated at various laser power and irradiation times to observe their effects on surface modifications. AFM was employed for morphological and topological characterization. Our results reveal a clear transition threshold between photoexpansion and photoevaporation phenomena under different femtosecond laser power regimes, occurring between 1 and 1.5 mW, irrespective of exposure time. Notably, the presence of a gold layer in the heterostructure minimally influenced this threshold. A maximum photoexpansion of 5.2% was obtained in As2S3 films, while the Au\As2S3 heterostructure exhibited a peak photoexpansion of 0.8%. The study also includes a comparative analysis of continuous-wave (cw) laser irradiation, confirming the efficiency of fs laser pulses in inducing photoexpansion effects.

Laser fabrication of diffractive optical elements based on detour-phase computer-generated holograms for two-dimensional Airy beams.

We have designed, fabricated, and tested an amplitude diffractive optical element for generation of two-dimensional (2D) Airy beams. The design is based on a detour-phase computer-generated hologram. Using laser ablation of metallic films, we obtained a 2 mm×2 mm diffractive optical element with a pixel of 5 µm×5 µm and demonstrated a fast, cheap, and reliable fabrication process. This device can modulate 2D Airy beams or it can be used as a UV lithography mask to fabricate a series of phase holograms for higher energy efficiency. Tests according to the premise and an analysis of the transverse profile and propagation are presented.

Laser-direct writing by two-photon polymerization of 3D honeycomb-like structures for bone regeneration.

A major limitation of existing 3D implantable structures for bone tissue engineering is that most of the cells rapidly attach on the outer edges of the structure, restricting the cells penetration into the inner parts and causing the formation of a necrotic core. Furthermore, these structures generally possess a random spatial arrangement and do not preserve the isotropy on the whole volume. Here, we report on the fabrication and testing of an innovative 3D hierarchical, honeycomb-like structure (HS), with reproducible and isotropic arhitecture, that allows in 'volume' migration of osteoblasts. In particular, we demonstrate the possibility to control the 3D spatial cells growth inside these complex architectures by adjusting the free spaces inside the structures. The structures were made of vertical microtubes arranged in a mulitlayered configuration, fabricated via laser direct writing by two photons polymerization of the IP-L780 photopolymer. In vitro tests performed in MG-63 osteoblast-like cells demonstrated that the cells migration inside the 3D structures is conducted by the separation space between the microtubes layers. Specifically, for layers separation between 2 and 10 µm, the cells gradually penetrated between the microtubes. Furthermore, these structures induced the strongest cells osteogenic differentiation and mineralization, with ALP activity 1.5 times stronger, amount of calcified minerals 1.3 times higher and osteocalcin secretion increased by 2.3 times compared to the other structures. On the opposite, for layers separation less than 2 µm and above 10 µm, the cells were not able to make interconnections and exhibited poor mineralization ability.

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.

Diode-laser pumping into the emitting level for efficient lasing of depressed cladding waveguides realized in Nd:YVO4 by the direct femtosecond-laser writing technique.

Depressed cladding waveguides have been realized in Nd:YVO(4) employing direct writing technique with a femtosecond-laser beam. It was shown that the output performances of such laser devices are improved by the reduction of the quantum defect between the pump wavelength and the laser wavelength. Thus, under the classical pump at 808 nm (i.e. into the (4)F(5/2) level), a 100-µm diameter circular waveguide inscribed in a 0.7-at.% Nd:YVO(4) outputted 1.06-µm laser pulses with 3.0-mJ energy, at 0.30 optical efficiency and slope efficiency of 0.32. The pump at 880 nm (i.e.directly into the (4)F(3/2) emitting level) increased the pulse energy at 3.8 mJ and improved both optical efficiency and slope efficiency at 0.36 and 0.39, respectively. The same waveguide yielded continuous-wave 1.5-W output power at 1.06 µm under the pump at 880 nm. Laser emission at 1.34 µm was also improved using the pump into the (4)F(3/2) emitting level of Nd:YVO(4).

Laser parallel nanofabrication by single femtosecond pulse near-field ablation using photoresist masks.

A new near-field processing method by femtosecond laser ablation using photoresist enhancing masks is numerically and experimentally investigated. Periodical structures with 2 µm pitch, 1 µm width and 300 nm height, created in polymethyl methacrylate photoresist by e-beam lithography, were used to intensify the incident laser radiation. The near-field distribution and the intensification factor of the optical radiation were computed using the Finite-Difference-Time-Domain numerical simulations. The pattern of the photoresist mask was imprinted on the surface of a silicon wafer. Using a single infrared femtosecond laser pulse, uniform and continuum grooves with the width in the range of 250 nm were obtained on large silicon surface.

Laser emission from diode-pumped Nd:YAG ceramic waveguide lasers realized by direct femtosecond-laser writing technique.

We report on realization of buried waveguides in Nd:YAG ceramic media by direct femtosecond-laser writing technique and investigate the waveguides laser emission characteristics under the pump with fiber-coupled diode lasers. Laser pulses at 1.06 µm with energy of 2.8 mJ for the pump with pulses of 13.1-mJ energy and continuous-wave output power of 0.49 W with overall optical efficiency of 0.13 were obtained from a 100-µm diameter circular cladding waveguide realized in a 0.7-at.% Nd:YAG ceramic. A circular waveguide of 50-µm diameter yielded laser pulses at 1.3 µm with 1.2-mJ energy.

One- and two-photon induced emission in heterobimetallic Zn(II)-Sm(III) and Zn(II)-Tb(III) complexes with a side-off compartmental ligand.

Heterobimetallic [Zn(II)Ln(III)] complexes have been obtained using a compartmental Schiff-base ligand, H(2)valdmpn, resulting from the 2:1 condensation between o-vanillin and 2,2-dimethyl-propilenediamine: [Zn(H(2)O)(valdmpn)Sm(O(2)NO)(3)] 1, [Zn(H(2)O)(valdmpn)Tb(O(2)NO)(3)] 2a, [Zn(H(2)O)(valdmpn)Tb(O(2)NO)(3)]·H(2)O 2b, and [Zn(H(2)O)(valdmpn)Gd(O(2)NO)(3)]·H(2)O 3. The crystal structures of 1, 2b, and 3 have been solved. Compounds 1 and 2a crystallize in a non-centrosymmetric space group (P2(1)2(1)2(1)), being isomorphous. Crystals 2b and 3 are also isomorphous (space group P1[combining macron]). The complex entities in the four crystals are similar and their structures consist of binuclear species with the pentacoordinated zinc(II) ion hosted into the N(2)O(2) compartment and the lanthanide(III) ion in the large, open compartment, with a coordination number of 10. The photophysical properties of the four compounds have been investigated. Strong visible excited (excitation tails extend up to 420-430 nm) one photon antenna sensitization was obtained with the samarium(III) and terbium(III) derivatives. Following femtosecond Ti:Sapphire laser at λ(ex) = 775 nm, both second-harmonic generation at λ(em) = 775/2 nm and two-photon induced emission in the VIS range were obtained, extending thus the excitation range of these complexes from the VIS to the NIR spectral range. The two-photon induced emission and second harmonic generation effect for a samarium(III) complex are reported for the first time.