Atlantic and Mediterranean populations of the widespread serpulid Ficopomatus enigmaticus: Developmental responses to carbon nanotubes.

Ficopomatus enigmaticus was adopted as model species for ecotoxicological bioassay, with its larval development as endpoint. Two different populations of the same species, collected in areas far from each other (Mediterranean Sea and Atlantic Ocean), were exposed to multi-walled carbon nanotubes, a class of emerging pollutants with a constantly increasing relevance in the landscape of nanomaterials production. Moreover, a molecular analysis based on Cyt b amplification and sequencing, was carried out to confirm that both populations belong to the same species. The aim of the present work was to strengthen existing results about F. enigmaticus relevance in ecotoxicological bioassays, adding the variable of population effect. For both populations the concentration-response curve of effect at different toxicant concentrations was similar and, at certain concentrations, overlapping, confirming the ecological relevance of the assay. These results posed an interesting acceptance on the introduction of this species as model in ecotoxicological bioassay scenery, underlining the relevance of a widespread wild species to compare effects of chemicals and environmental samples over large distances using the same bioassay.

Design and fabrication of novel polymeric biodegradable stents for small caliber blood vessels by computer-aided wet-spinning.

Biodegradable stents have emerged as one of the most promising approaches in obstructive cardiovascular disease treatment due to their potential in providing mechanical support while it is needed and then leaving behind only the healed natural vessel. The aim of this study was to develop polymeric biodegradable stents for application in small caliber blood vessels. Poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] (PHBHHx), a renewable microbial aliphatic polyester, and poly(ε-caprolactone), a synthetic polyester approved by the US Food and Drug Administration for different biomedical applications, were investigated as suitable polymers for stent development. A novel manufacturing approach based on computer-aided wet-spinning of a polymeric solution was developed to fabricate polymeric stents. By tuning the fabrication parameters, it was possible to develop stents with different morphological characteristics (e.g. pore size and wall thickness). Thermal analysis results suggested that material processing did not cause changes in the molecular structure of the polymers. PHBHHx stents demonstrated great radial elasticity while PCL stents showed higher axial and radial mechanical strength. The developed stents resulted able to sustain proliferation of human umbilical vein endothelial cells within two weeks of in vitro culture and they showed excellent results in terms of thromboresistivity when in contact with human blood.

Vitronectin absorbed on nanoparticles mediate cell viability/proliferation and uptake by 3T3 Swiss albino mouse fibroblasts: in vitro study.

We study the interaction of 3T3 Swiss albino mouse fibroblasts with polymeric nanoparticles (NPs) and investigate cellular behaviour in terms of viability/cytotoxicity, cell cycle, NPs uptake, MAP kinase (ERK1/2), and focal adhesion kinase (FAK) activation. After incubation of NPs with cell culture media, western blot analysis showed that Vitronectin is retained by NPs, while Fibronectin is not detected. From cytotoxicity studies (MTT and BrdU methods) an LD50 of about 1.5 mg/mL results for NPs. However, NPs in the range 0.01-0.30 mg/mL are able to trigger a statistically significant increase in proliferation and cell cycle progression in dose and time depending manner. Also, biochemical evaluation of ERK1/2 and FAK clearly shows an increasing phosphorylation in a dose and time depending manner. Finally, we found by transmission electron microscopy that NPs are internalised by cells. Competitively blocking VN-integrin receptors with echistatin (1 µg/mL) results in a decrease of viability/proliferation, cell cycle progression, cellular uptake, and FAK/ERK activation showing the involvement of Vitronectin receptors in signal transduction. In conclusion, our results show that cell surface NPs interactions are mediated by absorbed plasma proteins (i.e., Vitronectin) that represent an external stimuli, switched to the nucleus by FAK enzyme, which in turn modulate fibroblasts viability/proliferation.

Processing and characterization of innovative scaffolds for bone tissue engineering.

A new protocol, based on a modified replication method, is proposed to obtain bioactive glass scaffolds. The main feature of these samples, named "shell scaffolds", is their external surface that, like a compact and porous shell, provides both high permeability to fluids and mechanical support. In this work, two different scaffolds were prepared using the following slurry components: 59 % water, 29 % 45S5 Bioglass(®) and 12 % polyvinylic binder and 51 % water, 34 % 45S5 Bioglass(®), 10 % polyvinylic binder and 5 % polyethylene. All the proposed samples were characterized by a widespread microporosity and an interconnected macroporosity, with a total porosity of 80 % vol. After immersion in a simulated body fluid (SBF), the scaffolds showed strong ability to develop hydroxyapatite, enhanced by the high specific surface of the porous systems. Moreover preliminary biological evaluations suggested a promising role of the shell scaffolds for applications in bone tissue regeneration. As regards the mechanical behaviour, the shell scaffolds could be easily handled without damages, due to their resistant external surface. More specifically, they possessed suitable mechanical properties for bone regeneration, as proved by compression tests performed before and after immersion in SBF.

Fibrin acts as biomimetic niche inducing both differentiation and stem cell marker expression of early human endothelial progenitor cells.

OBJECTIVES: Transplantation of endothelial progenitor cells (EPCs) is a promising approach for revascularization of tissue. We have used a natural and biocompatible biopolymer, fibrin, to induce cell population growth, differentiation and functional activity of EPCs. MATERIALS AND METHODS: Peripheral blood mononuclear cells were cultured for 1 week to obtain early EPCs. Fibrin was characterized for stiffness and capability to sustain cell population expansion at different fibrinogen-thrombin ratios. Viability, differentiation and angiogenic properties of EPCs were evaluated and compared to those of EPCs grown on fibronectin. RESULTS: Fibrin had a nanometric fibrous structure forming a porous network. Fibrinogen concentration significantly influenced fibrin stiffness and cell growth: 9 mg/ml fibrinogen and 25 U/ml thrombin was the best ratio for enhanced cell viability. Moreover, cell viability was significantly higher on fibrin compared to being on fibronectin. Even though no significant difference was observed in expression of endothelial markers, culture on fibrin elicited marked induction of stem cell markers OCT 3/4 and NANOG. In vitro angiogenesis assay on Matrigel showed that EPCs grown on fibrin retain angiogenetic capability as EPCs grown on fibronectin, but significantly better release of cytokines involved in cell recruitment was produced by EPC grown on fibrin. CONCLUSION: Fibrin is a suitable matrix for EPC growth, differentiation and angiogenesis capability, suggesting that fibrin gel may be very useful for regenerative medicine.

Materials degradation in PVC medical devices, DEHP leaching and neonatal outcomes.

Polymeric materials play a key role in the production of medical and clinical devices thanks to their special features such as flexibility, easy processing and good price/performance ratio. Among the different polymeric matrixes, one of the most used is Poly(vinyl chloride) (PVC). At room temperature PVC is hard and brittle, thus great amounts (40-50%) of phthalate esters that act as plasticizers are added to the polymer to make it flexible and appropriate for medical use. Di-(2-ethylhexyl)-phthalate (DEHP) is the most widely used plasticizer in PVC medical devices. However, DEHP is not chemically bound to PVC and migrates from medical devices with time and use. The potential for DEHP to produce adverse effects in humans has been the subject of considerable discussion and debate in the scientific community. In particular, newborns in the new environment have to be considered at particularly increased risk, because of their small body size and the multiple medical device-related to the DEHP exposure. The major factors determining the degree to which DEHP migrates from medical devices are temperature, amount of DEHP in the device, storage time, shaking of the device while in contact with the medical solutions and degree of PVC degradation.

Investigation of interactions between poly-L-lysine-coated boron nitride nanotubes and C2C12 cells: up-take, cytocompatibility, and differentiation.

Boron nitride nanotubes (BNNTs) have generated considerable interest within the scientific community by virtue of their unique physical properties, which can be exploited in the biomedical field. In the present in vitro study, we investigated the interactions of poly-l-lysine-coated BNNTs with C2C12 cells, as a model of muscle cells, in terms of cytocompatibility and BNNT internalization. The latter was performed using both confocal and transmission electron microscopy. Finally, we investigated myoblast differentiation in the presence of BNNTs, evaluating the protein synthesis of differentiating cells, myotube formation, and expression of some constitutive myoblastic markers, such as MyoD and Cx43, by reverse transcription - polymerase chain reaction and Western blot analysis. We demonstrated that BNNTs are highly internalized by C2C12 cells, with neither adversely affecting C2C12 myoblast viability nor significantly interfering with myotube formation.

Preparation of stable dispersion of barium titanate nanoparticles: Potential applications in biomedicine.

Nanoscale structures and materials have been explored in many biological applications because of their extraordinary novel properties. Here we propose a study of cellular interactions with barium titanate nanoparticles, an interesting ceramic material that has received a lot of interest in the nanotechnology research, but without any attention about its biological potential. We introduced for the first time an efficient method for the preparation of stable aqueous dispersions of barium titanate nanoparticles, characterized with FIB, TEM and AFM imaging, light scattering, Z-potential and UV/vis analysis. Finally, we presented a systematic study of short-term cytotoxicity of the prepared dispersion based both on quantitative (metabolism, proliferation) and qualitative (apoptosis, viability, differentiation) assays.

Development of a bioactive glass fiber reinforced starch-polycaprolactone composite.

For bone regeneration and repair, combinations of different materials are often needed. Biodegradable polymers are often combined with osteoconductive materials, such as bioactive glass (BaG), which can also improve the mechanical properties of the composite. The aim of this work was to develop and characterize BaG fiber reinforced starch-poly-epsilon-caprolactone (SPCL) composite. Sheets of SPCL (30/70 wt %) were produced using single-screw extrusion. They were then cut and compression-molded in layers with BaG fibers to form composite structures with different combinations. Mechanical and degradation properties of the composites were studied. The actual amount of BaG in the composites was determined using combustion tests. Initial mechanical properties of the reinforced composites were at least 50% better than the properties of the nonreinforced specimens. However, the mechanical properties of the composites after 2 weeks of hydrolysis were comparable to those of the nonreinforced samples. During the 6 weeks hydrolysis the mass of the composites had decreased only by about 5%. The amount of glass in the composites remained as initial for the 6-week period of hydrolysis. In conclusion, it is possible to enhance initial mechanical properties of SPCL by reinforcing it with BaG fibers. However, mechanical properties of the composites are typical for bone fillers and strength properties need to be further improved for allowing more demanding bone applications.

Biodegradable nanomats produced by electrospinning: expanding multifunctionality and potential for tissue engineering.

With increasing interest in nanotechnology, development of nanofibers (n-fibers) by using the technique of electrospinning is gaining new momentum. Among important potential applications of n-fiber-based structures, scaffolds for tissue-engineering represent an advancing front. Nanoscaffolds (n-scaffolds) are closer to natural extracellular matrix (ECM) and its nanoscale fibrous structure. Although the technique of electrospinning is relatively old, various improvements have been made in the last decades to explore the spinning of submicron fibers from biodegradable polymers and to develop also multifunctional drug-releasing and bioactive scaffolds. Various factors can affect the properties of resulting nanostructures that can be classified into three main categories, namely: (1) Substrate related, (2) Apparatus related, and (3) Environment related factors. Developed n-scaffolds were tested for their cytocompatibility using different cell models and were seeded with cells for to develop tissue engineering constructs. Most importantly, studies have looked at the potential of using n-scaffolds for the development of blood vessels. There is a large area ahead for further applications and development of the field. For instance, multifunctional scaffolds that can be used as controlled delivery system do have a potential and have yet to be investigated for engineering of various tissues. So far, in vivo data on n-scaffolds are scarce, but in future reports are expected to emerge. With the convergence of the fields of nanotechnology, drug release and tissue engineering, new solutions could be found for the current limitations of tissue engineering scaffolds, which may enhance their functionality upon in vivo implantation. In this paper electrospinning process, factors affecting it, used polymers, developed n-scaffolds and their characterization are reviewed with focus on application in tissue engineering.

Development of diclofenac sodium releasing bio-erodible polymeric nanomats.

Application of nanofiber-based nanomats in medicine is attractive and thanks to the 3D nanostructure and the high surface to volume ratio they are excellent for local controlled drug delivery. The use of bioactive bioerodible polymers for developing drug delivery nanomats may allow for drug release and targeting control. Objective of the current study was to evaluate the suitability of bioerodible polymeric material based on n-butyl hemiester of [poly(maleic anhydride-alt-2-methoxyethyl vinyl ether)] (PAM14) for the preparation of nanomats for controlled administration of anti-inflammatory, diclofenac sodium (DS) drug. Samples were prepared using different polymer concentrations (5-10%) in either ethanol or acetic acid as solvent. Morphology was investigated by using scanning electron microscopy (SEM). Thermal analysis such as differential scanning calorimetry (DSC) was performed to detect effect on polymer arrangement. DS localization in electrospun nanomats was evaluated by using electron back scattering microanalysis, based on the detection of chlorine, and drug release kinetics was assessed using UV-Vis. Average fiber diameter resulted in the range of 100 nm to 1.0 microm and a homogeneous distribution of the loaded drug into the fibers was observed. The DS release was immediate and despite the preliminary nature of the performed electrospinning experiments, the achieved results appear promising for the future development of a novel system for the controlled and targeted administration of drug and active agent.

Nanoparticle systems for the targeted release of active principles of proteic nature.

The preparation and characterization of nanoparticles based on biodegradable/bioerodible polymers is reported. They have been designed for the controlled-targeted release of proteic drugs such as alpha-interferon and for the release of active principles in tissue engineering. The amenability of some of the prepared polymeric matrices to be used in the fabrication of micro and nano patterned scaffolds is also described.

Endothelial cell adhesion on bioerodable polymers.

This paper presents the results of a preliminary screening of a new class of bioerodable polymers, partial esters of alternating copolymers of maleic anhydride and mono-methoxyoligoethyleneglycol vinyl ethers (PAM) for use in engineered vascular tissue. Different initial concentrations of PAM and human serum albumin (HSA) were spin-coated onto glass substrates and the surface properties of the resulting films and their relationship to endothelial cell adhesion was examined. An optimum PAM/HSA blend for use as the cell contact surface of a bioerodable scaffold was identified.

A 3D model for the human hepatic asialoglycoprotein receptor (ASGP-R).

The human hepatic Asialoglycoprotein Receptor (ASGP-R) consists of two different types of liver specific membrane glycoproteins that bind to terminal galactose and N-acetylgalactosamine residues of serum glycoproteins. The two different polypeptide chains are referred to as two receptor subunits, HH1 and HH2, which are both involved in the activity of the functional receptor. This receptor has served as a model for understanding receptor-mediated endocytosis and carbohydrate mediated recognition phenomena. Here models for the C-terminal extracellular region of both HH1 and HH2 subunit are presented. The standard homology building procedure was modified in order to make it suitable for the modeling problem at hand. The models for the extracellular regions of HH1 and HH2 were initially constructed by exploiting several fragments, belonging to proteins of known 3D structure, and showing high local sequence similarity with respect to the glycoproteins of interest. Putative binding sites were first hypothesized on the basis of the comparison with other complexes of lectins, the crystal structure of which was available in the Protein Data Bank. A model for the complex involving the HH2 subunit and the typical high affinity ligand N-acetylgalactosamine (NacGal) was refined as the first by a suitable combination of MD simulations and Energy Minimization calculations, since it seemed to quickly converge to a plausible structure. An intermediate model for HH1 was then rebuilt on the basis of the refined model for HH2. It was then submitted to a sequence of molecular dynamics simulations with templates which took into account the secondary structure prediction for a final refinement. The structures of small regions of the models, located around the binding sites, were compared with more recent crystallographic data regarding a complex involving the mutant of Mannose Binding Protein QPDWGH (1BCH entry in the Protein Data Bank) and NacGal. This mutant shows high local sequence similarity with HH1 and HH2 at the binding sites. On the basis of the above comparison, different locations of the binding sites were also considered. In addition to other expected interactions, two hydrophobic interactions were observed in the models with Trp residues (positions 243 in HH1 and 181 or 267 in HH2 respectively) and His residues (positions 256 in HHI and 184 in HH2.respectively). The quality of the models was evaluated by the Procheck program and they seemed plausible. This observation together with analogies found between binding sites of the models and IBCH supported the validity of the models. A further validation element arose by comparison between experimental binding data available in the literature about the homologous rat receptor subunits and theoretical interaction energies evaluated, by means of the DOCK 3.5 program, in models for the rat subunits obtained from the corresponding human ones. The new modeling procedure used here appears to be a well-suited method for structural analysis of small regions, located around the ligands, in proteins of unknown 3D structure.

[Our experience in the treatment of idiopathic osteonecrosis of the femur head by évidement and filling with bone grafts]. / La nostra esperienza nel trattamento con svuotamento e borraggio della osteonecrosi idiopatica della testa femorale.

UNLABELLED: The authors relate their experience in treating idiopathic osteonecrosis of the femoral head with bone grafting. CASES: 18 hips under control for a period varying from 8 to 3 years. Radiographic and clinical assessment before and after surgery according to Ficat, Arlet and Merle D'Aubigné. Four patients were affected with idiopathic osteonecrosis of the femoral head. Results were very good in 4 cases and quite good in 2 cases. Only one third of the results could be regarded as positive. This modest achievement is partly due to the method employed. In order to obtain better results, however, an early diagnosis is required and patients who are operated on should have quite a good bone mineralization.