Fluctuation Spectroscopy Analysis of Glucose Capped Gold Nanoparticles.

In this work, we report the synthesis and biophysical studies carried out on a new kind of biocompatible and very stable gold nanoparticle (GNP) stabilized with glucose through a PEG linker (AuNP-PEG-Glu). The synthetic path was optimized to obtain nanoparticles of controlled sizes. ζ-potential and dynamic light scattering measurements allowed assessment of the nanodimension, dispersity, surface charge, and stability of our GNPs. Confocal microscopy demonstrated qualitatively that glucose molecules are successfully bonded to GNP surfaces. For our study, we selected nanoparticles with diameter in a range that maximizes the internalization efficiency in cells (40 nm). A detailed investigation about the biophysical proprieties of AuNP-PEG-Glu was carried out by means of fluorescence correlation spectroscopy (FCS) and orbital tracking techniques. This work gives new insights about the uptake mechanism of gold nanoparticles capped with glucose molecules.

A model study for tethering of (bio)active molecules to biomaterial surfaces through arginine.

A new approach for tethering of bioactive molecules via arginine is proposed and validated on collagen 2D matrices. The method involves the introduction of a methyl ketone on arginine side-chains, followed by reaction with model alkoxyamino derivatives.

Driving h-osteoblast adhesion and proliferation on titania: peptide hydrogels decorated with growth factors and adhesive conjugates.

Hydrogels from self-assembling ionic complementary peptides have been receiving much interest from the scientific community as mimetics of the extracellular matrix that can offer three-dimensional support for cell growth or become vehicles for the delivery of stem cells or drugs. These scaffolds have also been proposed as bone substitutes for small defects as they promote beneficial effects on human osteoblasts. In order to develop a novel bioactive titanium implant, we propose the introduction of a layer of ionic-complementary self-assembling peptides (EAbuK) on Ti whose surface has been previously sandblasted and acid etched. The peptide layer is anchored to the metal by covalent functionalization of titania with self-assembling sequences. The peptide layer has also been enriched by the insulin-like growth factor-1 incorporated to the layer and/or a conjugate obtained by chemoselective ligation between EAbuK and a sequence of 25 residues containing four GRGDSP motifs per chain. X-ray photoelectron spectroscopy studies confirmed a change in the surface composition in agreement with the proposed decorations. An evaluation of the contact angle showed a substantial change in wettability induced by the peptide layer. The human osteoblast adhesion and proliferation assays showed an increase in adhesion for the surfaces enriched with conjugate at a concentration of 3.8 × 10(-7)m and an enhanced proliferation for samples enriched with insulin-like growth factor-1 at the highest concentration tested (2.1 × 10(-5)m).

Silver nanoparticles linked by a Pt-containing organometallic dithiol bridge: study of local structure and interface by XAFS and SR-XPS.

Silver nanoparticles (AgNPs) functionalized with an organometallic bifunctional thiol containing Pt(ii) centers, generated in situ from trans-trans-[thioacetyl-bistributylphosphine-diethynylbiphenyl-diplatinum(ii)], were synthesized with different sulphur/metal molar ratios (i.e. AgNPs-1 and AgNPs-2) with the aim to obtain nanosystems of different mean size and self-organization behaviour. AgNPs spontaneously self-assemble, giving rise to 2D networks, as previously assessed. In this work a deeper insight into the chemico-physical properties of these AgNPs is proposed by means of synchrotron radiation induced X-ray photoelectron spectroscopy (SR-XPS) and X-ray absorption fine structure spectroscopy (XAFS) techniques. The results are discussed in order to probe the interaction at the interface between a noble metal and a thiol ligand at the atomic level and the aim of this study is to shed light on the chemical structure and self-organization details of nanosystems. The nature of the chemical interaction between the dithiol ligand and the Ag atoms on the nanoparticle surface was investigated by combining SR-XPS (S2p, Ag3d core levels) and XAS (S and Ag K-edges) analysis. UV-visible absorption and emission measurements were also carried out on all samples and compared with TD-DFT calculations so as to get a better understanding of their optical behavior and establish the nature of the excitation and emission processes.

Differential phytotoxic effect of silver nitrate (AgNO3) and bifunctionalized silver nanoparticles (AgNPs-Cit-L-Cys) on Lemna plants (duckweeds).

Duckweeds are aquatic plants often used in phytotoxic studies for their small size, simple structure, rapid growth, high sensitivity to pollutants and facility of maintaining under laboratory conditions. In this paper, induced phytotoxic effects were investigated in Lemna minor and Lemna minuta after exposition to silver nitrate (AgNO3) and silver nanoparticles stabilized with sodium citrate and L-Cysteine (AgNPs-Cit-L-Cys) at different concentrations (0, 20 and 50 mg/L) and times (7 and 14 days). Lemna species responses were evaluated analyzing plant growth (mat thickness, fresh and dry biomass, relative growth rate - RGR) and physiological parameters (chlorophyll - Chl, malondialdehyde - MDA, ascorbate peroxidase - APX and catalase - CAT). Ag content was measured in the fronds of the two Lemna species by inductively coupled plasma optical emission spectrometry. AgNO3 and AgNPs-Cit-L-CYs produced phytotoxic effects on both duckweed species (plant growth and Chl reduction, MDA increase) that enhanced in response to increasing concentrations and exposure times. AgNPs-Cit-L-Cys caused much less alteration in the plants compared to AgNO3 suggesting that the presence of bifunctionalized AgNPs-Cit-L-Cys have a reduced phytotoxic effect as compared to Ag+ released in water. Based on the physiological performance, L. minuta plants showed a large growth reduction and higher levels of chlorosis and stress in respect to L. minor plants, probably due to greater Ag+ ions accumulation in the fronds. Albeit with some differences, both Lemna species were able to uptake Ag+ ions from the aqueous medium, especially over a period of 14 days, and could be considered adapt as phytoremediation agents for decontaminating silver ion-polluted water.

Biofunctionalization of TiO2 surfaces with self-assembling oligopeptides in different pH and Ionic Strength conditions: Charge effects and molecular organization.

Self-assembling peptides (SAPs) were investigated by means of XPS and Angular Dependent NEXAFS spectroscopies, with the aim to probe the influence of pH and Ionic Strength conditions on the chemical structure and molecular organization of SAPs anchored on titania surfaces. XPS at the C1s, N1s, O1s core levels allowed to study surfaces and biomolecule/substrate interfaces. NEXAFS data allowed ascertaining that SAPs molecular structure is preserved upon grafting to the titania surface. Angular Dependent NEXAFS was used to investigate the influence of environmental conditions on the molecular organization behaviour. The objective of our study was to establish a set of methodologies for obtaining arrangements of well-organized biomolecules on scaffolds surfaces as a basic technology to develop and optimize cells adhesion and proliferation for tissue engineering applications.

Understanding the biomimetic properties of gallium in Pseudomonas aeruginosa: an XAS and XPS study.

Pyochelin (PCH) is a siderophore (extracellular chelator) produced by the pathogenic bacterium Pseudomonas aeruginosa (PAO). PCH is implicated in iron (Fe3+) transport to PAO, and is crucial for its metabolism and pathogenicity. Due to the chemical similarity with Fe3+, gallium (Ga3+) interferes with vital iron-dependent processes in bacterial cells, thereby opening new perspectives for the design of specific metal-based antibacterial drugs. However, the structural basis for the Fe3+-mimetic properties of Ga3+ complexed with the PCH siderophore is still lacking. A precise knowledge of the coordination chemistry at the metal site is one of the topmost issues in the production of novel biomimetic metal-based drugs. Elucidation of this issue by means of a deep structural spectroscopic investigation could lead to an improved interference with, or a specific inhibition of, relevant biological pathways. For this reason, we applied Synchrotron Radiation induced X-ray Photoelectron Spectroscopy (SR-XPS) and X-ray Absorption Spectroscopy (XAS) to probe the electronic nature and coordination chemistry of Fe3+ and Ga3+ coordinative sites in PCH metal complexes. Combined XAFS and SR-XPS studies allow us to demonstrate that both Fe and Ga have the same valence state in Fe-PCH and Ga-PCH, and have the same octahedral coordination geometry. Moreover, a similar next neighbour distribution for Fe and Ga, resulting from the EXAFS data analysis, strongly supports similar coordination chemistry at the origin of the biomimetic behaviour of Ga.

Synthesis of functionalized gold nanoparticles capped with 3-mercapto-1-propansulfonate and 1-thioglucose mixed thiols and "in vitro" bioresponse.

The synthesis, characterization and assessment of biological behavior of innovative negatively charged functionalized gold nanoparticles is herein reported, for potential applications in the field of radiotherapy and drug delivery. Gold nanoparticles (AuNPs) functionalized with two capping agents, i.e., the 3-mercapto-1-propansulfonate (3-MPS) and 1-ß-thio-D-glucose (TG), have been on purpose synthesized and fully characterized. Advanced characterization techniques including X-Ray Photoelectron Spectroscopy (XPS) were applied to probe the chemical structure of the synthesized nanomaterials. Z-potential and Dynamic Light Scattering measurements allowed assessing the nanodimension, dispersity, surface charge and stability of AuNPs. Transmission Electron Microscopy (TEM) and Flame Atomic Absorption Spectroscopy (FAAS) were applied to the "in vitro" HSG cell model, to investigate the nanoparticles-cells interaction and to evaluate the internalization efficiency, whereas short term cytotoxicity and long term cell killing were evaluated by means of MTT and SRB assays, respectively. In conclusion, in order to increase the amount of gold atoms inside the cell we have optimized the synthesis for a new kind of biocompatible and very stable negatively charged TG-functionalized nanoparticles, with diameters in a range that maximize the uptake in cells (i.e., ∼15nm). Such particles are very promising for radiotherapy and drug delivery application.

Bioconjugation of gold-polymer core-shell nanoparticles with bovine serum amine oxidase for biomedical applications.

Core-shell gold nanoparticles [AuNPs], stabilized with a hydrophilic polymer, poly(3-dimethylammonium-1-propyne hydrochloride) [PDMPAHCl], have been used for the immobilization of bovine serum amine oxidase [BSAO]. The functionalized surface of the hybrid nanoparticles is pH responsive, due to the presence of aminic groups that carry out a double role: on one hand they act as ligands for the gold nanoparticle surface, allowing the colloidal stabilization and, on the other hand, they give a hydrophilic characteristic to the whole colloidal suspension. The core-shell nanoparticles [Au@PDMPAHCl] have been characterized by using UV-vis and X-ray photoelectron spectroscopy, DLS, ζ-potential measurements and by FE-TEM microscopy. BSAO enzyme can be loaded by non-covalent immobilization onto Au@PDMPAHCl nanoparticles up to 70% in weight, depending on the pH values of the environmental medium. Activity tests on Au@PDMPAHCl-BSAO bioconjugates confirm an enzymatic activity up to 40%, with respect to the free enzyme activity. Moreover, our results show that loading and enzymatic activity are rather interrelated characteristics and that, under appropriate polymer concentration and pH conditions, a satisfactory compromise can be reached. These results, as a whole, indicate that Au@PDMPAHCl-BSAO bioconjugate systems are promising for future biomedical applications.

Urgent Carotid Surgery: Is It Still out of Debate?

Patients with symptomatic tight carotid stenosis have an increased short-time risk of stroke and an increased long-term risk of ischaemic vascular events compared with the general population. The aim of this study is to assess the safety, efficacy, and limitations of urgent CEA or CAS, in patients with carotid stenosis greater than 70% and clinically characterized by recurrent TIA or brain damage following a stroke (<2.5 cm). This study involved 28 patients divided into two groups. Group A consisted of sixteen patients who had undergone CEA, and group B consisted of twelve patients who had undergone CAS. Primary endpoints were mortality, neurological morbidity (by NIHSS) and postoperative hemorrhagic cerebral conversion, at 30 days. Ten patients (62.5%) of group A experienced an improvement in their initial neurological deficit while in 4 cases (26%) the deficit remained stable. Two cases of neurologic mortality are presented. At 1 month, 9 patients (75%) of group B experienced an improvement in their initial neurological deficit while 3 patients (25%) had a neurological impairment. Urgent or deferred surgical or endovascular treatment have a satisfactory outcome considering the profile in very high-risk patient population. Otherwise in selected patients CEA seems to be preferred to CAS.

Peptide/TiO2 surface interaction: a theoretical and experimental study on the structure of adsorbed ALA-GLU and ALA-LYS.

The adsorption on the TiO(2) surface of two dipeptides AE (L-alanine-L-glutamic acid) and AK (L-alanine-L-lysine), that are "building blocks" of the more complex oligopeptide EAK16, has been investigated both theoretically and experimentally. Classical molecular dynamics simulations have been used to study the adsorption of H-Ala-Glu-NH(2) and H-Ala-Lys-NH(2) dipeptides onto a rutile TiO(2) (110) surface in water solution. Several peptide conformers have been considered simultaneously upon the surface. The most probable contact points between the molecules and the surface have been identified. Carbonyl oxygens as well as nitrogen atoms are possible Ti coordination points. Local effects are responsible for adsorption and desorption events. Self-interaction effects can induce molecular reorientations giving less strongly adsorbed species. The chemical structure and composition of thin films of the two dipeptides AE and AK on TiO(2) were investigated by XPS (X-ray photoelectron spectroscopy) measurements at both O and N K-edges. Theoretical ab initio calculations (DeltaSCF) were also performed to simulate the spectra, allowing for a direct comparison between experiment and theory.

The haemodynamic effect of internal carotid artery stenosis on cerebral perfusion during aortic surgery.

OBJECTIVES: to determine the impact of the extracranial internal carotid stenosis on cerebral perfusion during aortoiliac surgery. DESIGN: prospective study. MATERIAL AND METHODS: of 432 consecutive patients undergoing aortoiliac reconstruction, 16/86 (18%) with >70% internal carotid artery stenosis, underwent inverted surgical timing (aortic reconstruction first and carotid endarterectomy second). Preoperative Transcranial Doppler (TCD) with and without acetazolamide was used to evaluate cerebrovascular reserve capacity (CRC). Intraoperatively, middle cerebral artery flow velocity (mean MCAv) and systemic blood pressure (SBP) were recorded. RESULTS: preoperatively, all 16 patients had good CRC (increase in mean MCAv: 66% right and 72% left). Intraoperatively, the mean MCAv (from 49+/-13 to 45+/-14 cm/s p=0.0249) and SBP decreased (from 127+/-25 to 113+/-22 mmHg p=0.0016). In patients with unilateral carotid disease, declamping had no effect on left mean MCAv despite a significant decrease of SBP (129+/-44 to 113+/-21 mmHg p=0.0211). In those with bilateral disease, declamping decreased both mean MCAv: from (48+/-12 to 39+/-10 cm/s p=0.011) and SBP (123+/-26 to 111+/-25 mmHg p=0.0479). No perioperative neurological deficit occurred. CONCLUSIONS: if CRC is normal or still effective, aortoiliac reconstruction does not impair cerebral perfusion.

Somatosensory evoked potentials versus locoregional anaesthesia in the monitoring of cerebral function during carotid artery surgery: preliminary results of a prospective study.

OBJECTIVE: to relate changes in somatosensory-evoked potentials (SEPs) with onset of neurological deficits in patients having carotid endarterectomy (CEA) under locoregional anaesthesia. METHODS: a prospective study of 50 consecutive patients. RESULTS: SEPs yielded an accuracy of 98%, specificity 100%, and sensitivity 89%. In all concordant cases the onset of a neurological deficit in awake patients corresponded to a 30--40% reduction in amplitude of N20-P25 waveforms. After shunting, the N20-P25 took 2--3 min to return to normal. CONCLUSIONS: SEPs are associated with a 2% false negative rate. Their threshold for detecting cerebral ischaemia is lower than the currently reported value for patients under general anaesthesia. The time needed for evoked potentials (2--3 min) to return to normal after shunting limits their usefulness in verifying effective shunting.