Bioinspired and Biointegrated Materials as Frontiers Nanomaterials V

Resume : Topography, the physical characteristics of an environment, is one of the most prominent stimuli neurons can encounter in the body. Many aspects of neurons and neuronal behavior are affected by the size, shape, and pattern of the physical features of the environment. A recent increase in the use of nanometric topographies, due to improved fabrication techniques, has resulted in new findings on neuronal behavior and development. Factors such as neuron adhesion, neurite alignment, and even the rate of neurite formation have all been highlighted through nanotopographies as complex phenomena that are driven by intricate intracellular mechanisms. The translation of physical cues is a biologically complex process thought to begin with recognition by membrane receptors as well as physical, cell-to-surface interactions, but the internal biological pathways that follow are still unclear. In this respect, nanotopography would be a more suitable platform on which to study receptor interfaces than microtopography because of the subcellular topographical features that are relevant in scale to the receptor activity. Ultimately, the characterization of this unknown network of pathways will unveil many aspects of the behavior and intracellular processes of neurons, and play an important role in the manipulation of neuronal development for applications in neural circuits, neuroregenerative medicine and prostheses, and much more.

Resume : Cells explore the surfaces of materials through membrane-bound receptors, such as the integrins, and use them to interact with extracellular matrix molecules adsorbed on the substrate surfaces, resulting in the formation of focal adhesions. With recent advances in nanotechnology, biosensors and bioelectronics are being fabricated with ever decreasing feature sizes. The performances of these devices depend on how cells interact with nanostructures on the device surfaces. However, the behavior of cells on nanostructures is not yet fully understood. Here we present a systematic study of cell-nanostructure interaction using polymeric nanopillars with various diameters. We also report the development of a well-defined stem cell culture system using surface functionalized polymeric nanopillars with tunable surface chemical and physical properties. Nanopillars, modified with peptides derived from vitronectin, can be used as defined substrates to maintain both human embryonic stem (hES) cells and human induced pluripotent stem (hiPS) cells for up to 3 months without differentiation. The surface chemical properties of the nanopillars were more critical than the physical properties in maintaining stem cells. However, the physical properties of the nanopillars influenced the stem cell fate. Softer nanopillars promoted the undifferentiated state, and more rigid nanopillars encouraged stem cell differentiation. Such nanopillar system also allows us to selectively sorting specific cells n

Resume : In this presentation, we will discuss our recent developments on the use of functional nanoparitcles in biomedical applications. We will mainly focus on the interactions of colloidal gold nanoparticles and endothelial cells. Endothelial cells are the major players in a vital biological process-the angiogenesis. Angiogenesis is the growth of new capillary vessels to bring oxygen and nutrients at the most distant areas of the body. Manipulation of this process could be the key to the development of several diseases including cancer. We will show our recent efforts towards the manipulation of angiogenesis using bio-functionalized nanoparticles.

Resume : Temperature-responsive cell culture surfaces, which is poly(N-isopropylacrylamide) (PIPAAm) modified tissue-culture polystyrene (TCPS) (PIPAAm-TCPS), has been used for the fabrication of various types of single monolayer cells sheet such as epithelium keratinocyte, corneal epithelial cell, oral mucosal epithelial cell, urothelial cell, periodontal ligament, cardiomyocyte, hepatocyte cell sheets, and so on. Various types of cell sheets have been directly transplanted to reconstruct tissue, and some of them are clinically applied. PIPAAm-TCPS has been conventionally prepared by using electron beam (EB) irradiation and is currently released as commercially available products. Although PIPAAm-TCPS has been elaborately characterized with FT-IR, XPS, AFM and cell detachment / attachment assay, the interaction between cells and PIPAAm-TCPS surface was not quantitatively estimated. To quantitatively estimate the interaction, we have newly prepared PIPAAm-TCPS having five parallel micro-channels for cell culture by using micro-fabrication technology. This construction allows concurrently generating five different shear forces to apply to cells in individual microchannels having various resistance of each channel and simultaneously gives an identical cell incubation condition to all test channels. Shear stress-dependent cell detachment process from PIPAAm-TCPS was evaluated at various shear stresses.　　EB irradiation method allows for mass-production of conventional PIPAAm-TCPS as a commercially-available product. However, the EB irradiation method is costly for the preparation of PIPAAm-TCPS. A facile method for the mass-production has been alternatively required to reduce the cost of mass-production without expensive and special EB irradiation system. In this presentation, we will show microfluidic-based analytical system for evaluating the interaction between cell and PIAPAm-TCPS surface and a facile method for the preparation of PIPAAm-TCPS.

Resume : Machine technology frequently puts magnetic or electrostatic repulsive forces to practical use, as in maglev trains, vehicle suspensions or non-contact bearings. In contrast, materials design overwhelmingly focuses on attractive interactions, such as in the many advanced polymer-based composites, where inorganic fillers interact with a polymer matrix to improve mechanical properties. However, articular cartilage strikingly illustrates how electrostatic repulsion can be harnessed to achieve unparalleled functional efficiency: it permits virtually frictionless mechanical motion within joints, even under high compression. Here we describe a composite hydrogel with anisotropic mechanical properties dominated by electrostatic repulsion between negatively charged unilamellar titanate nanosheets embedded within it. Crucial to the behaviour of this hydrogel is the serendipitous discovery of cofacial nanosheet alignment in aqueous colloidal dispersions subjected to a strong magnetic field, which maximizes electrostatic repulsion and thereby induces a quasi-crystalline structural ordering over macroscopic length scales and with uniformly large face-to-face nanosheet separation. We fix this transiently induced structural order by transforming the dispersion into a hydrogel using light-triggered in situ vinyl polymerization. The resultant hydrogel, containing charged inorganic structures that align cofacially in a magnetic flux, deforms easily under shear forces applied parallel to the embedded nanosheets yet resists compressive forces applied orthogonally. We anticipate that the concept of embedding anisotropic repulsive electrostatics within a composite material, inspired by articular cartilage, will open up new possibilities for developing soft materials with unusual functions.

Resume : Various calcium phosphate ceramics have been examined as scaffold materials for osteoblastic cells in bone tissue engineering. Octacalcium phosphate crystals, if synthesized in a specified condition, enhance mouse bone marrow derived stromal ST-2 cells inoculated on the surface, through increasing phosphorylation of p38 in the eosteoblastic signaling [1]. The change of inorganic ion composition around the crystals, caused by the conversion from OCP to hydroxyapatite (HA) crystals, could be one of factors involved in stimulating the osteoblastic cellular activity [2]. The physicochemical process during the material maturation into the thermodynamically stable phase (HA) could induce the surface morphological changes at nano-meter scale on the OCP material [3]. When OCP is mechanically mixed with amorphous calcium phosphate (ACP), which is a calcium phosphate salt soluble more than OCP, the conversion rate into HA from OCP increases with the modification of surface morphology of OCP [3]. Furthermore, the osteoconductivity of the mixture increases compared to the respective single phase if examined by its implantation into critical-sized rat calvaria defect [3]. Thus, it seems likely that calcium phosphate materials show the bioactivity through physicochemical process under physiological environment. [1] Nishikawa R et al. Dent Mater J 33:242 (2014); [2] Suzuki O et al. Biomaterials 27:2671 (2006); [3] Kobayashi K et al. ACS Appl Mater Interfaces 6:22602 (2014).

Resume : Hepatocyte sheet-based tissue engineering is an attractive method for the treatment of liver diseases. However, hepatocytes rapidly lose their viability and phenotypic functions on isolation from the native in vivo microenvironment of the liver. In this paper, heparin-functionalized poly(N-isopropylacrylamide) (PIPAAm)-grafted cell culture surface, which interacts with heparin-binding proteins such as heparin-binding EGF-like growth factor (HB-EGF), has been designed for maintaining hepatic functions during the cultivation. Heparin-functionalized thermoresponsive cell-culture surface induced the multivalent affinity binding of heparin-binding proteins in active form in the state of shrunken PIPAAm chains at 37 °C. By lowering temperature to 20 °C, the affinity binding between heparin-binding proteins and immobilized heparin was reduced with increasing the mobility of heparin and the swollen PIPAAm chains. When the medium contained less than 10 ng/cm2 of soluble HB-EGF, the hepatocytes were not able to adhere and form their cell sheets. Hepatocytes adhered well and formed their sheets on HB-EGF-bound heparin-functionalized thermoresponsive surface. The secretion of albumin was maintained and higher compared to that on PIPAAm-grafted surfaces with soluble HB-EGF. By lowering temperature to 20 °C, the cultured cell sheets were detached from the surface. The function of hepatic cell sheet was maintained by using advanced thermoresponsive cell culture surfaces.

Resume : Research in a new field, Hyper Bio Assembler for 3D Cellular Innovation (Bio Assembler) started in July 2011 as a five year project supported by Grants-in-Aid for Scientific Research on Innovative Areas from Japanese Ministry of Education (MEXT). The aim of project is to build 3D cellular systems capable of functioning in vitro and to propose an entirely new interdisciplinary area never previously explored. The Bio Assembler will elucidate the principles of ultrahigh speed measurement as well as the manipulation techniques and tissue function expression. The project consists of three research groups; first, ?Measurement and control of cell characteristics? which focuses on measuring physical properties of cells taken from living organisms at high speed and separating target cells useful in constructing cellular systems; second, ?3D cellular system assembly? which aims at shaping and assembling 3D cellular systems with complex morphologies and vascular networks inside; third, ?Analysis and evaluation of 3D cellular systems? which provides analysis and evaluation of growth, differentiation-inducing, morphogenesis controls and transplantation responses of created 3D cellular systems, and conducts functional elucidation as well as comparison with and verification of in vivo with looking for ways to bring our outcomes into the tissue life science such as regenerative medicine, cell assay, and so on. Our final goal is to establish three major academic achievements; ?Cell Sort Engineering?, ?3D Cellular System Design Logic? and ?Cell Sociology? as well as providing innovative measurement and control technologies based on micro/nano robotics.

Resume : A spherical multicellular aggregate can be used as an in vitro model for the 3D cellular construct in tissue engineering and tumors. Oxygen is one of the most important factors for cell survival and growth in three-dimensional culture. The increase in size of spheroid results in a hypoxia and causes cell necrosis in the core of large spheroids. We have developed an oxygen-permeable spheroid culture device using polydimethylsiloxane (PDMS) to prevent oxygen deprivation of the spheroid. In this study, we investigated whether this chip has advantages in the cell proliferation, viability, cellular function, and differentiation in comparison with the conventional non-oxygen-permeable chip. We found that the oxygen-permeable chip is useful for engineering 3D cellular constructs with high viability and functionality for tissue engineering.

Resume : Temperature-responsive cell culture surface (TRCS), poly(N-isopropylacrylamide) (PIPAAm) grafted tissue culture polystyrene (TCPS) (PIPAAm-TCPS) prepared by electron beam (EB) irradiation, has been used to fabricate various cells sheets. The fabricated cell sheets has been successfully transplanted to treat the damaged human tissues such as corneal, esophageal ulcerations, periodontitis and so on. Therefore, a lot of researchers are desiring for TRCS to research cell-sheet based tissue regeneration and/or to readily fabricate various tissues used for diagnosis. However, researchers in average laboratory can not prepare the TRCS because EB irradiation system is much expensive and not common equipment. Facile methods for preparing TRCS without EB system have been required. Based on the requirement, we newly developed a facile method for perpetrating TRCS without EB equipment. Advantage of the facile method is that commercially available polystyrene (PSt) surfaces is readily modified with thioxanthone based photo-initiator. In addition, PIPAAm was grafted on the photo-initiator modified PS surface by using visible light irradiation. These procedures were performed by commonly used chemical reaction and equipment. Researchers in the average laboratory also can readily prepare TRCS by this facile method as well as prepare polyacrylamide gel for the electrophoresis.

Resume : Microvasculatures are greatly involved in the disease such as inflammation, angiogenesis and cancer metastasis, and these events are highly complex, thus sophisticated model is required to understand these mechanism actions at cellular and tissue levels. Recent microtechnologies allow fabrication of three-dimensional (3D) microtissue structures and have a higher capability to control their microenvironments precisely. Since microvasculature is multicellular tissues and also microvascular disease is highly related to the chemical and mechanical factors such as extracellular matrices, shear stress and pressure, in vitro models enable analyzing various interactions from simple to complex state will be useful. Here, we introduce our 3D microvasculature models fabricated by a micro-molding method. Briefly, we prepared collagen microchannels by using collagen gels and PDMS hosting chamber including a needle (120 µm in diameter) as a channel mold. By using the device, continuous lumen structure of human umbilical vascular endothelial cells (HUVECs) surrounded by collagen gels can be formed. Our vasculature model allows continuous monitoring of the vascular permeability and sprouting responding to the angiogenic factors. Compared to the traditional 2D culture system, our model is very useful for the investigation of the vascular disease at both cellular and tissue levels.

Resume : Microfluidic technologies are capable of producing functional, micrometer-sized materials including particles, fibers, and sheets, composed of multiple small compartments with different physicochemical properties. In this presentation we briefly introduce our recent achievements on the preparation of anisotropic hydrogel materials and their applications to the construction of in vivo tissue-mimicking microenvironments. Hydrogel microfibers having anisotropic cross sections were prepared by utilizing the stable laminar flow in microfluidic channels, which were employed for tissue engineering applications including the guidance of cell proliferation, formation of heterotypic hepatic micro-organoids, and analysis of cancer cell migration in 3D environments. In addition, unique hydrogel micro-materials were produced such as stripe-patterned heterogeneous hydrogel sheets, yarn-ball-shaped hydrogel beads, and single micrometer-sized ECM beads. These results clearly demonstrated the usefulness of microfluidic technologies for preparing micrometer-sized, cell-encapsulating and/or cell-adhesive hydrogels.

Resume : Electrically conducting polymers (ECPs) were widely used to electrically couple with cells in implanted electronic devices and biosensors, mainly thanks to the long-term stability of their electrical properties in biological environments and their mechanical properties comparable with that of the extracellular matrix. To erase/diminish nonspecific interaction for ECPs would not only increase the selectivity of biosensors but also reduce the inflammation risk for implants. In this presentation, we show that with finely tuning the chemical structure, ECPs could specifically interact with cells on a protein-resistance background. Our recent works further demonstrated that with combining specific interaction with protein resistance on ECPs, they could differentiate cells, dynamically interact with and release cells, or even help to construct a three-dimensional platform to capture CTC cells with both high efficiency and selectivity.

Resume : LONGLIFE (“Advanced multifunctional zirconia ceramics for long-lasting implants”) coordinated by Prof. Jérôme Chevalier from INSA of Lyon, is a Collaborative European project in the frame of the 7th Framework Program. LONGLIFE aims at developing new zirconia-based oral and spine (lumbar inter-vertebral disc) implants, characterized by lifetime longer than 60 years and superior reliability as compared to previous biomedical devices. In order to reach this ambitious goal, efforts should be dedicated to improve the in-vivo stability of zirconia, by one side, and enhance the osseointegration capabilities of the implants in contact with bone, on the other. At the same time, the project aims at developing new ceramic-oriented designs for the implants as well as new multi-physic accelerated testing in vitro, able to reproduce more effectively the different degradation mechanisms and their interplay. In this work, the achievements concerning the design and development of new zirconia-based materials are presented, reached thanks to the synergies between Politecnico of Torino and INSA of Lyon. Among the technical oxide ceramics able to be used as structural biomaterials, zirconia-based ceramics are considered the best choice thanks to their excellent mechanical properties, biocompatibility and aesthetics. However, the current zirconia-based implants suffer from degradation under hydrothermal conditions (low temperature and presence of water), leading to surface degradation and possibly to fracture. Therefore, in order to increase the implants reliability, a strong effort is required to improve the zirconia stability in the presence of water, without decreasing its toughness and strength. The strategy chosen in LONGLIFE towards this goal is reported. Besides an innovative design of the composite material, a controlled synthesis process was used to produce multi-phase composites, having controlled architecture and composition. The results achieved demonstrate the effectiveness of this strategy towards the development of strong, tough and stable materials, fulfilling the objectives of the project.

Resume : This presentation will focus on the unprecedented impact nanotechnology has had across all of medicine, specifically focusing on improved disease prevention, diagnosis, and treatment. Disease systems will cover cancer, infection, inflammation, and poor tissue growth including bone, vascular, cardiovascular, bladder, nervous system and others. A particular focus will be placed on sensors that personalize medicine towards one's immune system. Toxicity will also be covered. A new field, picomedicine, will also be introduced concerning how further advances in medicine can be made. Here, the focus will be placed on controlling electron distributions (as opposed to atomic distribution in nanotechnology) to improve disease prevention, diagnosis, and treatment.

Resume : Molecular programming (also known as DNA computing) in its various implementations make it possible to implement complex instructions at the molecular scale. One of the DNA framework for molecular programming, the PEN framework developed by Y. Rondelez et al., provides a robust basis on which one can develop DNA based dynamic systems. It relies on a combination of three enzymes (a Polymerase, an Exonuclease and a Nickase) working on 22bp long DNA “templates” which encode relationships between 11bp strands. However the current communication paradigm is grounded in a classical molecular biology approach: inputting information with pipettes and reading states using fluorescence in a real time PCR machine. Such an approach is hard to miniaturize and parallelize and thus limits what can be done with this DNA framework. We report here our work done to use active surfaces for DNA computing. In fact, by working in microfluidics it is possible to benefit from an advantageous surface/volume ratio. This in turn makes it possible to design active surfaces which can release or expose sufficiently large quantity of DNA strand to effectively action our DNA dynamical system. We design these systems to be electrically driven to facilitate interfacing with classical computers. Not only do they provide greater control in giving instruction to DNA computers but they enable important new advances such as spatialisation of input.

Resume : The principles of self-organization of intelligent systems - is the basis of the designing of modern computers. The activation of hardware of any computer system implementation by running the bios-program and scan program environment of the operating system. This process has certain analogies with beginning of viability of living organisms from the genetic code of DNA, if we assume that the process bios - activation of computer is accompanied by searching conformity and agreement between a hardware and software system available and the formation of fixed relationships between them. This is especially true for modern branched and network systems, hot plug and auto identification of additional modules and peripherals. Systems that use elements of artificial intelligence and genetic algorithms are usually targeted at solving applied problems and can not influence on the architectural, structural and logical connections within the system. However, the range of tasks that need to be addressed when creating a modern bio integrated and cyber physical systems requires a multifunctional computer tools. On the other hand, the achievement of micro- and nanoelectronics, miniaturization units and of limited applicability in such systems, traditional architecture and system solutions. In this study substantiate the structural and functional and algorithmic methods and approaches to address these problems through the use of adaptive and self reconfigurable computer systems. The essence of adaptive re configuration in this sense is to use the principle of adapting the system to the new conditions of its operation, which is similar to the adaptation and variation in nature. The level of functional variability of the system shall be determined by the results of the knowledge environment functioning and formation of appropriate databases and knowledge. Such problems are currently solved by computer artificial neural network by learning and recognition of signals and images. The principle of self reconfiguration system provides in this case the possibility of correction and fixation of acquired knowledge and its application not only to improve the algorithm of the system, but also the partial modification of its architectural design. This is equivalent to the genetic modification of the DNA code of living organisms. Our results show that the proposed approach and the structural and algorithmic solutions can be the basis for the synthesis of multifunctional intelligent computer systems embedded in kiber physical devices and systems for various levels and purposes.

Resume : The dramatic increase in average life expectancy during the 20th century ranks as one of society’s greatest achievements. At the same time, an increased quality of life is expected as well. As a result, there is currently an ever increasing demand for biomedical devices with a perfect reliability and lifetime longer than 60 years. For this reason, new customized implants need to be developed, thus stimulating the research towards new materials, tailored macro- and micro-structures and advanced manufacturing methods. Commercial calcium phosphates are restricted to applications that require only moderate load bearing abilities. On the other hand, high toughness bioinert materials (such as alumina/zirconia composites) are used in the articulating couple of artificial joint implants, but they do not support direct bone/material interfaces or bond. To date, there is no tough and strong ceramic in regular clinical use with ability to create a strong, biologically relevant, interface with bone. New customized techniques, such as additive manufacturing, are currently under development leading to tailored material architecture and customized biomedical devices. However, most of the advances are currently achieved at the laboratory scale. The industrial development can be reached only if the gap between academic research, clinical use and commercial production is reduced, highlighting the need of continuous exchanges and cooperation between surgeons, industrials, engineers, material scientists and biologists. For this reason, this Cost Action, coordinated by Dr. Francis Cambier from Belgium Ceramic Research Centre of Mons, aims at creating the seed for the European research and industry collaboration, combining basic knowledge from academic laboratories, R&D centres, medical units from hospitals, and a significant number of companies. Currently, Institutions from 29 European Countries are involved in the Action, counting also on the participation of extra-Europe participants such as USA and Canada.

Resume : Over the recent years, a highly interdisciplinary field lying on the interface of Biology, Chemistry and Engineering has been developed concerning the design and the fabrication of bioinspired surfaces for the development of functional substitutes for damaged tissue. Towards this aim, the Direct Laser Writing (DLW) technique based on Multi-Photon Polymerization (MPP) of photosensitive materials has established a strong foothold, since it enables the fabrication of three-dimensional (3D) structures with sub-micron resolution. A decisive parameter for the accurate structures fabrication by DLW, besides the irradiation ones, is the structuring material choice. Indeed, it is the material properties that ineluctably and crucially determine definitely the success of the printing process and additionally the resolution of the laser fabricated structures as well as the potentiality of their bulk and surface functionalization. Here, we present the variety of the employed materials extended from non-biodegradable hybrid organic-inorganic ones to biodegradable synthetic or natural biopolymers. Other than that, the effect of the 3D structures topography on different cell responses are discussed. In particular, cell responses such as proliferation, migration and differentiation are investigated.

Resume : Previous studies have reported the use of 3D Polycaprolactone (PCL) macroporous scaffolds in bone tissue engineering applications, demonstrating their usefulness in promoting bone healing.1-3 In alternative studies conductive coating layers, used to provide electrical stimulation, were similarly shown to positively affect the proliferation of bone forming cells and their long-term functions.4 Here we report the use of vapor phase polymerization as a robust and reproducible technique for coating 3D printed PCL scaffolds with Poly(3,4-ethylenedioxythiophene):tosylate (PEDOT:tos). The surface and mechanical properties of the so developed conductive scaffolds have been investigated being these features crucial for their use in osteoinduction studies. Moreover, human fetal Mesenchymal Stem Cells (hfMSC) were showed to actively proliferate on the PEDOT:tos coated scaffolds. These preliminary results pave the way to further studies on the combined effects of electrical and topographical cues in bone tissue regeneration applications. References 1. Biomaterials (2014) 35, 5647-5659, 2. J. Biomed Mater Res A. (2010) 93, 4, 1358-1367, 3. Tissue Engineering: Part A (2011) 17, 19 2389-2397, 4. J. Mater. Chem. B, (2013) 1, 1439

Resume : Bombyx mori silk is of great interest to people for its outstanding mechanical and biological properties. However, the traditional electrospun regenerated silk fibroin (RSF) scaffolds from aqueous solution were weak and had limited applications. This study was to fabricate reinforced scaffolds with well-aligned RSF fibers electrospun on a layer of native extracellular matrix, bladder acellular matrix graft (BAMG). The silk fibroin fibers were well-aligned as a grill in multiple layers. Both the BAMG and the grill structure significantly improved the tensile properties and suture retention of the composite scaffolds. This made the scaffolds can be sutured well with tissue during implantation. In vitro assay indicates that the scaffolds had a good biocompatibility. Porcine iliac endothelial cells (PIECs) were attached and proliferated well on the vascular endothelial growth factor (VEGF) loaded scaffolds compared with those without VEGF. Moreover, the grill-like structure guides PIECs well along the aligned fiber.

Resume : Tissue engineering has potential to address this need through the combination of biomaterials, growth factors, and cells. Highly porous scaffolds are generally used as the substrate for anchorage dependent cells and to facilitate nutrient and metabolite distribution to guide cell growth leading to new bone tissue formation. Hydroxyapatite (HAP) has been investigated for bone replacement since this material mimics natural bone mineral features. HAP has been studied extensively in cell culture and possesses osteoconductivity. Regenerated Bombyx mori silk fibroin (SF) has excellent biological and mechanical properties, including biocompatibility, programmable biodegradability, and remarkable strength and toughness. One of the important physical forms for biomaterials is the formation of hydrogels, which has been extensively studied for a variety of polymers. The sol-gel transition depended on the concentration of the protein, temperature, and pH. In the SF hydrogel, random coil to β-sheet (physical cross-linking) structural transitions were noted during the process of hydrogelation. Due to the β-sheet formation, SF exhibits relatively slow degradation in vitro and in vivo, compared to collagen and many other biopolymers. In this study, the synthesis and characterization of bone-like mineral HAP into highly porous biodegradable silk fibroin scaffold with via chemical cross-linking reaction of SF by gamma-ray were investigated.

Resume : Calcium phosphates are widely used as resorbable bone substitutes under different forms, like Biphasic Calcium Phosphate (BCP) ceramics, Calcium Phosphate Cements (CPC), the latter being either purely inorganic cements or composite materials obtained by the addition of organic polymers and/or fibers. However, a number of technological issues still need to be addressed, in particular on the improvement of mechanical properties. The present lecture aims at reviewing recent works on the links between microstructure (porosity at different scales, nature and amount of polymer or fiber additives…) and several mechanical properties, may they concern elastic behaviour (Young’s modulus) or fracture behaviour (toughness and tolerance to damage), in relation with processing (strength and its statistical Weibull analysis). Ways to produce a variety of microstructures will be described (incorporating porogenic agents, changing the sintering temperature of BCP ceramics or the liquid-to-powder ratio in the CPC paste, adding cellulose ethers or fibers to CPC…) and their influence on mechanical properties will be discussed, in an attempt to propose strategies for material optimisation. In particular, it is shown that the addition of specific additives can improve fracture toughness, induce a good tolerance to damage, and may lead to interesting porous microstructures in view of biomedical applications.

Resume : We developed and patented a new method for improving osteointegration of titanium-based alloys [1]. Metallic artificial implants are widely used as medical devices to replace, support or enhance an existing biological structure. Titanium-based alloys possess advanced mechanical properties and excellent biocompatibility, but the bioactivity with the host has also to be improved. We obtained Ca-P /sodium titanate/ Ti-6Al-4V devices with good cell viability properties as well as good cohesion between the substrate and the coating. The implant surface was modified by mechanical, chemical and heat treatments, then a Ca-P layer was deposited onto the metal surface by sol-gel route. The coating has been characterized by X-ray diffraction, scanning electron microscopy performed on transverse section, and EDSX analysis. Nanoscratch tests showed a plastic deformation during the mechanical stress and no delamination of the coating was observed. SEM observations of residual grooves after the scratch with a maximal load of 100 mN showed the good adhesion of all Ca-P coatings onto the substrate. In vitro bioactivity of implants was test by Kokubo’s methods using the simulated body fluid. Finally, biological evaluations such as cell morphology and density as well as cell viability were performed. [1] Multilayer metallic materials with nanoporous calcium phosphate surface layers for medical implants Carrado et al. PCT Int. Appl. (2013), WO 2013068591 A1 20130516

Resume : Cellulose/calcium phosphate hybrid materials were synthesized via an ionic liquid-assisted route. Scanning electron microscopy, transmission electron microscopy, energy-dispersive X-ray spectroscopy, X-ray diffraction, infrared spectroscopy, and thermogravimetric analysis/differential thermal analysis show that, depending on the reaction conditions, cellulose/hydroxyapatite, cellulose/chloroapatite, or cellulose/monetite composites form. Preliminary studies with MC3T3-E1 pre-osteoblasts show that the cells proliferate on the hybrid materials suggesting that the ionic liquid-based process yields materials that are potentially useful as scaffolds for regenerative therapies.

Resume : LONGLIFE (“Advanced multifunctional zirconia ceramics for long-lasting implants”) coordinated by Prof. Jérôme Chevalier from INSA of Lyon, is a Collaborative European project in the frame of the 7th Framework Program. LONGLIFE aims at developing new zirconia-based oral and spine (lumbar inter-vertebral disc) implants, characterized by lifetime longer than 60 years and superior reliability as compared to previous biomedical devices. In order to reach this ambitious goal, efforts should be dedicated to improve the in-vivo stability of zirconia, by one side, and enhance the osseointegration capabilities of the implants in contact with bone, on the other. At the same time, the project aims at developing new ceramic-oriented designs for the implants as well as new multi-physic accelerated testing in vitro, able to reproduce more effectively the different degradation mechanisms and their interplay. In this work, the achievements concerning the design and development of new zirconia-based materials are presented, reached thanks to the synergies between Politecnico of Torino and INSA of Lyon. Among the technical oxide ceramics able to be used as structural biomaterials, zirconia-based ceramics are considered the best choice thanks to their excellent mechanical properties, biocompatibility and aesthetics. However, the current zirconia-based implants suffer from degradation under hydrothermal conditions (low temperature and presence of water), leading to surface degradation and possibly to fracture. Therefore, in order to increase the implants reliability, a strong effort is required to improve the zirconia stability in the presence of water, without decreasing its toughness and strength. The strategy chosen in LONGLIFE towards this goal is reported. Besides an innovative design of the composite material, a controlled synthesis process was used to produce multi-phase composites, having controlled architecture and composition. The results achieved demonstrate the effectiveness of this strategy towards the development of strong, tough and stable materials, fulfilling the objectives of the project.

Resume : Thanks to its biocompatibility, bioactivity, high osteoconductive and/or osteoinductive behaviour, hydroxyapatite (HA) has been deeply investigated for the production of bone substitutes. Besides several techniques currently used to synthesize HA powders, an innovative route implies the extraction of natural HA from bio-wastes. This method offers both biological and economic advances over the traditional synthetic techniques. Among bio-wastes, fish, bovine and pig bones have been explored as HA raw materials. The extraction of carbonated HA occurred through thermal, subcritical water or alkaline hydrothermal processes. In this work, natural HA powder was extracted from the cortical part of long pig bones, by treating at 100 °C with a NaOH water solution (4 M) for 48 hours. In order to remove the remaining sodium hydroxide, the material was then carefully washed into distilled water until a pH of 7 of the filtrate was reached. The dried powder was submitted to washing steps in distilled water, inducing de-carbonation by exploiting the high solubility of Ca(OH)2 in pure water. Washing steps were prolonged until a pH of 7 in the filtrate was reached. Starting from this natural HA powder, macro-porous ceramic materials have been prepared by a gel-casting process combined with a sacrificial template method. In particular, agar was used as natural gelling agent and polyethylene (PE) spheres as pore former. The sintered components were characterized by tailored macro- and micro-porosity features, as required for the development of engineered ceramic scaffolds.

Resume : The frequency of surgical septic postoperative complications in dirty wounds, which include a group of surgical interventions for purulent processes of maxillofacial area, averages more than 30-40%. This index is growing by every year due to increasing of widespread of multidrug-resistant pathogen strains and weakening of the immune status of patients. The same causes are among the leading in the case of the emergence and development of inflammatory complications in dental implantology because of a high risk of surgical wound infection by oral microflora. So, the prevention and treatment of such complications in the practice of maxillofacial surgery and dental implantology are important today. The effectiveness of silver nanoparticles (AgNP) and their combination with gold nanoparticles (Ag/AuNP) as new effective antimicrobial substances for maxillofacial surgery and dental implantology, in comparison with traditional antimicrobial substances has been studied. High bactericidal effectiveness of the studied metal nanoparticles (AgNP and Ag/AuNP) has been revealed in vitro against wide spectra of pathogens including multidrug-resistant ones (Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Klebsiella pneumoniae, Haemophilus influenzae, Micrococcus, Candida albicans etc.). All pathogens have been isolated from patients of the Department of Maxillofacial Surgery. High antimicrobial and anti-inflammatory effectiveness of AgNP and Ag/AuNP substances, in comparison with traditional antimicrobial drugs, has been shown in vivo on the model of abscesses of submandibular area using microbiological, biochemical and histological methods. Wistar rats have been used for modeling abscesses of submandibular area by means of subcutaneous injection in the submandibular area of the suspension containing clinical isolate of Staphylococcus aureus and active carbon powder.

Resume : By simulation of the biological process of synthesis and morphology of phosphates in the body the bioactive multiple doped nanoceramics have been developed which have 1000 - 10 000 times larger cross section of thermal neutron capture (or neutron with selected energy range), designed for neutron capture therapy of cancer tumors. The nanoceramics can be fixed at a predetermined position of the body through biochemical bonding and is fully bioresobable during 6 to 12 months after implantation.

Resume : This presentation is aimed at demonstrating the progress with the synthesis and characterization of new inorganic nanotubes (INT) and fullerene-like (IF) nanoparticles (NP) from 2-D layered compounds. Two important categories of new IF/INT nanostructures will be discussed in particular: 1. Synthesis of Doped IF/INT of WS2 (MoS2) by rhenium and niobium; 2. Synthesis of IF and in particular INT from the ternary misfit compounds, like PbS-TaS2, GdS-CrS2. The synthesis of 1-D nanostructures from this vast group of layered materials is particularly promising. Re-doped IF-MoS2 NP exhibit superior solid lubrication behavior in different environment and can find numerous applications in e.g. medical technology, which will be briefly demonstrated. Major progress has been achieved in elucidating the structure of INT and IF using advanced microscopy techniques, like aberration corrected TEM and electron tomography. Applications of the IF/INT as superior solid lubricants and for reinforcement of polymer nanocomposites, which gained a lot of momentum in recent times, will be briefly discussed. Few recent studies indicate that this brand of nanoparticles is not-toxic and biocompatible. With expanding product lines, manufacturing and sales, this generation of superior lubricants is becoming gradually a commodity.

Resume : Gene silencing via RNA interference (RNAi) has been demonstrated as a great potential therapeutic agent for cancer treatment. The emerging RNAi therapy delivers gene-silencing RNAs to the cytoplasm where they recognize and degrade the complementary target mRNA in a sequence-specific manner at a post-transcriptional level. However, safe and efficient transduction of siRNAs to target tissues and cells remains a major challenge for clinical trials. Therefore, we have developed several functional lipid-based nanoparticles as carriers for siRNA delivery to overcome the disadvantages mentioned above. For instance, many kinds of nanosized cationic delivery systems have been developed for siRNAs delivery via electrostatic interaction, such as cationic polymers, cationic lipids and so on. However, due to low charge density and stiff backbone structure, siRNA with 21-23 bp nucleotides has inherently poor binding ability to cationic polymers and lipid carriers, which results in low siRNA loading efficiency. In order to achieve high loading efficiency for gene transfection, cationic polymers and lipids must be used in an excess amount, whereas they often exhibit severe cytotoxicity. In this study, we report chemically conjugated siPlk1-phospholipids enveloped hybrid nanoparticles (siPlk1-PCNPs) for siRNA delivery to overcome the siRNA’s stiff backbone structures and enhance siRNA loading efficiency.

Resume : Regenerated silk fibroin (RSF)/graphene oxide (GO) hybrid silk fibers were dry-spun from a mixed dope of GO suspension and RSF aqueous solution. The rheological properties of the RSF/GO spinning dope were varying with the GO content. Synchrotron Radiation Wide Angle X-ray Diffraction and FTIR results showed that the addition of GO could confine the crystallization of silk fibroin which lead to the decrease of crystallinity, the smaller crystallite size and more formation of interphase zones in the artificial silks. Synchrotron Radiation Small Angle X-ray Scattering were also applied in the hybrid silks and hybrid solutions to study the interphase zones formed by the complex interaction between silk fibroin and GO flakes. An outstanding reinforcement was observed for the dry-spun hybrid silk fibers, of which the breaking stress reached 341.5 MPa.

Resume : Magnetoelectronics at the nanoscale provides new strategies to tackle issues concerning to life sciences and healthcare: from recognition of molecules to therapeutic health treatments. Magnetic structures can be exploited to detect molecules, bacteria and cells or to improve the functionalities of artificial biological tissues and finally may be directly employed to drug delivery and to kill cancers. However, this is only possible through a smart bio-functionalization of the magnetic objects, and having control onto their magnetic properties. Nanomagnetism applied to Nanomedicine appears thus as an emerging and multidisciplinary discipline that aims to provide personalized and more efficient tools to detect and treat health diseases, as delivering cancer treatments at the right place, at the right dose and at the right moment. This novel technology offers clear advantages over conventional techniques that can suppose a real breakthrough in biomedical research, and health care. Within this context, I illustrate different examples of bio-sensing and bio-medical applications based on magnetic nanomaterials.

Resume : Nanoparticles combining optical and magnetic signals are promising for the detection of sentinel nodes by hand-held probes in cancer surgery. Now, radioactive colloids (RuS labelled with 99mTc) or/and Vital Blue dye are injected around the primary tumour and detected by nuclear probe or the eye respectively. The aim of our work is to replace the radioactive colloids by magnetic colloids by developing and optimizing dendronized iron oxide nanoparticles. The nanoparticles were produced in an aqueous basic media and sorted by varying the ionic force, temperature and magnetic field parameters. The dendrons are either of first or second generation and bear a Patent Blue dye and/or a fluorescent dye. The suspensions have been characterized by dynamic light scattering, magnetic susceptibility and the amount of incorporated Patent Blue/fluorescent dye analyzed. We show that the structure and the composition of the aggregates strongly impact the optical and magnetic properties of the suspensions and therefore are critical for optimizing their detection sensitivity. Major attention was given to the effect of the dendron length, the grafting ratios, as well as to the electrostatic interactions between the dendron and the dye. Finally, in-vivo experiments on mice have been performed to determine the biodistribution. This work was supported by the European Union in the framework of the program «Nano@matrix» INTERREG IV Upper Rhine Valley.

Resume : Currently, therapy of cancer is mainly based on different modalities, which are distinctly selected depending on the cancer stage and tumor entity to be treated. Most of them are not highly specific, leading to side effects as well as long and painful patient recovery. Therefore, new treatment modalities have been proposed. Among them hyperthermia is considered to be a promising tool which basically addresses the transient rise of temperature above 37 °C leading to tumor cell death due to distinct changes of the tumor pathophysiology. Two different concepts of heating have been followed so far: hyperthermia (temperatures between 41 and 45 °C Experience from the clinical and preclinical studies has shown that a dedicated combination of hyperthermia with established oncologic modalities could markedly increase the therapeutic outcome and patient compliance (e.g. [6]). In this context, the synergy of therapeutic modalities with regard to breast and pancreatic cancer cells at early stages of tumor development are being evaluated in a collaborative European project (Multifunctional Nanotechnology for selective detection and Treatment of cancer, MultiFun, 7th framework program). Hereto, dedicated nanoparticle formulations in terms of their heating capabilities, morphologic and magnetic features, in vitro biocompatibility in isolated cells, pharmacokinetic properties and their therapeutic efficacy in vivo are being investigated.

Resume : Designing and synthesis of multifunctional nanoparticles for biological application is an actual challenge in which many research groups are now involved. Most of these studies are focused on the lowering the detection limit of cell expressions with respect to current diagnosis techniques. Our results start from the synthesis of naked AuNPs and FeOxNPs obtained with Laser Ablation synthesis in water. Nanoparticles are then combined in a core-shell-satellite type structure using MPTMS. SERS properties of these nanostructures are obtained with a dye embedded in the MPTMS shell. To perform immunomagnetic sorting, satellite AuNP are functionalized with an antibody for membrane prostatic antigen (PSMA). This type of nanostructures show promising results in immunomagnetic sorting and detection of prostate cancer cells. Associating different dyes with different antibodies there is also the possibility of performing SERS measurement with a multiplexed approach. 1 M. Meneghetti, et al. Small,(2012) 8, 3733 2 F. Bertorelle, et al. J. Phys. Chem. C (2014) 118, 14534 3 V. Amendola, et al. Small (2014) 10, 2476

Resume : The hydrothermally synthesized Fe3O4 NPs usually have a controllable size, narrow size distribution, and tunable magnetic properties, offering many opportunities in the field of biomedical imaging applications. This talk will be focused on the recent progress of hydrothermal synthesis and functionalization of iron oxide nanoparticles for magnetic resonance (MR) imaging applications developed in our group. Our work has been involved in the use of modified hydrothermal routes to generate surface-functionalized Fe3O4 NPs with good water dispersibility, colloid stability, and relatively high r2 relaxivity. In particular, the hydrothermally synthesized Fe3O4 NPs coated with branched polyethyleneimine were able to be further surface functionalized with targeting ligands (e.g., folic acid and hyaluronic acid), enabling effective tumor MR imaging with high specificity via receptor-mediated active targeting pathway. What’s more, the hydrothermal route was also used to synthesize Fe3O4@Au composite nanoparticles for dual mode MR and computed tomography (CT) imaging and photothermal therapy of tumors.

Resume : In addition to the increasing number of nanotechnology applications in medical diagnostics and therapeutics, the utilization of nanomaterials in consumer products has recently become more important. Among them, the spinel ferrite materials have widely been paid attention due to their current and future applications not only in the field of catalysts but also in the field of biology and medicine, such as diagnostics, biosensing, therapeutics, drug delivery and targeting. In this study, biocompatible multifunctional particles (CoFe2O4@HP) of well-defined sizes (60, 133, 245, 335 nm) were fabricated to be used as photodynamic therapeutic agents for cancer cells. Functionality of photodynamic therapy (PDT) is provided by hematoporphyrin (HP) which generates singlet oxygen with high quantum yield by photodynamic treatment. The HP molecules are covalently bonded to the surface of CoFe2O4 nanoparticle in various sizes of CoFe2O4 nanoparticle.The magnetic properties of cobalt ferrite (CoFe2O4) particles have been finely adjusted by controlling the size of the primary CoFe2O4 nanograins and the secondary superstructure composited particles formed by an aggregation of the nanograins. The prepared CoFe2O4@HP particles with various sizes exhibit high water solubility, good magnetic resonance imaging (MRI) and biocompatibility without any cytotoxicity. Especially, anticancer activities of CoFe2O4@HP present remarkable photodynamic anticancer efficiency via apoptosis in PC-3 cells, which showed particle size and dose dependent toxicity. This size dependent effect is determined by the particle specific surface area. These results indicate that CoFe2O4@HP are suitable for effective photodynamic therapy (PDT) and have potential as therapeutic agents for MRI based PDT, because they have a high value of saturation-magnetization and superparamagnetism

Resume : The understanding of the magnetoresistance (MR) responses as related to the way how the magnetization reverses in any kind of magnetic material is of great interest, not only for fundamental study, but importantly also for the improvement of the current spintronics technology. In this work, we demonstrate that by having direct experimental access to the magnetization vector during the reversal (by using a vectorial-Kerr magnetometry) we can predict the magnetoresistive signals. We demonstrate so in spin-valve [1] and in its basic constituents, i.e. single FM layer [2] and bilayer FM / AFM. We performed RT angular and field resolved M-H loops and the corresponding simultaneous resistance changes, and demonstrate that the R(H) curves depend exclusively on the reversal processes (ultimately dictated by magnetic anisotropy of the system). Very importantly, from the vectorial-resolved M-H we can experimentally determine the relative angle between the magnetization vectors of the two FM layers in spin-valve and the magnetic torque and the angle enclosed between the magnetization vector and the applied current direction in a single FM layer. This allows for the simulation and prediction of the giant magnetoresistance (GMR), anisotropic magnetoresistance (AMR) and Planar Hall Effect (PHE) in the whole angular range and for any magnetic field values. [1] P. Perna et al. Phys. Rev. B 86, 024421 (2012) [2] P. Perna et al. Appl. Phys. Lett. 104, 202407 (2014)

Resume : Photocatalyst semiconductor titanium dioxide is widely utilized as a self-cleaning and self disinfecting material for surface coating in many applications. It has a more helpful role in our environmental purification due to its nontoxicity, photoinduced super hydrophobicity and antifogging effect. Titanium dioxide used frequently in cosmetics, pharmaceutical, paint and paper industry. Recently the interest of TiO2 increased to the environmental, medical and biological application. The materials based on titanium dioxide can be used to create of antibacterial ceramic, paint and varnish coating and packing with antibacterial properties. It was studied the antibacterial activity of samples highly porous titanium dioxide. They were obtained of high temperature (700-1100 oC) by hydrolysis of titanium tetrachloride vapor in the air-hydrogen flame. Research of antibacterial activity of TiO2 performed by a diffusion method (disk methods) according to ISO 27447: 2009. It was shown that the samples of suspensions titanium dioxide (0.1-10.0 %) are exhibit antibacterial activity to bacterias Escherichia coli and Staphylococcus aureus. It was observed a complete neutralization of colonies of microorganisms Escherichia coli while lesion area of bacteria Staphylococcus aureus was lower. In both cases, the effect of antibacterial action increases with the concentration of TiO2.

Resume : Animal silks from spider and silkworm attract many people’s attention due to their outstanding mechanical properties and the smart spinning method of the animals. In this paper, several biomimetic spinning methods for spider silk and silkworm silk were designed. By mimicking the dry-spinning method of silkworm and spider, regenerated silk fibroin (RSF) aqueous solution was used as spinning dope and spun in air directly at room temperature. The as-spun fiber was then post-drawn in ethanol aqueous solution to induce the conformation transition of RSF. The oriented crystalline and amorphous regions of the silk fibers contribute to the remarkable mechanical properties of the artificial silk. By mimicking the functions of the spinning apparatus of spider and silkworm, ion and protein concentrations in the RSF aqueous solution were adjusted in microfluidic chips with multiple channels. Inspired by the shape and dimensions of the natural spinning apparatus, a microfluidic chip was designed and applied to the studies of aggregation mechanism of silk fibroin in micro-channel. Moreover, the dry-spun/electrospun fibers of RSF were also toughened or reinforced by mimicking the core-shell structure of natural animal silks, adding silk sericin, graphene oxide, TiO2 nanoparticles in spinning dopes and changing the collecting method in electrospinning process.

Resume : Insertion of endoscopes and other medical devices to the human body are ubiquitous, especially among aged males. The applied force for the insertion/extraction of the device from the urethra must overcome the endoscope (or catheter)-surface human-tissue interaction. In daily practice a gel is applied on the endoscope surface, in order to facilitate its entry to the urethra, providing also a local anesthesia. In the present work MoS2 nanoparticles, with fullerene-like structure and MoS2 doped with minute amounts of rhenium atoms, have been added to a commercial gel, in order to reduce the metal\catheter-urethra interaction and alleviate the potential damage to the epithelial tissue. In order to compare between the different gels a urethra model was designed and fabricated, which allowed a quantitative assessment of the applied force for extraction of the endoscope from a soft polymer-based ring. It is shown that the Re-doped nanoparticles reduce the traction force used to retrieve the metallic lead of the endoscope or the catheters from the soft ring by a factor close to three times with respect to the original gel. The mechanism of the mitigation of both friction and adhesion forces in these systems by the nanoparticles is discussed.

Resume : Here we develop a universal solution-processing approach for producing three dimensional (3D) conducting polymer-based bioelectronic interfaces (BEIs), which can be integrated on chips for rare circulating tumor cell (CTC) isolation and detection. Based on the chemical oxidative polymerization and modified poly(dimethylsiloxane) (PDMS) transfer printing technology, the poly(3,4-ethylenedioxythiophene) (PEDOT)-based micro/nanorod array films can be fabricated with topographical and chemical control through the chemical oxidative polymerization. This 3D PEDOT-based BEI film features the advantageous characteristics: (1) diverse dimensional structures (tunable from the microscale to the nanoscale), (2) varied surface chemical properties (tunable from nonspecific to specific), (3) high electrical conductivity, and (4) reversible electrochemical switching, and (5) high optical transparency. Furthermore, we integrated this 3D PEDOT-based BEI on chips, which exhibited optimal cell-capture efficiency from MCF7 cells was approximately 85%; featured highly efficient performance for the cell isolation of rare CTCs with minimal contamination from surrounding nontargeted cells (e.g., EpCAM-negative cells, white blood cells); preserved the cell viability with negligible effect on cells. According to the electric cell-substrate impedance sensing concept, the 3D BEI-based device was also demonstrated as a high sensitive and specific liquid biopsy tool for rapid CTC purification, detection and characterization. Therefore, it is conceivable that use of this platform will meet the requirements on developing for the next-generation bioelectronics for biomedical applications.

Resume : Circulating tumor cells (CTCs) in the bloodstream are exceptionally rare. As a result, efficient and specific isolation and capture of CTCs from peripheral blood are important not only for disease diagnosis, but also for patient treatment at early stages. In this work, we reported a platform comprised of electrospun nanofibers and a microfluidic device to isolate CTCs in whole blood with high efficiency and specificity. The nanofibers with high surface-to-volume ratio were aligned or randomly electrospun and embedded in the microchannels. The fiber diameter and biotin conjugation were investigated and optimized by studying their effects on the CTCs capture efficiency. In addition, both microfluidic devices contained aligned and random nanofibers showed low yield of nonspecific binding of CTCs. As compared with the aligned device, the random device yielded higher capture efficiency and recovery of 97% from whole blood. Results showed that high numbers of captured CTCs were viable, making them an ideal platform for subsequent molecular- or cellular-level studies.

Resume : Electrical conducting polymers (ECPs) are promising as interfacial materials to provide a combination of the electrical, biochemical and topological requirements for neural interfaces. ECPs are electrical/ionic conductive and effective at electron-ion transition over device-tissue boundary. Pioneer works have demonstrated the efficient utilization of ECPs to modulate cellular activities (including cell adhesion, migration, DNA synthesis, protein secretion, etc.) upon electrical stimulation. Nowadays, many research interests on bioelectronic devices have shifted to ethylenedioxythiophene (EDOT)-based polymers because of its long-term electro-stability in aqueous solution. In the present research, hydrophilic and bio-functionalized ethylenedioxythiophene (EDOT) building blocks were designed on mimicking natural bio-membranes, and chemically synthesized. The functionalized EDOT building blocks were electrodeposited as cell-nonstick or stick polymer thin films. The surface properties of the thin films could be easily switched from cell binding to cell resistant by deposition of a new layer of EDOT polymer onto the existing surface. This feature allow us to integrate the material assembly process with top-down lithographic process, thereby fabricating patterned all conducting polymer biointerface to spatially define the cell behaviors. The spatial control on attachment of cell could be available at a single cell if the binding domain was small enough. Finally, this platform was further employed to integrate spatial arrangements, surface functionality and electric stimulation to direct cell polarity, proliferation and differentiation. The spatial definition of cells at cellular level on all-conducting-polymer interface would not only help us to understand how electrical stimuli modulates cellular activities of neurons, but also could be adopted to guide nerve regeneration.

Resume : In recent years, a new form of carbon nanotube electrode spread like promising nanoelectronic devices based on carbon and for biological and medical applications. Because of their physical property, chemical and electrical. Carbon nanotubes can be effectively used as electrochemical sensors. The integration of carbon nanotubes provides a good solid support for the immobilization of enzymes. Determining glucose levels using biosensors, especially in medical diagnostics and in food industries. This action provides high accuracy and rapid detection rate. The purpose of this study includes the presentation of the transistor-based biosensor field effect structure with a grid of carbon nanotubes for detection of glucose with an analytical study of biosensor, and in this model we present a relationship combines between the glucose concentration and one of transistor parameters. .

Resume : Organic acids and their salts (sorbic acid (E200) and sodium sorbate, potassium sorbate, calcium sorbate (E201, E202, E203), benzoic acid (E210) and sodium benzoate, potassium benzoate, calcium benzoate (E211, E212, E213), acetic acid (E 260) and sodium acetate, calcium acetate, ammonium acetate (E262, E263, E264), ascorbic acid (E300) and sodium ascorbate, calcium ascorbate, potassium ascorbate (E301, E302, E303), etc.) are widely used as food additives (preservatives, antioxidants) in the food industry and in cosmetics, pharmacy and medicine. However, beyond the limiting doses they can have harmful effects on the human body that actualizes on developing rapid methods of control of their content in food products. When sampling for analysis important point is to ensure the necessary concentration of controlled substances according to the sensitivity of the measuring channel information-measuring system. The purpose of this study was to investigate the sorption properties of porous silicon on some of these organic acids and their salts. Using double-stage processing of silicon wafers for the active porous silicon surface: 1) chemical treatment in ammoniac-peroxide and acid-peroxide solutions for various combination options; 2) formation of anodic porous silicon surface at different values of anode current. The research of the sensory properties of porous silicon surface was performed photometric, spectral and other methods. The sensitivity of the optical solar cells was provided at a pico Ampere. For rapid analysis of the results used mobile computerized measuring system. The error of measurements and data processing did not exceed 8%. According to the research proposed and optimized structure of the measuring cell and method of rapid analysis of some organic acids and their salts-food additives. It is shown that in the study of the sorption properties of porous silicon on organic compounds there is a relationship between the presence of active -O-O- and -O-H groups and chemical surface treatment method of silicon wafers in ammoniac-peroxide or acid-peroxide solutions.

Resume : An optically transparent poly(3,4-ethylenedioxythiophene) (PEDOT) based organic electronic devices have been developed to investigate the behavior of human mesenchymal stem cell (hMSC) and to sense and to capture circulating tumor cells. We first conducted a control experiment on electrical cell-substrate impedance sensing (ECIS) device by culturing the hMSC on the chip. According to our result, the impedance increased reflected the hMSC proliferation, attachment and motility during the first 16 hours of cell culture. In order to control the differentiation of human mesenchymal stem cell (hMSC) on chip, we also developed all-solution-processed multifunctional organic devices, comprising reduced graphene oxide (rGO) and dexamethasone 21-phosphate disodium salt (DEX) drug loaded poly(3,4-ethylenedioxythiophene) (PEDOT) microelectrode arrays on indium tin oxide glass, that can be used to manipulate differentiation. In our devices, the rGO micropatterns were used as the adhesive coating to attract the adhesion of hMSC cells whereas PLL-g-PEG coated PEDOT electrodes served as the anti-adhesive coating where no hMSC cells can attach. In addition, the PEDOT electrodes also work as drug releasing components where control DEX release from PEDOT matrix can be achieved via cyclic potential stimulation (CPS). To capture the circulating tumor cells, we fabricated 3D PEDOT-based micro/nanorod array, which can be further surface-grafted with capture agents for directed specific recognition to study the cell–substrate interactions on bioelectronics interfaces (BEIs). This BEI platform features the advantageous characteristics: (1) diverse dimensional structures (tunable from the microscale to the nanoscale), (2) varied surface chemical properties (tunable from nonspecific to specific), (3) high electrical conductivity, and (4) reversible chemical redox switching. Furthermore, through systematic studies of PEDOT systems, we explore the effects of both chemistry and topography on the circulating tumor cell (CTC)-capture performance. The 400 nm PEDOT pillars exhibited the optimal cell-capture efficiency; it could be used to isolate CTCs with minimal contamination from surrounding nontargeted cells (e.g., EpCAM-negative cells, white blood cells) and negligible disruption of the CTCs’ viability and functions. It is conceivable that PEDOT-based micro/nanorod array films function as a critical therapeutic intervention for monitoring tumor progression and metathesis, providing valuable insight into the use of electronics for tissue engineering and regenerative medicine.

Resume : Immunogenic scar formation would insulate electrodes from targeted cells and tissues, thereby reducing the lifetime of the bioelectronic devices. Nonspecific interactions could make electrodes binding non-targeted cells, and lead to the poor selectivity of biosensors. Ideally, an electrode material capable of electrically interfacing with cells selectively and efficiently would be integrated without being recognized by the immune system or nonspecific interaction to cells. We developed a series of biomimetic conducting polymer from zwitterionic and cell-targeted ethylenedioxythiophenes (EDOTs), which specifically interact with cells on a protein-resistance background. The predominant presence of zwitterionic side-groups on surface prevent enzyme and cell binding, allowing the polymers to functionalize in aqueous buffer for an extended period of time and preventing nonspecific interaction. The cell-targeted peptides, proteins or polymer chains on surface ensure specific interaction with desired cells. The further introduction of stimuli-responsive bio-conjugation linkage or polymer chains allows a stimuli-responsive cell capture and release, which could be used to build noninvasively removed bioelectronic devices or cell-detachable sensors. Combining the molecular design features, these materials further demonstrated the following superior material performances. They displayed low impedance at low frequency range, and this is ideal for biological application. The biofunctions presented on the surface could also provide biochemical stimulation to the differentiation of cells attached. Stable binding and electrical communication could be created between biomimetic PEDOTs and cells attached. The cell-substrate interaction could be largely enhanced by Nano-assemble of PEDOTs or extension of polymer chains on surfaces.

Resume : Preventive health care is an emerging trend requiring easy to use methods and tools. Health monitoring on daily or weekly basis would benefit from fast and label-free detection of for example saliva, urine or blood, and from portable units with connectivity via internet or mobile phones. Most of the biomolecules carry net charge detectable with charge sensitive devices, such as field effect transistors, leaving the problem to achieve selectivity and immobilisation of the receptors. Hydrophobins are protein amphiphiles with a hydrophobic patch at one end facilitating formation of well-ordered monolayers on hydrophobic surfaces and, consequently, providing means to immobilize receptors [1]. Hydrophobins have been used to exfoliate and in-situ functionalise graphene flakes [2] and for directed self-assembly of gold nanoparticles on silicon surfaces [3]. Hydrophobins can be genetically engineered to realise receptor modules specific to various analytes. In these experiments graphene field effect transistors (GFET) have been used as charge detectors and the non-linear characteristics near the Dirac point provide very high charge sensitivity. The fusion proteins form dense monolayer on the hydrophobic graphene channel [4] and immobilise the receptors enabling selective bio-recognition. We have tested the approach with hydrophobin-ZE - ZR amino acid zipper pair and hydrophobin-protein A – IgG immunoglobulin antibody and measured sensitivities at the level of femtomols with a few seconds response time. In the presentation we will discuss the possible applications and the potential limitations of the concept. [1] Linder, M. B., Szilvay, G. R.,Nakari-Setälä, T., Penttilä, M. E., FEMS Microbiology Reviews 29 (2005) 877–896. [2] Laaksonen, P., Kainlauri, M., Laaksonen, T., Shchepetov, A., Jiang, H., Ahopelto, J., and Linder, M. B., Angew. Chem. Int. Ed., 49, (2010), 4946-4949. [3] Laaksonen, P., Kivioja, J., Paananen, A., Kainlauri, M., Kontturi, K., Ahopelto, J., and Linder, M.B., Langmuir 25 (2009) 5185–5192. [4] Kivioja, J.; Kurppa, K.; Kainlauri, M.; Linder, M. B.; Ahopelto, J., Appl. Phys. Lett. 94 (2009) 183901.

Resume : It may be expected that methods for investigating cell-cell interactions in an in vitro tissue model will facilitate to gain deeper insights into operating principles underlying organogenesis during development and tissue regeneration. Here we present our approach in which a cell culture substrate was fabricated through micrometer-scale two-dimensional antibody display for permitting to investigate dynamics of heterotypic cellular interactions. In this study, special attention was paid to an epithelial-mesenchymal interaction that is ubiquitously seen in tissue morphogenesis. For the preparation of a cell culture substrate, the surface of a glass plate was cationically modified and then used as a substrate for two-dimensional antibody display. Using a poly(dimethylsiloxane) stamp with a microfluidics, two antibodies against distinct surface markers of epithelial and mesenchymal cells were immobilized in a site-addressable manner: Two antibodies were immobilized in separate stripes in close proximity to each other. To demonstrate the feasibility of the substrate for micrometer-scale positioning of heterotypic cells, populations of epithelial and mesenchymal cells were simultaneously seeded to the substrate that displayed antibodies. It was shown that our method using the stamp with a microfluidics allowed us to immobilize two different antibodies in a site-addressable manner. On this substrate, gingival epithelial cells and mesenchymal cells could be successfully positioned to the regions where antibodies specific for either of the two cell types were immobilized. These cells could be further cultured in situ on the substrate for analyzing cellular proliferation, migration and morphological changes. The results obtained in our study suggest that the two-dimensional antibody display serves to establish the spatially-controlled co-culture of two different cell populations, providing an initial condition for further study on the dynamic evolution of heterotypic cellular systems.

Resume : To understand life it is important to clarify various phenomena that occur in the cell at the molecular level. Several methods have been developed to monitor cellular processes, especially, imaging of specific signaling molecules including metal ions that may provide spatiotemporal information on their location in the cell. Since potassium ion (K+) plays an important role in many physiological events such as homeostasis in the heart muscle and hyper polarization of neurons, it is important to develop not only a detection method for this cation, but also a fluorescence imaging technique. However, specific detection of K+ is hampered because of the existence of sodium (Na+) and other ions in the cell. Oligonucleotides with sequences of human telomeric DNA or thrombin binding aptamer (TBA) are known to form tetraplex structures upon K+ ion binding. We successfully synthesized a novel fluorescent reagent, PSO-5, showing preference for K+ on the basis of a conformational change in G-rich DNA. PSO-5 consisted of thrombin binding aptamer (TBA) carrying FAM at the 5’-end conjugated with a peptide carrying TAMRA and biotin at the middle and at its C-terminus, respectively. The ternary complex of streptavidin with PSO-5 and biotinylated nuclear export signal peptide in a 1:1:3 stoichiometry had the dissociation constant of 2.24 mM for K+ and its preference for K+ is 236 times over Na+. K+ in the cell was visualized based on the FRET ratio of this complex. Changes in the FRET signal of PSO-5 was observed to indicate a decrease in K+ concentration in the cell upon addition of amphotericin B and ouabain which facilitate K+ efflux from the cell.1 [1] K. Ohtsuka, S. Sato, Y. Sato, K. Sota, S. Ohzawa, T. Matsuda, K. Takemoto, N. Takamune, B. Juskowiak, T. Nagai, S. Takenaka, Chem. Comm., 2012, 48, 4740-4742.

Resume : Circulating tumor cells (CTCs) have become an emerging ?biomarker? for monitoring cancer metastasis and prognosis. Although there are existing technologies available for isolating/counting CTCs, the most common of which using immunomagnetic beads, they are limited by their low capture efficiencies and low specificities. By introducing a three-dimensional (3D) nanostructured substrate ? specifically, a silicon-nanowire (SiNW) array coated with anti-EpCAM ? we can capture CTCs with much higher efficiency and specificity. The conventional methods of isolating CTCs depend on biomolecular recognitions, such as antigen-antibody interaction. Unlikely, we here proposed that nanoscaled local topographic interactions besides biomolecular recognitions inspired by natural immuno-recognizing system. This cooperative effect of physical and chemical issues between CTCs and substrate leads to increased binding of CTCs, which significantly enhance capture efficiency. Recently, we have also developed a 3D cell capture/release system triggered by aptamer enzyme, electrical potential and Temperature, which is effective and of ?free damage" to capture and release cancer cells. The bio-inspired interfaces of cell capture and release open up a light to rare-cell based diagnostics, such as CTCs, fetal cells, stem cell and so on.

Resume : Genetically encoded green fluorescent protein and its homologs opened the avenue for a wide applications of these intrinsic biological labels representing specific biological proteins. In this work we report on a new visible blue/green fluorescent phenomenon found in -sheet nanowires of ultrashort bionspired peptides. Di- and tri-peptide nanostructures self-assembled from biomolecules of different compositions and origin such as aromatic, aliphatic, linear and cyclic (diphenylalanine, FF-; dileucine, LL-; triphenylalanine, FFF- monomers) have been studied. At elevated temperature 140-180?C these supramolecular structures of diverse morphologies undergone the same irreversible thermally-induced phase transformation. This reconstruction process is followed by deep modification at all levels: molecular, electronic, elementary symmetry, peptide secondary structure, displaying new nanowire morphology and completely new common physical properties. The thermally-induced nanowire supramolecular ensembles acquire -sheet secondary structure with new building blocks and new non-covalent hydrogen bonds. In this phase the -sheet nanowires, irrespective of their native biomolecules origin, exhibit similar profound alteration of optoelectronic properties and appearance of visible (blue and green) fluorescence ascribed to non-covalent hydrogen bonds. Observed visible fluorescent of self-assembled dyes (hydrogen bonds) in thermally-induced peptide fiber nanostructures can be used as intrinsic optical labels in biomedicine as well for a new generation of novel optoelectronic biocompatible nanomaterials for emerging nanophotonic applications such as bio-lasers, integrated optics and biocompatible markers.

Resume : Screen-printed electrodes (SPEs) are used as cost-effective (bio)sensor platforms, which have been successfully utilised for rapid in situ analysis of biomarkers, enviromental pollutants, biorecognition interactions such as nucleic acids, or proteins due to their advantageous material properties, disposability, simplicity, and rapid responses (1-10). Screen printed electrodes have been fabricated as the miniaturized and multi channel forms of the electrochemical analysis systems (8-10). These disposable (bio)sensors can easily be modified with different nanomaterials such as carbon nanotubes, nanoparticles and dendrimers [4,7,10]. They are appropriate candidates for on-line measurements of numerous biological samples due to require low sample volumes. Multi channel screen-printed electrochemical array systems tested for biomolecular interactions have been overviewed herein for monitoring of spesific biomolecular recognitions; such as, nucleic acids and aptamer-protein interactions with their advantages and further applications. Acknowledgements. A.E would like to express her gratitude to the Turkish Academy of Sciences (TUBA) as the associate member of TUBA for its partial support. References: 1- J. Wang, Electroanalysis 17, 2005, 7-14. 2- M. Li, Y.-T. Li, D.-W. Li, Y.-T. Long, Analytica Chimica Acta, 734, 2012, 31?44 3- O. Dom?nguez Renedo,M.A. Alonso-Lomillo, M.J. Arcos Mart?nez, Talanta 73, 2007, 202?219. 4- D. Kumar, B.B. Prasad, Sensors and Actuators B: Chem. , 171, 2012, 1141-1150. 5- J. Wang, D. Xu, A. Erdem, R. Polsky, M. Salazar, Talanta, 56, 2002, 931-938 6- J. Wang, J.-W. Mo, A. Erdem, Electroanalysis, 14, 2002, 1365-1368. 7- F. Rohrbach, H. Karadeniz, A. Erdem, M. Famulok, G. Mayer, Analytical Biochemistry, 421, 2012, 454-459. 8- A. Erdem, G. Congur, E. Eksin, Sensors and Actuators B: Chem., 188, 2013, 1089-1095. 9- A. Erdem, G. Congur, Talanta, 118, 2014, 7-13. 10- A. Erdem, G. Congur, Sensors and Actuators B: Chem., 196, 2014, 168-174.

Resume : Cells are known to probe different physical properties of their environment. In the mechanotransduction field many tools are used to study these signaling events, such as engineered substrates that allow cellular traction forces measurements. Our study introduces a novel nanopillar platform, which revealed that the forces acting on perinuclear nanopillars are rich in α5β1-integrins and significantly higher than those acting on αvβ3 integrin-rich peripheral focal adhesions. These perinulear forces increased upon integrin activation using Mn2 and decreased with pharmacological Latrunculin B* treatement. LMNA knockouts allowed to manipulate peripheral versus perinuclear traction forces and confirmed that these central forces originate from actin fibers that span across the cell nucleus.