Influence of the type of substrate on the properties of carbon nanotubes layer studied by Raman spectroscopy.

The development of nanomaterials technology allows to design a novel medical strategies, and could also be useful in the field of regenerative medicine. The paper presents a study on the functionalized multi-walled carbon nanotubes (MWCNTs-f) layers deposited by electrophoretic method (EPD) on the surfaces of two types of substrates: titanium (Ti) and stainless steel. SEM and EDS analyses confirm that incubation in a simulated body fluid (SBF) caused a formation of hydroxyapatite on the surface of the Ti/MWCNTs-f. Raman micro-spectroscopy was a method of choice to study presented materials. The MWCNTs-f layer on the surface of the titanium plate shows better layer order than the corresponding layer deposited on the stainless steel. The structure and ordering of the nanocarbon layer play a key role in the biological activity of the materials. This was confirmed by the incubation of the plates with deposited layer of carbon nanotubes in SBF. A titanium substrate with a MWCNTs-f layer supports the deposition of some components from the environment, while a stainless steel substrate promotes the formation of a carbon film that inhibits the deposition of certain components from the environment. A two-trace two-dimensional (2T2D) analysis confirmed a different effect of SBF on the MWCNTs-f layer depending on the type of substrate. The MWCNTs-f layer on titanium substrate seems to represent an interesting proposition for novel bioactive strategies.

A study of the interactions between human osteoblast-like cells and polymer composites with functionalized graphene derivatives using 2D correlation spectroscopy (2D-COS).

In response to the growing need for development of modern biomaterials for applications in regenerative medicine strategies, the research presented here investigated the biological potential of two types of polymer nanocomposites. Graphene oxide (GO) and partially reduced graphene oxide (rGO) were incorporated into a poly(ε-caprolactone) (PCL) matrix, creating PCL/GO and PCL/rGO nanocomposites in the form of membranes. Proliferation of osteoblast-like cells (human U-2 OS cell line) on the surface of the studied materials confirmed their biological activity. Fluorescence microscopy was able to distinguish the different patterns of interaction between cells (depending on the type of material) after 15 days of the test run. Raman micro-spectroscopy and two-dimensional correlation spectroscopy (2D-COS) applied to Raman spectra distinguished the nature of cell-material interactions after only 8 days. Combination of these two techniques (Raman micro-spectroscopy and 2D-COS analysis) facilitated identification of a much more complex cellular response (especially from proteins) on the surface of PCL/GO. The presented approach can be regarded as a method for early study of the bioactivity of membrane materials.

Corrosion Resistance and Electrical Conductivity of Hybrid Coatings Obtained from Polysiloxane and Carbon Nanotubes by Electrophoretic Co-Deposition.

Nanocomposites developed based on siloxanes modified with carbon nanoforms are materials with great application potential in the electronics industry, medicine and environmental protection. This follows from the fact that such nanocomposites can be endowed with biocompatibility characteristics, electric conductivity and a high mechanical durability. Moreover, their surface, depending on the type and the amount of carbon nanoparticles, may exhibit antifouling properties, as well as those that limit bacterial adhesion. The paper reports on the properties of polysiloxane (PS) and carbon nanotubes (CNT) nanocomposite coatings on metal surfaces produced by the electrophoretic deposition (EPD). A comparison with coatings made of pure PS or pure CNT on the same substrates using the same deposition method (EPD) is provided. The coatings were examined for morphology and elemental composition (SEM, EDS), structural characteristics (confocal Raman spectroscopy), electrical conductivity and were tested for corrosion (electrochemical impedance spectroscopy-EIS, potentiodynamic polarization-PDP). The results obtained in this study clearly evidenced that such hybrid coatings conduct electricity and protect the metal from corrosion. However, their corrosion resistance differs slightly from that of a pure polymeric coating.

Early Recognition of the PCL/Fibrous Carbon Nanocomposites Interaction with Osteoblast-like Cells by Raman Spectroscopy.

Poly(ε-caprolactone) (PCL) is a biocompatible resorbable material, but its use is limited due to the fact that it is characterized by the lack of cell adhesion to its surface. Various chemical and physical methods are described in the literature, as well as modifications with various nanoparticles aimed at giving it such surface properties that would positively affect cell adhesion. Nanomaterials, in the form of membranes, were obtained by the introduction of multi-walled carbon nanotubes (MWCNTs and functionalized nanotubes, MWCNTs-f) as well as electro-spun carbon nanofibers (ESCNFs, and functionalized nanofibers, ESCNFs-f) into a PCL matrix. Their properties were compared with that of reference, unmodified PCL membrane. Human osteoblast-like cell line, U-2 OS (expressing green fluorescent protein, GFP) was seeded on the evaluated nanomaterial membranes at relatively low confluency and cultured in the standard cell culture conditions. The attachment and the growth of the cell populations on the polymer and nanocomposite samples were monitored throughout the first week of culture with fluorescence microscopy. Simultaneously, Raman microspectroscopy was also used to track the dependence of U-2 OS cell development on the type of nanomaterial, and it has proven to be the best method for the early detection of nanomaterial/cell interactions. The differentiation of interactions depending on the type of nanoadditive is indicated by the ν(COC) vibration range, which indicates the interaction with PCL membranes with carbon nanotubes, while it is irrelevant for PCL with carbon nanofibers, for which no changes are observed. The vibration range ω(CH2) indicates the interaction for PCL with carbon nanofibers with seeded cells. The crystallinity of the area ν(C=O) increases for PCL/MWCNTs and for PCL/MWCNTs-f, while it decreases for PCL/ESCNFs and for PCL/ESCNFs-f with seeded cells. The crystallinity of the membranes, which is determined by Raman microspectroscopy, allows for the assessment of polymer structure changes and their degradability caused by the secretion of cell products into the ECM and the differentiation of interactions depending on the carbon nanostructure. The obtained nanocomposite membranes are promising bioactive materials.

Carbon nanotube/iron oxide hybrid particles and their PCL-based 3D composites for potential bone regeneration.

This study describes the preparation, and evaluates the biocompatibility, of hydroxylated multi-walled carbon nanotubes (fCNTs) functionalized with magnetic iron oxide nanoparticles (IONs) creating hybrid nanoparticles. These nanoparticles were used for preparing a composite porous poly(ε-caprolactone) scaffolds for potential utilization in regenerative medicine. Hybrid fCNT/ION nanoparticles were prepared in two mass ratios - 1:1 (H1) and 1:4 (H4). PCL scaffolds were prepared with various concentrations of the nanoparticles with fixed mass either of the whole nanoparticle hybrid or only of the fCNTs. The hybrid particles were evaluated in terms of morphology, composition and magnetic properties. The cytotoxicity of the hybrid nanoparticles and the pure fCNTs was assessed by exposing the SAOS-2 human cell line to colloids with a concentration range from 0.01 to 1 mg/ml. The results indicate a gradual increase in the cytotoxicity effect with increasing concentration. At low concentrations, interestingly, SAOS-2 metabolic activity was stimulated by the presence of IONs. The PCL scaffolds were characterized in terms of the scaffold architecture, the dispersion of the nanoparticles within the polymer matrix, and subsequently in terms of their thermal, mechanical and magnetic properties. A higher ION content was associated with the presence of larger agglomerates of particles. With exception of the scaffold with the highest content of the H4 nanoparticle hybrid, all composites were superparamagnetic. In vitro tests indicate that both components of the hybrid nanoparticles may have a positive impact on the behavior of SAOS-2 cells cultivated on the PCL composite scaffolds. The presence of fCNTs up to 1 wt% improved the cell attachment to the scaffolds, and a content of IONs below 1 wt% increased the cell metabolic activity.

Reconstruction of Ovine Trachea with a Biomimetic Composite Biomaterial.

The aim of this study was to evaluate a novel composite material for tracheal reconstruction in an ovine model. A polymer containing various forms of carbon fibers (roving, woven, and nonwoven fabric) impregnated with polysulfone (PSU) was used to create cylindrical tracheal implants, 3 cm in length and 2.5 cm in diameter. Each implant, reinforced with five rings made of PSU-impregnated carbon-fiber roving, had three external layers made of carbon-fiber woven fabric and the inner layer formed of carbon-fiber nonwoven fabric. The inner surface of five implants was additionally coated with polyurethane (PU), to promote migration of respiratory epithelium. The implants were used to repair tracheal defects (involving four tracheal rings) in 10 sheep (9-12 months of age; 40-50 kg body weight). Macroscopic and microscopic characteristics of the implants and tracheal anastomoses were examined 4 and 24 weeks after implantation. At the end of the follow-up period, outer surfaces of the implants were covered with the tissue which to various degree resembled histological structure of normal tracheal wall. In turn, inner surfaces of the prostheses were covered only with vascularized connective tissue. Inner polyurethane coating did not improve the outcomes of tracheal reconstruction and promoted excessive granulation, which contributed to moderate to severe stenosis at the tracheal anastomoses. The hereby presented preliminary findings constitute a valuable source of data for future research on a tracheal implant being optimally adjusted for medical needs.

Raman studies of the interactions of fibrous carbon nanomaterials with albumin.

Adsorption or immobilization of proteins on synthetic surfaces is a key issue in the context of the biocompatibility of implant materials, especially those intended for the needs of cardiac surgery but also for the construction of biosensors or nanomaterials used as drug carriers. The subject of research was the analysis of Raman spectra of two types of fibrous carbon nanomaterials, of great potential for biomedical applications, incubated with human serum albumin (HSA). The first nanomaterial has been created on the layer of MWCNTs deposited by electrophoretic method (EPD) and then covered by thin film of pyrolytic carbon introduced by chemical vapor deposition process (CVD). The second material was formed from carbonized nanofibers prepared via electrospinning (ESCNFs) of polyacrylonitrile (PAN) precursor and then covered with pyrolytic carbon (CVD). The G-band blue-shift towards the position of about 1600cm-1, observed for both studied surfaces, clearly indicates the albumin (HSA) adhesion to the surface. The G and G' (2D) peak shift was employed to assess the stress build up on the carbon nanomaterials. The surface nano- and micro-topography as well as the method of ordering the carbon nanomaterial has a significant influence on the mode of surface-protein interaction.

The use of calcium ions instead of heat treatment for ß-1,3-glucan gelation improves biocompatibility of the ß-1,3-glucan/HA bone scaffold.

The aim of this study was to determine which procedure for ß-1,3-glucan gelation - newly developed dialysis against calcium salt or described in the literature thermal technique - is more appropriate for fabrication of a biomaterial designed for bone tissue engineering applications. Thus, ß-1,3-glucan/hydroxyapatite scaffolds were prepared based on two different methods and their physicochemical, microstructural, and biological properties were compared. Obtained results demonstrated that unlike thermal method-prepared ß-1,3-glucan/hydroxyapatite material (glu/HAT), bone scaffold fabricated via dialysis method (glu/HA D) possessed rough surface resulting from the presence of CaCl2 precipitates as proven by SEM and EDS analysis. As a consequence, glu/HA D scaffold released Ca2+ ions to the surrounding environment positively affecting osteoblast behaviour and biomineralization in vitro. Since glu/HA D material exhibited better bioactivity and biocompatibility compared to the glu/HA T scaffold, it may be concluded that the dialysis method is more suitable for ß-1,3-glucan/hydroxyapatite biomaterial fabrication.

Hybrid chitosan/ß-1,3-glucan matrix of bone scaffold enhances osteoblast adhesion, spreading and proliferation via promotion of serum protein adsorption.

Initial protein adsorption to the material surface is crucial for osteoblast adhesion, survival, and rapid proliferation resulting in intensive new bone formation. The aim of this study was to demonstrate that modification of a chitosan matrix of chitosan/hydroxyapatite (chit/HA) biomaterial for bone tissue engineering applications with linear ß-1,3-glucan (curdlan) leads to promotion of serum protein adsorption to the resultant scaffold (chit/glu/HA) and thus in enhancement of osteoblast adhesion, spreading and proliferation. Fabricated biomaterials were pre-adsorbed with different protein solutions and then protein adsorption and osteoblast behavior on the scaffolds were compared. Moreover, surface chemical composition, wettability and surface energy of biomaterials were compared. Modification of the chitosan matrix with ß-1,3-glucan introduces a greater polarpart in the resultant chitosan/ß-1,3-glucan matrix presumably resulting from more OH groups within the curdlan structure. Moreover, FTIR-ATR results suggest that there might be some sort of chemical interaction between the NH group of chitosan and the OH group of ß-1,3-glucan. As a consequence, the chit/glu/HA scaffold adsorbs significantly more adhesion proteins that are crucial for osteoblasts compared to the chit/HA material, providing a higher density culture of well-spread osteoblasts on its surface. Obtained results revealed that not only is chit/glu/HA biomaterial a promising scaffold for bone tissue engineering applications, but the specific polysaccharide chit/glu matrix itself is promising for use in the biomedical material field to modify various biomaterials in order to enhance osteoblast adhesion and proliferation on their surfaces.

Haemocompatibility and cytotoxic studies of non-metallic composite materials modified with magnetic nano and microparticles.

PURPOSE: Preventing the formation of blood clots on the surface of biomaterials and investigation of the reasons of their formation are the leading topics of the research and development of biomaterials for implants placed into the bloodstream. Biocompatibility and stability of a material in body fluids and direct effect on blood cell counts components are related both to the structure and physico-chemical state of an implant surface. The aim of this study was to determine haemocompatibility and cytotoxicity of polysulfone-based samples containing nano and micro particles of magnetite (Fe3O4). METHODS: The polysulfone-based samples modified with nanometric and micrometric magnetite particles were examined. Physicochemical properties of the composites were determined by testing their wettability and surface roughness. The action of haemolytic, activation of coagulation system and cytotoxicity of composites was evaluated. RESULTS: Wettability and roughness of materials were correlated with nanoparticles and microparticles content. In the tests of plasma coagulation system shortening of activated partial thromboplastin time for polysulfone with nano magnetite and with micro magnetite particles was observed in comparison with pure polysulfone. Prothrombine time and thrombine time values as well as fibrinogen concentration were unchanged. Haemolysis values were normal. Morphology and viability of cells were normal. CONCLUSIONS: Composites made from polysulfone modified with nanoparticles and microparticles of magnetite cause neither haemolytic nor cytotoxic reaction. These composites evoke plasma endogenous system activation.

On the influence of various physicochemical properties of the CNTs based implantable devices on the fibroblasts' reaction in vitro.

Coating the material with a layer of carbon nanotubes (CNTs) has been a subject of particular interest for the development of new biomaterials. Such coatings, made of properly selected CNTs, may constitute an implantable electronic device that facilitates tissue regeneration both by specific surface properties and an ability to electrically stimulate the cells. The goal of the presented study was to produce, evaluate physicochemical properties and test the applicability of highly conductible material designed as an implantable electronic device. Two types of CNTs with varying level of oxidation were chosen. The process of coating involved suspension of the material of choice in the diluent followed by the electrophoretic deposition to fabricate layers on the surface of a highly biocompatible metal-titanium. Presented study includes an assessment of the physicochemical properties of the material's surface along with an electrochemical evaluation and in vitro biocompatibility, cytotoxicity and apoptosis studies in contact with the murine fibroblasts (L929) in attempt to answer the question how the chemical composition and CNTs distribution in the layer alters the electrical properties of the sample and whether any of these properties have influenced the overall biocompatibility and stimulated adhesion of fibroblasts. The results indicate that higher level of oxidation of CNTs yielded materials more conductive than the metal they are deposited on. In vitro study revealed that both materials were biocompatible and that the cells were not affected by the amount of the functional group and the morphology of the surface they adhered to.

Titanium coated with functionalized carbon nanotubes--a promising novel material for biomedical application as an implantable orthopaedic electronic device.

The aim of the study was to fabricate titanium (Ti) material coated with functionalized carbon nanotubes (f-CNTs) that would have potential medical application in orthopaedics as an implantable electronic device. The novel biomedical material (Ti-CNTs-H2O) would possess specific set of properties, such as: electrical conductivity, non-toxicity, and ability to inhibit connective tissue cell growth and proliferation protecting the Ti-CNTs-H2O surface against covering by cells. The novel material was obtained via an electrophoretic deposition of CNTs-H2O on the Ti surface. Then, physicochemical, electrical, and biological properties were evaluated. Electrical property evaluation revealed that a Ti-CNTs-H2O material is highly conductive and X-ray photoelectron spectroscopy analysis demonstrated that there are mainly COOH groups on the Ti-CNTs-H2O surface that are found to inhibit cell growth. Biological properties were assessed using normal human foetal osteoblast cell line (hFOB 1.19). Conducted cytotoxicity tests and live/dead fluorescent staining demonstrated that Ti-CNTs-H2O does not exert toxic effect on hFOB cells. Moreover, fluorescence laser scanning microscope observation demonstrated that Ti-CNTs-H2O surface retards to a great extent cell proliferation. The study resulted in successful fabrication of highly conductive, non-toxic Ti-CNTs-H2O material that possesses ability to inhibit osteoblast proliferation and thus has a great potential as an orthopaedic implantable electronic device.

Fibrous polymeric composites based on alginate fibres and fibres made of poly-ε-caprolactone and dibutyryl chitin for use in regenerative medicine.

This work concerns the production of fibrous composite materials based on biodegradable polymers such as alginate, dibutyryl chitin (DBC) and poly-ε-caprolactone (PCL). For the production of fibres from these polymers, various spinning methods were used in order to obtain composite materials of different composition and structure. In the case of alginate fibres containing the nanoadditive tricalcium phosphate (TCP), the traditional method of forming fibres wet from solution was used. However in the case of the other two polymers the electrospinning method was used. Two model systems were tested for biocompatibility. The physicochemical and basic biological tests carried out show that the submicron fibres produced using PCL and DBC have good biocompatibility. The proposed hybrid systems composed of micrometric fibres (zinc and calcium alginates containing TCP) and submicron fibres (DBC and PCL) meet the requirements of regenerative medicine. The biomimetic fibre system, the presence of TCP nanoadditive, and the use of polymers with different resorption times provide a framework with specific properties on which bone cells are able to settle and proliferate.

In vitro and in vivo studies on biocompatibility of carbon fibres.

In the present study we focused on the in vitro and in vivo evaluation of two types of carbon fibres (CFs): hydroxyapatite modified carbon fibres and porous carbon fibres. Porous CFs used as scaffold for tissues regeneration could simultaneously serve as a support for drug delivery or biologically active agents which would stimulate the tissue growth; while addition of nanohydroxyapatite to CFs precursor can modify their biological properties (such as bioactivity) without subsequent surface modifications, making the process cost and time effective. Presented results indicated that fibre modification with HAp promoted formation of apatite on the fibre surface during incubation in simulated body fluid. The materials biocompatibility was determined by culturing human osteoblast-like cells of the line MG 63 in contact with both types of CFs. Both tested materials gave good support to adhesion and growth of bone-derived cells. Materials were implanted into the skeletal rat muscle and a comparative analysis of tissue reaction to the presence of the two types of CFs was done. Activities of marker metabolic enzymes: cytochrome c oxidase (CCO) and acid phosphatase were examined to estimate the effect of implants on the metabolic state of surrounding tissues. Presented results evidence the biocompatibility of porous CFs and activity that stimulates the growth of connective tissues. In case of CFs modified with hydroxyapatite the time of inflammatory reaction was shorter than in case of traditional CFs.

Comparative in vivo biocompatibility study of single- and multi-wall carbon nanotubes.

Carbon nanotubes are expected to be of use in both genetic engineering and biomaterials engineering. In each of these potential areas of application, nanoparticles are introduced into a living organism either in the form of active biomolecule carriers or as a result of the degradation process of an implant. In the present study we focus on the in vivo behavior of two types of carbon nanotubes (single- and multi-wall nanotubes). Raman and Fourier transform infrared spectroscopy, thermogravimetric analysis and differential scanning calorimetry techniques are used to characterize the materials before introducing them into the living system. The nanotubes were implanted into the skeletal rat muscle. A comparative analysis of the tissue reaction to the presence of the two types of carbon nanotubes was made. It was observed that multi-wall carbon nanotubes were found to form large aggregates within the living tissue, while distinctly smaller particles consisting of single-wall nanotubes were easily phagocytosed by macrophages and transported to local lymph nodes.

[Study of selected biomaterials for reconstruction of septal nasal perforation]. / Ocena wybranych biomaterialów do rekonstrukcji perforacji przegrody nosa.

INTRODUCTION: The septal nasal perforation is an important problem for the laryngologists and plastic surgeons. The reasons of septal nasal perforations are injuries, neoplasm, self-mutilation, chronic rhinitis, allergy, Wegener granuloma, sarcoidosis, tuberculosis, toxic metals (arsenic, chrome), some drugs (steroids), narcotizing agents (cocaine) and complications after endoscopic and septal nasal operations. The surgical treatment, especially in the cases of large septal perforation, is often difficult because of the atrophy of nasal mucosa and lack of suitable material for reconstruction. In the surgical treatment many of methods and reconstructive materials have been used. The following autogenous tissues were used in the reconstruction of septal perforation: alloderm, temporal fascia, septal and auricle cartilage, cranial periosteum, perichondrium, ethmoidal and hip bone. The defect of such materials is progressive resorption. For many years the suitable synthetic material for septal nasal reconstruction has been searched for. Among the biomaterials the following have been used without success: Dacron, porous polyethylene, dolomite, bioglass. The rejection of synthetic material was the reason of failure. The aim of our study was to evaluate two different biomaterials with proper mechanical and biological features for nasal cartilage replacement. MATERIAL AND METHODS: We studied two types of biomaterials: biostable terpolymer PTFE/PVDF/PP and resorbable copolymer of glycolide and L-lactide (PGLA). The pilot studies were performed on two experimental animals (rabbits). The animals were operated in the general anesthesia. The biomaterials were implanted in the rabbit auricular cartilage because of its similarity to the septum and easy surgical access. Subperichondrically 1 x 1 cm fragment of the cartilage was removed. This fragment was then replaced with the biomaterial. The rabbits were painlessly sacrificed after 4 months of observation. RESULTS: A very good integration of PGLA implant with auricular cartilage was observed. In the histological examination the lack of excessive inflammatory reaction as well as no cartilage necrosis were observed. CONCLUSIONS: 4 months after implantation of PGLA in the rabbit auricular cartilage very good macroscopic and histological results were achieved.

[New composite implant for tracheal reconstruction--preliminary study]. / Nowy kompozytowy implant do rekonstrukcji tchawicy--badania wstepne.

Stenosis of trachea's diameter occurs the most often as complications after intubation and tracheotomy. Among the other reasons of narrowing of this organ the following are being named: mechanical injuries, chemical damages, primary and metastasis tumors. The therapy of trachea's stenosis includes both alternative and radical treatments. The radical treatment consists of surgical excision of the narrowed segment followed by reconstruction of trachea (anastomosing of free ends or filling of lost segment with use of auto- or allogenic material). There is no appropriate alloplastic material developed to date, the use of which in reconstructive surgery of large segment trachea defects would bring about positive long-term experimental results. The success of alloplastic material implantation in the reconstruction of large tracheal defects is limited by the fact that the biomechanical characteristics of replaced tissue are distinctly different from synthetic material. The work is devoted to evaluation of complex mechanical characteristic of natural sheep trachea before designing the proper synthetic material for large tracheal defects reconstruction. Various mechanical tests were conducted to study the mechanical behavior of total trachea. Samples of tissue cut out from trachea were also examined. The results revealed strong directional--dependent mechanical properties of trachea. Composite constituents, namely carbon fibers and biocompatible and biostable polysulfone were used to manufacture the implant.

In vitro response of macrophages to a new carbon-polylactide composite for the treatment of periodontal diseases.

The purpose of the study was to examine the response of macrophages and the concentration of selected released cytokines following contact with a new carbon-polylactide composite. The macrophages were grown on samples of the materials and on each of its components separately. Viability of the cells as well as concentrations of interleukins IL-6, IL-10, IL-12 and TNF-alpha were then determined. Some differences in the viability of the cells were demonstrated. They varied according to the kind of material used. After incubation with the serum, the composite and its components induced the release of IL-6, IL-12 and TNF-alpha which did not differ significantly from one another.