Spatio-temporal control over destabilization of Pickering emulsions stabilized by light-sensitive dextran-based nanoparticles.

The implementation of light-sensitive Pickering emulsions with spatio-temporal responsiveness in advanced applications like drug-delivery, colloidal or reaction engineering would open new avenues. However, curiously, light-sensitive Pickering emulsions are barely studied in the literature and their biocompatibility and/or degradability scarcely addressed. Thus, their development remains a major challenge. As an original strategy, we synthesized light-sensitive nanoparticles based on biocompatible Poly(NitroBenzylAcrylate) grafted dextran (Dex-g-PNBA) to stabilize O/W Pickering emulsions. The produced emulsions were stable in time and could undergo time and space-controlled destabilization under light stimulus. Irradiation time and alkaline pH-control of the aqueous phase were proved to be the actual key drivers of destabilization. As the nanoparticles themselves were photolyzed under light stimulus, possible harmful effects linked to accumulation of nanomaterials should be avoided. In addition to UV light (365 nm), visible light (405 nm) was successfully used for the spatio-temporal destabilization of the emulsions, offering perspectives for life science applications.

Magnetically-responsive, multifunctional drug delivery nanoparticles for elastic matrix regenerative repair.

Arresting or regressing growth of abdominal aortic aneurysms (AAAs), localized expansions of the abdominal aorta are contingent on inhibiting chronically overexpressed matrix metalloproteases (MMPs)-2 and -9 that disrupt elastic matrix within the aortic wall, concurrent with providing a stimulus to augmenting inherently poor auto-regeneration of these matrix structures. In a recent study we demonstrated that localized, controlled and sustained delivery of doxycycline (DOX; a tetracycline-based antibiotic) from poly(lactic-co-glycolic acid) nanoparticles (PLGA NPs), enhances elastic matrix deposition and MMP-inhibition at a fraction of the therapeutically effective oral dose. The surface functionalization of these NPs with cationic amphiphiles, which enhances their arterial uptake, was also shown to have pro-matrix regenerative and anti-MMP effects independent of the DOX. Based on the hypothesis that the incorporation of superparamagnetic iron oxide NPs (SPIONs) within these PLGA NPs would enhance their targetability to the AAA site under an applied external magnetic field, we sought to evaluate the functional effects of NPs co-encapsulating DOX and SPIONs (DOX-SPION NPs) on elastic matrix regeneration and MMP synthesis/activity in vitro within aneurysmal smooth muscle cell (EaRASMC) cultures. The DOX-SPION NPs were mobile under an applied external magnetic field, while enhancing elastic matrix deposition 1.5-2-fold and significantly inhibiting MMP-2 synthesis and MMP-2 and -9 activities, compared to NP-untreated control cultures. These results illustrate that the multifunctional benefits of NPs are maintained following SPION co-incorporation. Additionally, preliminary studies carried out demonstrated enhanced targetability of SPION-loaded NPs within proteolytically-disrupted porcine carotid arteries ex vivo, under the influence of an applied external magnetic field. Thus, this dual-agent loaded NP system proffers a potential non-surgical option for treating small growing AAAs, via controlled and sustained drug release from multifunctional, targetable nanocarriers. STATEMENT OF SIGNIFICANCE: Proactive screening of high risk elderly patients now enables early detection of abdominal aortic aneurysms (AAAs). There are no established drug-based therapeutic alternatives to surgery for AAAs, which is unsuitable for many elderly patients, and none which can achieve restore disrupted and lost elastic matrix in the AAA wall, which is essential to achieve growth arrest or regression. We have developed a first generation design of polymer nanoparticles (NPs) for AAA tissue localized delivery of doxycycline, a modified tetracycline drug at low micromolar doses at which it provides both pro-elastogenic and anti-proteolytic benefits that can augment elastic matrix regenerative repair. The nanocarriers themselves are also uniquely chemically functionalized on their surface to also provide them pro-elastin-regenerative & anti-matrix degradative properties. To provide an active driving force for efficient uptake of intra-lumenally infused NPs to the AAA wall, in this work, we have rendered our polymer NPs mobile in an applied magnetic field via co-incorporation of super-paramagnetic iron oxide NPs. We demonstrate that such modifications significantly improve wall uptake of the NPs with no significant changes to their physical properties and regenerative benefits. Such NPs can potentially stimulate structural repair in the AAA wall following one time infusion to delay or prevent AAA growth to rupture. The therapy can provide a non-surgical treatment option for high risk AAA patients.

Investigating the effect of tumor vascularization on magnetic targeting in vivo using retrospective design of experiment.

Nanocarriers take advantages of the enhanced permeability and retention (EPR) to accumulate passively in solid tumors. Magnetic targeting has shown to further enhance tumor accumulation in response to a magnetic field gradient. It is widely known that passive accumulation of nanocarriers varies hugely in tumor tissues of different tumor vascularization. It is hypothesized that magnetic targeting is likely to be influenced by such factors. In this work, magnetic targeting is assessed in a range of subcutaneously implanted murine tumors, namely, colon (CT26), breast (4T1), lung (Lewis lung carcinoma) cancer and melanoma (B16F10). Passively- and magnetically-driven tumor accumulation of the radiolabeled polymeric magnetic nanocapsules are assessed with gamma counting. The influence of tumor vasculature, namely, the tumor microvessel density, permeability and diameter on passive and magnetic tumor targeting is assessed with the aid of the retrospective design of experiment (DoE) approach. It is clear that the three tumor vascular parameters contribute greatly to both passive and magnetically targeted tumor accumulation but play different roles when nanocarriers are targeted to the tumor with different strategies. It is concluded that tumor permeability is a rate-limiting factor in both targeting modes. Diameter and microvessel density influence passive and magnetic tumor targeting, respectively.

Structure and antioxidant activity of soy protein isolate-dextran conjugates obtained by TiO2 photocatalysis.

The aim of this study was to investigate the structural characteristics and antioxidant activities of soy protein isolate- (SPI-) dextran conjugates obtained by TiO2 photocatalysis treatment. Results revealed that the UV-vis absorption and the fluorescence intensity increased as the photocatalytic power increased (P < 0.05). Higher photocatalytic power could promote the extent of glycation and the formation of high molecular weight SPI-dextran conjugates, which were evidenced by free amino group content and sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The Fourier transform infrared (FT-IR) spectra suggested that the amide I, II, and III bands of SPI were altered by the glycation induced by TiO2 photocatalysis. Moreover, significant changes of secondary structure occurred in SPI-dextran conjugates. The α-helix, ß-sheet, ß-turns, and random coil were changed from approximately 10.6%, 37.9%, 12.9%, and 38.6% to 3.8%, 10.4%, 17.7%, and 68.8%, respectively, after treatment at photocatalytic power of 1000 W. In addition, SPI-dextran conjugates obtained by TiO2 photocatalysis treatment exhibited high hydroxyl radical scavenging activity and possessed increased reducing power. All data indicated that TiO2 photocatalysis was an efficient method for promoting protein-polysaccharide copolymerisation.

Radiation cross-linked collagen/dextran dermal scaffolds: effects of dextran on cross-linking and degradation.

Ionizing radiation effectively cross-links collagen into network with enhanced anti-degradability and biocompatibility, while radiation-cross-linked collagen scaffold lacks flexibility, satisfactory surface appearance, and performs poor in cell penetration and ingrowth. To make the radiation-cross-linked collagen scaffold to serve as an ideal artificial dermis, dextran was incorporated into collagen. Scaffolds with the collagen/dextran (Col/Dex) ratios of 10/0, 7/3, and 5/5 were fabricated via (60)Co γ-irradiation cross-linking, followed by lyophilization. The morphology, microstructure, physicochemical, and biological properties were investigated. Compared with pure collagen, scaffolds with dextran demonstrated more porous appearance, enhanced hydrophilicity while the cross-linking density was lower with the consequence of larger pore size, higher water uptake, as well as reduced stiffness. Accelerated degradation was observed when dextran was incorporated in both the in vitro and in vivo assays, which led to earlier integration with cell and host tissue. The effect of dextran on degradation was ascribed to the decreased cross-linking density, looser microstructure, more porous and hydrophilic surface. Considering the better appearance, softness, moderate degradation rate due to controllable cross-linking degree and good biocompatibility as well, radiation-cross-linked collagen/dextran scaffolds are expected to serve as promising artificial dermal substitutes.

Dual stimuli-responsive multi-drug delivery system for the individually controlled release of anti-cancer drugs.

A dual stimuli-responsive multi-drug delivery system was developed for "cancer cocktail therapy". Upon UV irradiation, microcapsules could rapidly release the small-molecule drugs, and thereafter the macromolecular drugs would be released in the presence of MMP in the tumor cells. This system will find great potential as a novel chemotherapeutic combination for cancer treatment.

Electron-beam generated porous dextran gels: experimental and quantum chemical studies.

PURPOSE: The aim of this work was to investigate the reaction mechanism of electron-beam generated macroporous dextran cryogels by quantum chemical calculation and electron paramagnetic resonance measurements. METHODS: Electron-beam radiation was used to initiate the cross-linking reaction of methacrylated dextran in semifrozen aqueous solutions. The pore morphology of the resulting cryogels was visualized by scanning electron microscopy. Quantum chemical calculations and electron paramagnetic resonance studies provided information on the most probable reaction pathway and the chain growth radicals. RESULTS: The most probable reaction pathway was a ring opening reaction and the addition of a C-atom to the double-bond of the methacrylated dextran molecule. CONCLUSIONS: First detailed quantum chemical calculation on the reaction mechanism of electron-beam initiated cross-linking reaction of methacrylated dextran are presented.

Biocompatible polysaccharide-based cryogels.

This study focuses on the development of novel biocompatible macroporous cryogels by electron-beam assisted free-radical crosslinking reaction of polymerizable dextran and hyaluronan derivatives. As a main advantage this straightforward approach provides highly pure materials of high porosity without using additional crosslinkers or initiators. The cryogels were characterized with regard to their morphology and their basic properties including thermal and mechanical characteristics, and swellability. It was found that the applied irradiation dose and the chemical composition strongly influence the material properties of the resulting cryogels. Preliminary cytotoxicity tests illustrate the excellent in vitro-cytocompatibility of the fabricated cryogels making them especially attractive as matrices in tissue regeneration procedures.

Photocrosslinkable bioadhesive based on dextran and PEG derivatives.

In this study, a hybrid photopolymeric bioadhesive system consisting of urethane methacrylated dextran (Dex-U) and 3, 4-Dihydroxyphenyl-l-alanine (DOPA) modified three-arm poly (ethylene glycol) s (PEG-DOPAs) was designed. The process of photopolymerization was detected by Photo-Differential Scanning Calorimetry (Photo-DSC). The adhesion strength was evaluated by the lap shear tests. The surface tension of the solutions, burst pressures and the cytotoxicity assays were also investigated. The addition of PEG-DOPAs significantly improved the properties of Dex-U especially in the field of adhesion strength and burst pressure. And materials variation could be tailored to match the demands for tissue repair. Compared to the Dex-U systems, the maximum adhesion strength of the copolymeric system increased from 2.7±0.1 MPa to 4.0±0.6 MPa. Owing to its strong adhesion strength, rapid curing rate and good biocompatibility, such photocrosslinkable hydrogelsa could be applied to the areas of bioadhesive.

A novel approach for the synthesis of superparamagnetic Mn3O4 nanocrystals by ultrasonic bath.

In this study, the synthesis of Mn(3)O(4) (husmannite) nanoparticles was carried out in two different alkali media under sonication by ultrasonic bath and conventional method. Manganese acetate was used as precursor, sodium hydroxide and hexamethylenetetramine (HMT) as basic reagents in this synthesis. An ultrasonic bath with low intensity was used for the preparation of nanomaterials. The as prepared samples were characterized with X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, transmission electron microscopy (HRTEM, TEM), energy-dispersive spectrum (EDS), and superconducting quantum interference device (SQUID) analysis. The XRD patterns exhibit the nanocrystals are in pure tetragonal phase. The chemical composition was obtained by EDS analysis and confirmed the presence of Mn and O in the sample. According to the TEM and HRTEM results, both nanorods and nanoparticles of Mn(3)O(4) were obtained in the presence of ultrasonic irradiation. The average size of nanoparticles was 10nm, and the size of nanorods was 12 nm in diameter and 100-900 nm in length for the samples prepared in basic medium with sodium hydroxide. In the conventional method with the same basic medium, the nanorod was not observed and the nearly cubic nanoparticles was appeared with an average size of 2.5 nm. The selected area electron diffraction (SAED) patterns revealed that the nanocrystals are polycrystalline in nature. When HMT was used as a basic reagent in the presence of ultrasonic irradiation, it was led to a higher size of nanoparticles and nanorods than when sodium hydroxide was used as a basic reagent. The average size of nanoparticles was about 15 nm and its shape was nearly cubic. The diameter for nanorods was 50 nm and the length was about a few micrometers. The magnetic measurements were carried out on the sample prepared in sodium hydroxide under ultrasonic irradiation. These measurements as a function of temperature and field strength showed a reduction in ferrimagnetic temperature (T(c) = 40K) as compared to those reported for the bulk (T(c) = 43K). The superparamagnetic behavior was observed at room temperature with no saturation magnetization and hysteresis in the region of measured field strength.

Evaluation of uptake and transport of ultrasmall superparamagnetic iron oxide nanoparticles by human brain-derived endothelial cells.

AIM: Ultrasmall superparamagnetic iron oxide nanoparticles (USPIO-NPs) are under development for imaging and drug delivery; however, their interaction with human blood-brain barrier models is not known. MATERIALS & METHODS: The uptake, reactive oxygen species production and transport of USPIO-NPs across human brain-derived endothelial cells as models of the blood-brain tumor barrier were evaluated for either uncoated, oleic acid-coated or polyvinylamine-coated USPIO-NPs. RESULTS: Reactive oxygen species production was observed for oleic acid-coated and polyvinylamine-coated USPIO-NPs. The uptake and intracellular localization of the iron oxide core of the USPIO-NPs was confirmed by transmission electron microscopy. However, while the uptake of these USPIO-NPs by cells was observed, they were neither released by nor transported across these cells even in the presence of an external dynamic magnetic field. CONCLUSION: USPIO-NP-loaded filopodia were observed to invade the polyester membrane, suggesting that they can be transported by migrating angiogenic brain-derived endothelial cells.

Photoregulation of drug release in azo-dextran nanogels.

A simple photoresponsive azo-dextran polymer has been investigated for its ability to act as a nanogel drug carrier. Self aggregation of the azo-dextran polymer leads to the formation of nanogels, AD (5 and 10) in aqueous media, which were characterized by TEM and DLS. When examined under UV light (365 nm), the unloaded nanogels, which were observed to be in the range of 120-290 nm, show dependence on the degree of crosslinking, pH and ionic concentration of the dispersed media. Nanogels, AD (5 and 10), have been loaded with a model fluorophore, rhodamine B and a drug, aspirin, by freeze drying an aqueous dispersion of the nanogels in the presence of the substrate dissolved in water or PBS buffer. The release pattern of the encapsulated bio-active molecules from these nanogels was regulated by (trans-cis) photoisomerization of the azobenzene moiety present in the crosslinker. A comparison of the release behavior of the loaded (rhodamine, aspirin) AD (5 and 10) nanogels reveal that the rate of release of the encapsulated active molecules from the nanogels was slower when the azo moiety was in E-configuration as compared to that the azo in the Z-configuration. The in vitro release behavior of drug from these polymeric micellar systems is revelative of the potential of the nanogels for targeted drug delivery in nanomedicine.

Photocrosslinking of dextran and polyaspartamide derivatives: a combination suitable for colon-specific drug delivery.

The aim of this study was to prepare and characterize novel hydrogels with polysaccharide-polyaminoacid structure, able to undergo an enzymatic hydrolysis in the colon and potentially useful for treating inflammatory bowel diseases (IBD). Starting materials were methacrylated dextran (DEX-MA) and methacrylated alpha,beta-poly(N-2-hydroxyethyl)-dl-aspartamide (PHM). These polymers were photocrosslinked by exposure of their aqueous solutions at 313 nm without photoinitiators. Different samples, shaped as microparticles, were obtained as a function of polymer concentration and irradiation time. FT-IR analysis confirmed the occurrence of a co-crosslinking between DEX-MA and PHM in all experimental conditions. Size analysis evidenced a narrow particle distribution and swelling studies, performed in twice-distilled water and simulated gastrointestinal fluids, showed a good affinity of these hydrogels towards the aqueous medium. DEX-MA/PHM based hydrogels undergo a negligible chemical hydrolysis, whereas they are partially degraded by dextranase. In vitro biological assays showed cell compatibility of these samples. Beclomethasone dipropionate (BDP), a drug recently proposed for the treatment of IBD was entrapped into a DEX-MA/PHM based hydrogel and its release was evaluated in the absence or in the presence of dextranase. Obtained release profiles suggest the potential use of BDP loaded DEX-MA/PHM based hydrogels for the treatment of IBD.

Hydrogels of dextran containing nonsteroidal anti-inflammatory drugs as pendant agents.

The oral administration of nonsteroidal anti-inflammatory drugs (NSAIDs) is often associated with upper gastrointestinal tract side effects. To reduce these effects and improve the therapeutic efficacy, NSAIDs are often formulated as controlled release systems. We have prepared a new formulation consisting of dextran hydrogels containing NSAIDs as pendant agents, through ultraviolet irradiation of solutions of dextran functionalized with methacrylic groups in the presence of the drug derivatized in the same way. Release studies of different drugs from this system, carried out in media simulating the gastrointestinal tract, have demonstrated that the amount of released drug is strictly related to the concentration of the polymer in the solution submitted to irradiation as well as to its derivatization degree. Our obtained data confirm that the system is able to realize a colon-specific drug delivery.

Use of microphotohemolysis to distinguish differences in erythrocyte treatments.

A new microhemolytic assay was used to determine if this assay could distinguish between normal erythrocytes and those which had been experimentally altered. Solutions of erythrocytes (4% hematocrit in buffer with fluorescein isothiocyanate tagged to 150,000 MW dextran, FITC-DEX) were placed in a hemacytometer and epi-illumination with a Leitz fluorescent microscope was used to activate the fluorochrome. The resultant hemolysis that occurred only in the discrete area of activation was dependent on the total light energy used for activation (45-180 J/cm2). It was quantitated by an analysis of the amount of light transmittance through that area. The presence of glucose in the buffers decreased the rate of hemolysis and increased the time to reach 50% of the maximal response (T50). Erythrocytes treated with diamide had up to a fourfold increase in the rate of hemolysis and a 48% decrease in the T50, while chlorpromazine produced a 51% decrease in the T50 but had no effect on the rate of hemolysis. Gluteraldehyde produced a graded suppression of the hemolysis. These results demonstrate that the microphotohemolytic assay can be used with energy response curves to provide a relatively quick, quantitative determination of altered erythrocytes.

Quantitation of erythrocyte photohemolysis by light microscopy.

In a solution of erythrocytes and a photo-active compound, light activation of the compound produces hemolysis of the cells. This study describes a new assay in which focused light through a microscope is used to induce a circumscribed hemolysis of erythrocytes that have been mixed with fluorescein isothiocyanate-dextran 150,000 (FITC-DEX) and placed in a hemacytometer. The hemolytic response was monitored by detecting transmitted light intensity in the area (1.8 x 10(-4) cm2) of activation. Only cells within the specific microscopic field of activation hemolyze. This allows for multiple sites of activation within one sample. The hemolytic response was dependent on the concentration of FITC-DEX (0.5-4 mg/ml) and on light intensity (53-210 J/cm2) but not on small changes in hematocrit (3%-5%) or on the presence of platelets and leukocytes. Rabbit erythrocytes, however, were almost twice as sensitive as those from guinea pigs. Since the photohemolytic response will depend on the composition and strength of the erythrocyte membrane and presence of oxidant defense mechanisms, we suggest that this assay could be used to detect drug- or disease-induced changes in the red blood cell membrane.

Antithrombotic effects of recombinant hirudin in different animal models.

The effect of recombinant hirudin (r-hirudin; HBW 023) was studied in four different models of thrombosis as well as on bleeding time. Arteriolar thrombus formation was induced in rats either by argon laser injury or by photochemical reaction. r-Hirudin, given by intravenous infusion, significantly inhibited arteriolar thrombus formation and arterial bleeding time at a dose of 40 micrograms/kg.min. In rabbit jugular veins and carotid arteries, occluding thrombi were produced by stenosis and endothelial damage. r-Hirudin reduced the incidence of thrombosis dose dependently (ED50 0.7 and 1.0 mg/kg i.v. in arteries and veins, respectively). Caval vein thrombosis was initiated in rats by insertion of a stainless steel coil. This thrombosis was dose dependently reduced by the injection of r-hirudin (ED50 0.16 mg/kg i.v.). The duration of the antithrombotic effect of 0.25 mg/kg of r-hirudin in caval veins was about 60 min, whereas the arterial bleeding time was significantly prolonged in these rats for only 20 min. The plasma level of r-hirudin had dropped to 0.12 micrograms/ml 60 min after injection. These results suggest that the duration of the effect on bleeding time may be shorter than the antithrombotic action of r-hirudin.