Experimental observation of tunable Wood type resonances in an all-ferrodielectric periodical metasurface.

In this Letter, a completely ferrodielectric metasurface consisting of an array of cylinders on a substrate is studied. All structural elements are made of ferrodielectric material. The conditions for the excitation of Wood's anomaly mode, obtained for different geometric parameters of the metasurface, are revealed. By continuously changing the structure parameters, we can change the position of the resonance at the Wood anomaly, thereby setting the position of the resonance at the frequency we need. It is shown that there is a resonant increase in the polarization plane rotation of the transmitted waves at the corresponding resonant frequency of the lattice mode excitation. Such polarization rotation is demonstrated both experimentally and theoretically.

Development of two-photon polymerised scaffolds for optical interrogation and neurite guidance of human iPSC-derived cortical neuronal networks.

Recent progress in the field of human induced pluripotent stem cells (iPSCs) has led to the efficient production of human neuronal cell models for in vitro study. This has the potential to enable the understanding of live human cellular and network function which is otherwise not possible. However, a major challenge is the generation of reproducible neural networks together with the ability to interrogate and record at the single cell level. A promising aid is the use of biomaterial scaffolds that would enable the development and guidance of neuronal networks in physiologically relevant architectures and dimensionality. The optimal scaffold material would need to be precisely fabricated with submicron resolution, be optically transparent, and biocompatible. Two-photon polymerisation (2PP) enables precise microfabrication of three-dimensional structures. In this study, we report the identification of two biomaterials that support the growth and differentiation of human iPSC-derived neural progenitors into functional neuronal networks. Furthermore, these materials can be patterned to induce alignment of neuronal processes and enable the optical interrogation of individual cells. 2PP scaffolds with tailored topographies therefore provide an effective method of producing defined in vitro human neural networks for application in influencing neurite guidance and complex network activity.

New microorganism isolation techniques with emphasis on laser printing.

The study of biodiversity, growth, development, and metabolism of cultivated microorganisms is an integral part of modern microbiological, biotechnological, and medical research. Such studies require the development of new methods of isolation, cultivation, manipulation, and study of individual bacterial cells and their consortia. To this end, in recent years, there has been an active development of different isolation and three-dimensional cell positioning methods. In this review, the optical tweezers, surface heterogeneous functionalization, multiphoton lithography, microfluidic techniques, and laser printing are reviewed. Laser printing is considered as one of the most promising techniques and is discussed in detail.

Laser printing of microbial systems: effect of absorbing metal film.

Recently, it was shown that laser-induced forward transfer (LIFT) technology and the laser engineering of microbial systems (LEMS) technique (based on LIFT method) are effective for isolation of micro-organisms from different complex substrates. These techniques frequently utilize Au as an absorbing layer material. The purpose of this study was to investigate the influence of absorbing film materials (Au, Ti and Cr) on the effectiveness of laser printing of micro-organisms to improve LEMS and LIFT techniques. It was shown that application of Ti and Cr absorbing layers activates bacterial growth after laser printing and is significantly more effective in comparison to Au films, which actually show a suppressing effect on bacterial cells. Results of this study can be applied for LEMS and LIFT protocols for improving bacterial isolation and microbial growth. SIGNIFICANCE AND IMPACT OF THE STUDY: Laser-induced forward transfer technique (LIFT) is currently used for printing of micro-organisms and in biosensor techniques, for single-cell isolation, and for culturing of micro-organisms from complex substrates. We have studied the influence of absorbing film materials (Au, Ti and Cr) on the effectiveness laser printing of micro-organisms. It was shown that application of Ti and Cr absorbing layers activates bacterial growth and is more effective in LIFT compared to Au films, which actually have a suppressive effect on bacteria cells. The results can improve LIFT protocols for bacteria isolation and culturing of microbial systems.

Multicomponent nanocrystals with anti-Stokes luminescence as contrast agents for modern imaging techniques.

Lanthanide-doped upconversion nanoparticles (UCNPs) have recently attracted great attention in theranostics due to their exceptional optical and physicochemical properties, which enable the design of a novel UCNP-based nanoplatform for luminescent imaging, temperature mapping, sensing, and therapy. In addition, UCNPs are considered to be ideal building blocks for development of multimodal probes for cells and whole body imaging, exploiting simple variation of host matrix, dopant ions, and surface chemistry. Modalities responsible for magnetic resonance imaging (MRI), computed tomography (CT), and positron emission tomography (PET)/single-photon emission computed tomography (SPECT) are embedded in a single UC nanocrystal, providing integrating effect over any modality alone in terms of the efficiency and sensitivity for clinical innovative diagnosis through multimodal bioimaging. In particular, we demonstrate applications of UCNPs as a new nanoplatform for optical and multimodal cancer imaging in vitro and in vivo and extend discussions to delivery of UCNP-based therapeutic agents for photodynamic and photothermal cancer treatments.

Surface micromorphology of cross-linked tetrafunctional polylactide scaffolds inducing vessel growth and bone formation.

In the presented study, we have developed a synthetic strategy allowing a gradual variation of a polylactide arms' length, which later influences the micromorphology of the scaffold surface, formed by a two-photon polymerization technique. It has been demonstrated that the highest number of cells is present on the scaffolds with the roughest surface made of the polylactide with longer arms (PLA760), and osteogenic differentiation of mesenchymal stem cells is most pronounced on such scaffolds. According to the results of biological testing, the PLA760 scaffolds were implanted into a created cranial defect in a mouse for an in vivo assessment of the bone tissue formation. The in vivo experiments have shown that, by week 10, deposition of calcium phosphate particles occurs in the scaffold at the defect site, as well as, the formation of a new bone and ingrowth of blood vessels from the surrounding tissues. These results demonstrate that the cross-linked microstructured tetrafunctional polylactide scaffolds are promising microstructures for bone regeneration in tissue engineering.

Formation of Neural Networks in 3D Scaffolds Fabricated by Means of Laser Microstereolithography.

We developed and tested new 3D scaffolds for neurotransplantation. Scaffolds of predetermined architectonic were prepared using microstereolithography technique. Scaffolds were highly biocompatible with the nervous tissue cells. In vitro studies showed that the material of fabricated scaffolds is not toxic for dissociated brain cells and promotes the formation of functional neural networks in the matrix. These results demonstrate the possibility of fabrication of tissue-engineering constructs for neurotransplantation based on created scaffolds.

EVALUATION OF VASCULOGENIC POTENTIAL OF MODIFIED FIBRIN HYDROGEL.

In recent years, engineering of blood vessels, which can provide the effective transport of nutrients and various metabolites, is one of the major challenges in tissue reconstruction. Many researches are carried out to develop cell-seeded bioconstructs based on natural polymers, particularly on PEGylated fibrin. Therefore, the aim of this study was to reveal the optimal component ratio for modified fibrin hydrogels in order to provide favorable conditions for vascular development of endothelial and mesenchymal stem cell co-culture. It has been found out that the PEGylated fibrin hydrogels can support 3D cell growth in HUVECs and hASCs co-culture. The microporous filamentous hydrogel prepared from PEGylated 5 : 1 fibrinogen and using the 1 : 0.2 protein to thrombin ratio had the most favorable microenvironment for cell distribution, growth and development in the studied co-culture that resulted in high levels of expression of proteins required for angiogenesis.

[Compatibility of cells of the nervous system with structured biodegradable chitosan-based hydrogel matrices].

Hydrogel matrices for cell cultivation have been generated by two-photon laser polymerization of unsaturated chitosan derivatives and methacrylated hyaluronic acid. The adhesive and toxic properties of the matrices have been assessed, and the matrices have been shown to have a good compatibility with primary hippocampal cell cultures. The formation of morphologically normal neural networks by cells of the nervous system cultured on the surface of hydrogel matrices has been observed. The metabolic status of dissociated hippocampal cells cultured on the matrices was similar to that of the control cultures, as shown by the results of MTT reductase activity assay. Thus, matrices based on unsaturated polysaccharide derivatives crosslinked by laser irradiation showed good compatibility with differentiated cells of the nervous system and considerable potential for use in neurotransplantation.

[Implant Design by Means of Multiphoton Polymerization]. / Implantatdesign mittels Multiphotonen-Polymerisation.

BACKGROUND: Additive manufacturing and 3D printing create new paths for the design and manufacturing of implants. Technologies with high resolution are required for the development of microstructured eye implants. In the present study, we demonstrate how these technologies can be used during the design development and manufacturing of a multifocal diffractive aspheric intraocular lens. MATERIAL AND METHODS: Multiphoton polymerisation (MPP) is used to manufacture a diffractive relief with resolution in the sub-micrometer range. The relief is applied to the moulded body of a refractive lens, forming a trifocal lens. Propagation of light behind the lens is visualised in water with fluorescein. RESULTS: Multifocal lenses were successfully manufactured with this approach. The optical design with three foci is confirmed by the light propagation images. The images even clearly demonstrate the impact of the refractive and diffractive elements and may provide information on artefacts and aberrations. CONCLUSIONS: Multiphoton polymerisation is an interesting tool for the flexible manufacturing of complex multifocal lenses. With future technological progress in 3D printing with MPP, this is a promising method for on-demand manufacturing of patient individual intraocular lenses.

[Optimisation of the visualisation technique for optical paths through intraocular lenses for characterisation of multifocal imaging properties of Fresnel-zone plates]. / Optimierung der Visualisierungstechnik für Strahlenverläufe durch Intraokularlinsen zur Charakterisierung multifokaler Abbildungseigenschaften von Fresnel-Zonenplatten.

The utilisation of the diffractive properties of Fresnel zone plates offers the possibility of intraocular lens designs with multiple foci. Such intraocular lenses can be manufactured by two-photon polymerisation (2PP). This paper explains the underlying concept and shows the principles for visualisation of the focus properties of such implants.

[New concepts for pressure-controlled glaucoma implants]. / Neue Konzepte für druckgesteuerte Glaukomimplantate.

In industrialized countries glaucoma is one of the most common causes that leads to blindness. It is also the most common cause of irreversible blindness worldwide. In addition to local treatment of intraocular pressure and filtering glaucoma surgery, alloplastic implants are increasingly being used in glaucoma therapy. As long-term results published in the literature of commonly used implants are unsatisfactory, it seems useful to search for new concepts. In order to avoid the well-known short-term and long-term postoperative complications a pressure-controlled microstent with antiproliferative surface modifications was developed. Additionally, the functionality of such a microstent should be investigated using an animal glaucoma model. This paper describes the concept of a microstent which drains aquous humour from the anterior chamber into the suprachoroidal space. In addition, the glaucoma models described in the literature are discussed. Unfortunately, none of the methods could be reproduced permanently. First results show a correct implantation of a coated microstent with valve where the anti-proliferative effect could be demonstrated histologically. The promising results should lead to further investigations and the final goal will be the testing of the stent in the human eye.

Laser assisted cell printing.

Computer assisted biofabrication of fully functional living tissue for regenerative medicine involves generation of complex three-dimensional constructs consisting of living cells and biomaterials. Laser BioPrinting (LaBP) based on laser-induced forward-transfer provides unique possibilities for the deposition of different living cells and biomaterials in a well-defined 3D structure. LaBP can be applied to generate scaffold-free 3D cell systems through a layer-by-layer technique by combining cell solutions with materials that are able to form stable gels. Also, it is used to precisely populate scaffolds with different cells and different cell densities. It was proven that printed cells are not affected by the laser printing procedure and that a differentiation of printed stem cells is not induced. Thus, LaBP is demonstrated as a promising tool for the ex vivo generation of tissue replacements.

Air-directed attachment of coccoid bacteria to the surface of superhydrophobic lotus-like titanium.

Superhydrophobic titanium surfaces fabricated by femtosecond laser ablation to mimic the structure of lotus leaves were assessed for their ability to retain coccoid bacteria. Staphylococcus aureus CIP 65.8T, S. aureus ATCC 25923, S. epidermidis ATCC 14990T and Planococcus maritimus KMM 3738 were retained by the surface, to varying degrees. However, each strain was found to preferentially attach to the crevices located between the microscale surface features. The upper regions of the microscale features remained essentially cell-free. It was hypothesised that air entrapped by the topographical features inhibited contact between the cells and the titanium substratum. Synchrotron SAXS revealed that even after immersion for 50 min, nano-sized air bubbles covered 45% of the titanium surface. After 1 h the number of cells of S. aureus CIP 65.8T attached to the lotus-like titanium increased to 1.27×10(5) mm(-2), coinciding with the replacement of trapped air by the incubation medium.

Two-photon polymerization-generated and micromolding-replicated 3D scaffolds for peripheral neural tissue engineering applications.

In this study, we explore the production of well-defined macroscopic scaffolds with two-photon polymerization (2PP) and their use as neural tissue engineering scaffolds. We also demonstrate that these 3D scaffolds can be replicated via soft lithography, which increases production efficiency. Photopolymerizable polylactic acid (PLA) was used to produce scaffolds by 2PP and soft lithography. We assessed the biocompatibility of these scaffolds using an SH-SY5Y human neuronal cell line and primary cultured rat Schwann cells (of direct relevance to the repair of nerve injuries). A Comet assay with SH-SY5Y human neuronal cells revealed minimal DNA damage after washing the photocured material for 7 days in ethanol. Additionally, thin films and 3D scaffolds of the photocured PLA sustained a high degree of Schwann cell purity (99%), enabled proliferation over 7 days and provided a suitable substrate for supporting Schwann cell adhesion such that bi-polar and tri-polar morphologies were observed. Evidence of orthogonally aligned and organized actin thin filaments and the formation of focal contacts were observed for the majority of Schwann cells. In summary, this work supports the use of PLA as a suitable material for supporting Schwann cell growth and in turn use of 3D soft lithography for the synthesis of neural scaffolds in nerve repair.

Adipogenic differentiation of laser-printed 3D tissue grafts consisting of human adipose-derived stem cells.

Laser-assisted bioprinting (LaBP) allows the realization of computer-generated 3D tissue grafts consisting of cells embedded in a hydrogel environment. In this study, human adipose-derived stem cells (hASCs) were printed in a free-scalable 3D grid pattern by means of LaBP. We demonstrate that neither the proliferation ability nor the differentiation behaviour of the stem cells was affected by the LaBP procedure. Furthermore, the 3D grafts were differentiated down the adipogenic lineage pathway for 10 days. We verify by quantitative assessments of adipogenic markers that the 3D grafts resemble cell lineages present in natural adipose tissue. Additionally, we provide the proof that even pre-differentiated hASCs could be utilized for the generation of 3D tissue grafts. These results indicate that the biofabrication of living grafts resembling their complex native origin is within reach.

Three-dimensional laser micro- and nano-structuring of acrylated poly(ethylene glycol) materials and evaluation of their cytoxicity for tissue engineering applications.

The natural cell environment is characterized by complex three-dimensional structures, which contain features at multiple length scales. Many in vitro studies of cell behavior in three dimensions rely on the availability of artificial scaffolds with controlled three-dimensional topologies. In this paper, we demonstrate fabrication of three-dimensional scaffolds for tissue engineering out of poly(ethylene glycol) diacrylate (PEGda) materials by means of two-photon polymerization (2PP). This laser nanostructuring approach offers unique possibilities for rapid manufacturing of three-dimensional structures with arbitrary geometries. The spatial resolution dependence on the applied irradiation parameters is investigated for two PEGda formulations, which are characterized by molecular weights of 302 and 742. We demonstrate that minimum feature sizes of 200nm are obtained in both materials. In addition, an extensive study of the cytotoxicity of the material formulations with respect to photoinitiator type and photoinitiator concentration is undertaken. Aqueous extracts from photopolymerized PEGda samples indicate the presence of water-soluble molecules, which are toxic to fibroblasts. It is shown that sample aging in aqueous medium reduces the cytotoxicity of these extracts; this mechanism provides a route for biomedical applications of structures generated by 2PP microfabrication and photopolymerization technologies in general. Finally, a fully biocompatible combination of PEGda and a photoinitiator is identified. Fabrication of reproducible scaffold structures is very important for systematic investigation of cellular processes in three dimensions and for better understanding of in vitro tissue formation. The results of this work suggest that 2PP may be used to polymerize poly(ethylene glycol)-based materials into three-dimensional structures with well-defined geometries that mimic the physical and biological properties of native cell environments.