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Preparation of biodegradable, surface engineered PLGA nanospheres with enhanced lymphatic drainage and lymph node uptake.

PURPOSE: Nanospheres can be utilised for the targeting of drugs and diagnostic agents to the regional lymph nodes. The surface modification of model polystyrene, (PS), and poly(lactide-co-glycolide),(PLGA), nanospheres by poly(lactide)-poly(ethylene glycol), (PLA:PEG), copolymers has been assessed by in vitro characterisation and in vivo biodistribution studies following subcutaneous administration of the nanospheres to the rat. METHODS: Three PLA:PEG copolymers were investigated, with PEG chain lengths of 750, 2000 and 5000 Da. The PLA:PEG copolymers were either coated onto the surface of PS and PLGA nanospheres or used as a co-precipitate in the formation of PLGA-PLA:PEG nanospheres. Coating of the nanospheres was confirmed by an increase in their particle size and a corresponding decrease in the surface potential. The kinetics of injection site drainage and lymph node retention was determined over a 24 hour time course for naked, coated and co-precipitated nanosphere systems. RESULTS: Dependent on the surface characteristics, the distribution of the nanospheres can be significantly modified and the lymph node localisation dramatically enhanced by coating their surfaces with PLA:PEG copolymers or by producing co-precipitate nanospheres of PLGA and PLA:PEG. CONCLUSIONS: A fully biodegradable nanosphere system has been developed with excellent lymph node targeting characteristics.

Animals↗

Oligosaccharides on living human neuroblastoma cells of dissimilar degrees of differentiation. A flow-cytometric study with sugar-specific lectins and glycosidases.

The present study is an attempt to correlate cell-surface saccharide composition and/or disposition with malignant behavior and differentiation of two established human neuroblastoma sublines. The methodology applied was quantitative flow-cytometric evaluation of binding data for sugar-specific lectins in conjunction with cell-surface modification by specific glycosidases. The relevant parameters were both the number of binding sites and their apparent affinity constants for the respective lectins on native cells as well as the expected shift of those values after sequential treatment with specific glycosidases. The main conclusions from the findings may be summarized as follows: 1. There appears to exist a correlation between differentiation and/or maturation of neural cells and their cell-surface oligosaccharide patterns, as deduced indirectly by the biophysical approach of quantitative evaluation of lectin-binding data. More specifically, our findings support the hypothesis of a strong correlation between the degree of sialylation of terminal saccharide structures and the relative immaturity and/or lack of differentiation of the respective cells by morphological and biochemical criteria. 2. The combined application of specific lectins and glycosidases should be further exploited for similar purposes since it yields unequivocal information, provided that all biochemical and biophysical methods are scrutinized for their specificity. 3. Flow cytometry with fluorescence-labeled lectins is especially suited for the purposes mentioned since it allows quantitative binding studies to be conducted in a quick and uncomplicated manner. Most importantly, these data can be derived from intact living cells.

Binding Sites↗

High-density assembly of gold nanoparticles on multiwalled carbon nanotubes using 1-pyrenemethylamine as interlinker.

In this article, we describe the formation of carbon nanotube (CNT)-gold nanoparticle composites in aqueous solution using 1-pyrenemethylamine (Py-CH2NH2) as the interlinker. The alkylamine substituent of 1-pyrenemethylamine binds to a gold nanoparticle, while the pyrene chromophore is noncovalently attached to the sidewall of a carbon nanotube via pi-pi stacking interaction. Using this strategy, gold nanoparticles with diameters of 2-4 nm can be densely assembled on the sidewalls of multiwalled carbon nanotubes. The formation of functionalized gold nanoparticles and CNT-Au nanoparticle composites was followed by UV-vis absorption and luminescence spectroscopy. After functionalization of gold nanoparticles with 1-pyrenemethylamine, the distinct absorption vibronic structure of the pyrene chromophore was greatly perturbed and its absorbance value was decreased. There was also a corresponding red shift of the surface plasmon resonance (SPR) absorption band of the gold nanoparticles after surface modification from 508 to 556 nm due to interparticle plasmon coupling. Further reduction of the pyrene chromophore absorbance was observed upon formation of the CNT-Au nanoparticle composites. The photoluminescence of 1-pyrenemethylamine was largely quenched after attaching to gold nanoparticles; formation of the CNT-Au nanoparticle composites further lowered its emission intensity. The pyrene fluoroprobe also sensed a relatively nonpolar environment after its attachment to the nanotube surface. The present approach to forming high-density deposition of gold nanoparticles on the surface of multiwalled carbon nanotubes can be extended to other molecules with similar structures such as N-(1-naphthyl)ethylenediamine and phenethylamine, demonstrating the generality of this strategy for making CNT-Au nanostructure composites.

Journal Article↗

Effect of alkaline treatment of pure titanium and its alloys on the bonding strength of dental veneering resins.

Commercially pure titanium (cpTi), Ti6Al4V, an experimental beta-type titanium (Ti 53.4 wt%, Nb 29 wt %, Ta 13 wt %, and Zr 4.6 wt %), and 12% AuPdAg alloy plates were sandblasted, cleaned in water, and dried. cpTi plates were treated with nine alkaline treatments that differed in the type of alkali, alkaline concentration, soaking temperature, soaking time, and heating temperature. cpTi plates that were only sandblasted or sandblasted and oxidized at 600 degrees C for 1 h in air were also prepared. Finally, the bonding strengths of 11 kinds of surface-treated cpTi to resin were measured using a pull-shear bonding method after immersion in physiologic saline solution at 37 degrees C for 24 h. The bonds of the standard alkaline-treated cpTi and two titanium alloys to resins were 1.5-1.9 times stronger than those of sandblasted specimens (p < 0.01), but no significant effects of the alkaline treatment were observed on the 12% AuPdAg alloy. The greatest bonding strengths were found for cpTi treated with NaOH and KOH and then heated at 600 degrees C (p < 0.01). In conclusion, alkaline treatment is a simple, effective surface modification of titanium that improves bonding to veneering resin.

Alloys↗

In vitro growth and differentiation of osteoblast-like human bone marrow cells on glass reinforced hydroxyapatite plasma-sprayed coatings.

Human osteoblastic bone marrow cells were cultured for periods of up to 28 days in control conditions and on the surface of a glass reinforced hydroxyapatite composite (HA/G1) and commercial hydroxyapatite (HA) plasma-sprayed coatings, in the "as-received" condition and after immersion treatment in culture medium for 21 days. Cultures were characterized for total protein content and alkaline phosphatase activity. Scanning electron microscope analyses were performed on control cultures, seeded materials and materials incubated in the absence of cells. Culture media were analyzed for total and ionized calcium and phosphorus concentrations throughout the incubation period. Immersion of HA/G1 and HA coatings in culture medium resulted in significant alterations to the levels of calcium and phosphorus in the medium, leading to surface modifications. However, seeded material samples showed significant differences in the pattern of variation of the levels of these species. Cell proliferation was observed in the "as-received" HA/G1 composite, but cell mediated formation of mineral deposits was not proved. In contrast, "as-received" HA hardly supported cell growth. Previously immersed material samples showed cell proliferation and evidence of biological formation of mineral deposits. However, the HA/G1 composite presented better surface characteristics for cell growth as the behavior of bone marrow cells was closer to that observed in control cultures.

Journal Article↗

Fretting wear study of surface modified Ni-Ti shape memory alloy.

A combination of shape memory characteristics, pseudoelasticity, and good damping properties make near-equiatomic nickel-titanium (Ni-Ti) alloy a desirable candidate material for certain biomedical device applications. The alloy has moderately good wear resistance, however, further improvements in this regard would be beneficial from the perspective of reducing wear debris generation, improving biocompatibility, and preventing failure during service. Fretting wear tests of Ni-Ti in both austenitic and martensitic microstructural conditions were performed with the goal of simulating wear which medical devices such as stents may experience during surgical implantation or service. The tests were performed using a stainless steel stylus counter-wearing surface under dry conditions and also with artificial plasma containing 80 g/L albumen protein as lubricant. Additionally, the research explores the feasibility of surface modification by sequential ion implantation with argon and oxygen to enhance the wear characteristics of the Ni-Ti alloy. Each of these implantations was performed to a dose of 3 x 10(17) atom/cm(2) and an energy of 50 kV, using the plasma source ion implantation process. Improvements in wear resistance were observed for the austenitic samples implanted with argon and oxygen. Ion implantation with argon also reduced the surface Ni content with respect to Ti due to differential sputtering rates of the two elements, an effect that points toward improved biocompatibility.

Journal Article↗

Osteoblast growth promotion by protein electrostatic self-assembly on biodegradable poly(lactide).

Extracellular matrix (ECM)-like coating was developed on biodegradable biomaterials based on the electrostatic self-assembly (ESA) technique to promote osteoblast growth. Poly(ethylenimine) (PEI) was first employed to obtain a stable positively charged surface on poly (DL-lactide) (PDL-LA) substrate. Gelatin was selected as ECM-like biomacromolecule to deposit on the activated PDL-LA substrate using the ESA technique. zeta-Potential results showed alternating charge of polyelectrolytes (PEI/gelatin) layering on PDL-LA microspheres. Quartz crystal microbalance (QCM) measurement further verified the gradual deposition of PEI/gelatin on PDL-LA thin film. Osteoblast cells (MC3T3) were chosen to test the cell behavior on modified PDL-LA substrates. The osteoblast test about cell activity, intracellular total DNA content, total protein content and cell morphology by SEM investigation on ECM-like multilayer-modified PDL-LA substrate showed to promote osteoblast growth. Comparing conventional coating methods, polyelectrolyte multilayers are easy and stable to prepare. It may be a good choice for the surface modification of complex biomedical devices. These very flexible systems allow broad medical applications for drug delivery and tissue engineering.

3T3 Cells↗

Dual mode bioreactions on polymer nanoparticles covered with phosphorylcholine group.

We investigated the preparation of polymer nanoparticles covered with phosphorylcholine (PC) groups and the immobilization of proteins in order to observe dual mode bioreactions on the nanoparticles. For the surface modification on the nanoparticles, a water-soluble amphiphilic phospholipid polymer with PC groups as a hydrophilic moiety was synthesized. In this polymer, an active ester group, which can immobilize proteins, was introduced. Using the phospholipid polymer as a solubilizer, poly(L-lactic acid) nanoparticles were prepared from its methylene chloride solution in an aqueous medium by the solvent evaporation method. The diameter of the nanoparticles was ca. 200 nm and the surface was covered with the PC groups and active ester groups. Proteins could immobilize on the nanoparticles under mild conditions by the reaction between the active ester group and amino group in the proteins. Both an antibody and enzyme were immobilized on the nanoparticles and bioreactions such as the antigen/antibody reaction and enzymatic reaction were observed. When an antigen was added to the suspension of the nanoparticles, aggregation of the nanoparticles occurred and then they precipitated. Also, the enzymatic reaction proceeded well when the enzyme substrate was added to the suspension. Based on these results, we provided polymer nanoparticles functionalized with both the antibody and enzyme, and the dual mode bioreactions could occur. We concluded that the novel polymer nanoparticles could be used for nano-/micro-scaled diagnostic and medical treatment systems.

Antibodies, Monoclonal↗

Differential conjugation of tat peptide to superparamagnetic nanoparticles and its effect on cellular uptake.

Surface modification of superparamagnetic contrast agents with HIV-1 tat peptide has emerged as a promising means for intracellular magnetic labeling and noninvasive tracking of a large number of cell types with MRI. To achieve efficient intracellular delivery of the nanoparticles, we investigated the effect on cellular uptake of superparamagnetic iron oxide particles by varying the number of attached tat peptides. First, we report here a modified P2T method in measuring the numbers of surface attachments per particle through disulfide linkage. The method was shown to have desirable simplicity and reproducibility. With the P2T method as a tool, conjugates with progressively higher ratios of peptide-to-particle were synthesized. We were able to demonstrate that higher numbers of tat peptide facilitate the cellular uptake of iron oxide nanoparticles in a nonlinear fashion. Cells labeled with these optimized preparations were readily detectable by MR imaging. The increase in sensitivity could allow in vivo tracking of 100-fold lower cell concentration than currently described.

Animals↗

Acid etching of titanium for bonding with veneering composite resins.

Commercially pure titanium (cpTi) was etched using three concentrated acids: 18% HCl, 43% H3PO4, and 48% H2SO4. The bond strengths between five types of veneering composite resin and eight cpTi treatments (involving combinations of sandblasting, acid etching in 48% H2SO4, and vacuum firing) were determined before and after 10,000 and 20,000 thermal cycles. There were no significant differences in the bond strength of resin to cpTi after etching in 48% H2SO4 at 90 degrees C for 15 minutes, at 60 degrees C for 15, 30, or 60 minutes, and after sandblasting with and without vacuum firing (p > 0.05); moreover, these treatments yielded the highest values. As for vacuum firing, it had no significant effect on resin bond strength to cpTi before or after 10,000 and 20,000 thermal cycles. We therefore concluded that acid etching in concentrated H2SO4 is a simple and effective surface modification method of titanium for bonding to veneering composite resins.

Acid Etching, Dental↗

Platelet adhesion and contact activation time tests on HEMA coated cellulose acetate membranes.

Surface modification of cellulose acetate dialysis membranes was carried out by 60Co radiation induced graft copolymerization of the hydrogel, hydroxyethyl methacrylate (HEMA). The degree of grafting was controlled by varying the HEMA monomer concentration in the grafting solution and the radiation dose. A continuous flow platelet adhesion test was designed which allows testing under conditions more closely approximating hemodialysis than other small scale in vitro tests. Platelet adhesion on treated membranes fell substantially with increasing surface HEMA concentration. The presence of HEMA on the membrane surface did not affect the membrane activated clotting times significantly.

Acrylates↗

Nanofabrication for micropatterned cell arrays by combining electron beam-irradiated polymer grafting and localized laser ablation.

Most methods reported for cell-surface patterning are generally based on photolithography and use of silicon or glass substrates with processing analogous to semiconductor manufacturing. Herein, we report a novel method to prepare patterned plastic surfaces to achieve cell arrays by combining homogeneous polymer grafting by electron beam irradiation and localized laser ablation of the grafted polymer. Poly(N-isopropylacrylamide) (PIPAAm) was covalently grafted to surfaces of tissue culture-grade polystyrene dishes. Subsequent ultraviolet ArF excimer laser exposure to limited square areas (sides of 30 or 50 microm) produced patterned ablative photodecomposition of only the surface region (approximately 100-nm depth). Three-dimensional surface profiles showed that these ablated surfaces were as smooth and flat as the original tissue culture-grade polystyrene surfaces. Time-of-flight secondary ion mass spectrometry analysis revealed that the ablated domains exposed basal polystyrene and were surrounded with PIPAAm-grafted chemistry. Before cell seeding, fibronectin was adsorbed selectively onto ablated domains at 20 degrees C, a condition in which the non-ablated grafted PIPAAm matrix remains highly hydrated. Hepatocytes seeded specifically adhered onto the ablated domains adsorbed with fibronectin. Because PIPAAm, inhibits cell adhesion and migration even at 37 degrees C when the grafted density is > 3 microg/cm2, all the cells were confined within the ablated domains. A 100-cell domain array was achieved by this method. This surface modification technique can be utilized for fabrication of cell-based biosensors as well as tissue-engineered constructs.

Acrylic Resins↗

Heparin surface modified intraocular lenses implanted in the monkey eye.

The biocompatibility of heparin surface modified poly(methyl methacrylate) intraocular lenses (IOLs) was evaluated in two experiments following implantation in the anterior and posterior eye chambers of adult cynomolgus monkeys. Throughout the study, large inflammatory cells and prominent pigment deposits were seen on the unmodified lenses, whereas the heparin surface modified IOLs remained almost free of precipitates. Similarly, fewer posterior synechias were observed in eyes implanted with surface modified IOLs in the posterior chamber than in eyes implanted with control lenses. Histopathological examination of enucleated eyes confirmed the clinical findings. These experiments strongly support the idea that surface modification with heparin is a useful way to reduce clinical complications following cataract surgery with IOL implantation.

Animals↗

The effect of a PEG versus a chitosan coating on the interaction of drug colloidal carriers with the ocular mucosa.

The influence of the surface characteristics of colloidal drug carriers in their interaction with different biological surfaces is becoming increasingly evident. In order to investigate the importance of these characteristics in their interaction with the ocular mucosa, we developed three types of nanocapsules that differ in their surface properties: poly- epsilon -caprolactone (PECL) nanocapsules, chitosan (CS)-coated PECL nanocapsules and poly(ethylene glycol) (PEG)-coated PECL nanocapsules. Two different approaches were used to form these polymer coated nanocapsules: (i) the electrostatic anchorage of the coating onto the PECL nanocapsules-in the case of CS-and (ii) the use of the previously synthesized copolymer PECL-PEG for the formation of the nanocapsules. In both cases, the systems, prepared by the interfacial deposition technique, were loaded with a fluorescent dye (rhodamine) in order to quantify and visualize their interaction with the ocular surface ex vivo and in vivo. An important conclusion from the ex vivo studies is that the developed systems, and specially the CS-coated ones, enhanced the penetration of the encapsulated dye through the cornea. This effect was not simple due to the physical presence of the nanocapsules but to their ability to carry the encapsulated compound. The second conclusion from the confocal laser scanning microscopy (CLSM) studies is that the systems were able to enter the corneal epithelium by a transcellular pathway and that the penetration rate was dependent on the coating composition. The images suggest that the PEG coating accelerates the transport of the nanocapsules across the whole epithelium, whereas the CS coating favours the retention of the nanocapsules in the superficial layers of the epithelium. The specific behaviour of CS-coated systems was also corroborated in vivo. These results indicate that the surface composition of colloidal drug carriers affects their biodistribution in the eye. Therefore, this surface modification approach can be used as a targeting strategy in ocular drug delivery.

Animals↗

Correlation of topography to bond strength of etched ceramic.

PURPOSE: Bonding techniques are commonly used today to ensure the durability of porcelain laminate veneers and ceramic inlay-onlays, which are esthetically pleasing solutions to the problem of dental restoration. Acid etching and silanization of porcelain have been widely used to enhance the retention between bonding resins and ceramic restorations. The purpose of this study was to evaluate the effect of surface modifications with 10% hydrofluoric acid gel and a coupling agent (a dual component silane) on the bond strength of 2 dental feldspathic ceramics (GC and PVS) bonded with an unfilled resin (Super-Bond). MATERIALS AND METHODS: Eighty samples of GC and eighty samples of PVS were built for a mechanical study. Mechanical tests were conducted using a Lloyd T 6000 R tensile machine, which determined the bond strengths of the 2 ceramics after different surface treatments (etching or silanization). In the next part of the study, 10 surfaces of GC and 10 surfaces of PVS were studied with a scanning mechanical microscope to evaluate the action of hydrofluoric acid gel on the roughness of the 2 ceramics. RESULTS: Etching ceramic with hydrofluoric acid gel increased the developed surface of feldspathic ceramics, especially for PVS, but this treatment was not sufficient to obtain the highest shear bond strength. The highly positive influence of silanization was shown, particularly for GC ceramic bonded with an unfilled resin. CONCLUSION: Silane combined with the action of hydrofluoric acid gel is the most effective surface treatment for ceramic.

Analysis of Variance↗

Hemocompatibility of layer-by-layer hyaluronic acid/heparin nanostructure coating on stainless steel for cardiovascular stents and its use for drug delivery.

In order to develop drug-eluting cardiovascular stents, stainless steel (SUS316L) sheets were coated with hyaluronic acid (HA) and heparin (HEP), and their in vitro characteristics and drug release pattern were investigated. The surface of stainless steel (SS) was treated with nitric acid and followed by anchoring aminotrimethoxysilane (ATMS), then a nanolayer of HA was covalently immobilized onto the surface. Heparin was then covalently bonded to the HA-immobilized SS substrate. After repeating 1 to 5 cycles, 1 to 5 layers polyelectrolyte complex (PEC) nanobrush of HA/HEP were resulted with the thickness ranging from 280 to 630 nm (measured with ellipsometry). A model drug (sirolimus) was loaded in the HA/HEP layers at a density ranging from 1.02 to 3.12 microg/cm2. The SS-ATMS-HA-HEP substrates were evidenced by X-ray photoelectron spectroscope (XPS), contact angle, and AFM measurement. The effect of this surface modification on the coagulation time of the resulting SS substrates was investigated. The results show that the multilayer HA/HEP stainless steel would exhibit higher anticoagulant activity than pure SS substrates. In addition, the results of the in vitro drug delivery study showed that release of sirolimus from the 5-layer-HA-HEP stainless steel was able to maintain more than 30 days. Thus layer-by-layer HA/HEP PEC can improve the hemocompatibility of SS surface and control the drug released rate by multiple layers of HA/HEP PEC. These results indicate that the multi-layer HA/HEP PEC coated stainless steel would be suitable for drug eluting stents.

Adjuvants, Immunologic↗

Chitosan surface-modified hydroxycamptothecin loaded nanoparticles with enhanced transport across Caco-2 cell monolayer.

This study explored the feasibility of using surface-modified nanoparticulate drug delivery system to enhance the transepithelial transport of antitumor drugs. An antitumor drug, 10-hydroxycamptothecin, was encapsulated into nanoparticles made of biodegradable poly(caprolactone-co-lactide)-PEG-poly(caprolactone-co-lactide) by a novel two-step nano-precipitation method. The obtained nanoparticles had a drug loading content of 10.4% and a size of 256.3 nm, exhibiting a steady and sustained in vitro release profile. By incubation in chitosan containing medium, the drug-loaded nanoparticles could be subsequently surface-modified with chitosan. The surface modification was monitored by dynamic light scattering method, zeta potential observation, and transmission electron microscopy, and its degree could be easily adjusted by varying the concentration of chitosan in the incubation medium. Caco-2 cell monolayer was used as an in vitro model to evaluate the intestinal 10-hydroxycamptothecin absorption. The absorptive transport of 10-hydroxycamptothecin could be improved to some extent by drug loaded nanoparticles and could be further enhanced in the case of surface-modified nanoparticles, suggesting that chitosan surface-modified nanoparticles may be a promising oral delivery system for antitumor drugs.

Antineoplastic Agents, Phytogenic↗

Carbon materials in the treatment of soft and hard tissue injuries.

Carbon-based implant materials are of interest because they are well accepted by the biological environment. Carbon fibrous materials developed in the Department of Special Ceramics of the University of Mining and Metallurgy in Cracow were tested in in vivo studies to determine their influence on the living body. For comparative purposes, different carbon fibers were prepared and subjected to different surface modifications. Carbon materials prepared in the form of braids were implanted in subcutaneous tissue of rabbits and into skeletal muscle of rats. Carbon fabrics were examined as scaffolds in reconstruction of bone defects. The present study examined the synthesis-structure-property relationships of fibrous carbon samples with respect to the tissue response. It was shown that the tissue response depends on the form of the material form, the degree of order of the crystallites, the surface state and microstructural parameters. Carbon fibers with higher crystallinity and a better-organized graphite structure were assimilated by the body with more difficulty and small particles coming from these materials were found in the regional lymph nodes. Low-carbonized carbon fibers (small crystallite size) underwent partial fragmentation and reacted with the biological environment by being gradually resorbed in the implantation site. The presence of acidic groups on the surface of the carbon fibers enhanced phagocytosis of the carbon material by macrophages. Depending on the surface state of carbon fibers different rates of bone wound healing were observed.

Journal Article↗