In vivo toxicity study of quatro stimuli nanocontainers in pregnant rats: Gestation, parturition and offspring evaluation.

The aim of the present study was to investigate the impact of intravenous administration of newly fabricated nanocontainers (NCs) on the last third of pregnancy in rats. Fifteen pregnant 3-month-old Wistar rats were separated into 3 groups. On the 15th and 17th day of pregnancy all animals received an intravenous administration of 1 ml of 15 mg of NCs (Group A), 1 ml of 5 mg NCs (Group B) while Control group received 1 ml of 0.9% NaCl. On the 14th and 17th of pregnancy ultrasonography was performed and the parameters evaluated were the width of placenta, the length and width of the embryonic sac, the foetus length and the heart rate. On parturition the number of pups per dam was evaluated. Half of the pups were euthanised the day after parturition and their liver and kidney was histologically evaluated and for the rest of the pups the body growth curve was evaluated until the age of 14 week. At the end of the 14th week the remaining pups were euthanised and their liver and kidney was histologically evaluated. At weaning the dams were euthanised and their liver and kidney was histologically evaluated. Ultrasonography: Baseline measurements of the width of placenta, the length and width of embryonic sac, the foetus length and the heart rate on the 14th day of pregnancy, revealed no statistical significant differences between groups. Comparison of the same values on the 17th day of pregnancy after 2 intravenous administrations of NCs showed no statistical significant effect on the respective parameters. The administration of NCs had no impact on the mean number of pups per dam. Additionally, no impact of the NCs on the body weights of the pups was observed on the 1st day after parturition. Moreover, comparisons between groups, for both sexes showed no difference on growth rate. During the histological evaluation no inflammatory, degenerative or neoplastic lesions were observed as far as the newborn, adult offspring and dams were concerned. According to our results no toxic impact of the low and high doses of the NCs was observed on the parameters selected to be evaluated.

Synthesis of novel quaternary silica hybrid bioactive microspheres.

PURPOSE: To survey the preparation of novel hybrid microspheres of quaternary silicate glassy composition (SiO2 P2 O5 CaONa2 O) and the prospect of using them as an osteogenic system with enhanced bioactive properties for the development of hydroxyapatite. METHOD: In line with our previous synthetic procedure a two-step process was followed, wherein polystyrene (PS) microspheres were prepared by the emulsifier free-emulsion polymerization method and constituted the core for the sol-gel coating of the silicate inorganic shell. The development of the hybrid microspheres was based on silane and phosphate precursors and was assesses at different ratio of ethanol/water (of 9/1, 4/1, and 2/1, in mL) and at varied ammonia concentration of 4.8-1.0 mL. RESULTS: The hybrid microspheres had an average size ranged between 350 and 550 nm according to SEM, depending on the ethanol/water solution rate and ammonia content. The final microspheres probably exhibited a porous-like structure through the formation of diffused voids along with the low carbon content of the EDX analysis, which could be regulated by the catalyst content. The hybrid microspheres exhibited effective in vitro bioactivity assessed in simulated body fluids (SBF). CONCLUSION: Quaternary hybrid silica microspheres were effectively synthesized. The bioassay evaluation of the final microspheres revealed the rapid in vitro formation of a bone-like apatite layer. The results verify the bioactivity of the microspheres and promote further research of their suitability on regenerative treatment of bone abnormalities. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 112-120, 2018.

Reducing the layer number of AB stacked multilayer graphene grown on nickel by annealing at low temperature.

Controlling the number of layers of graphene grown by chemical vapor deposition is crucial for large scale graphene application. We propose here an etching process of graphene which can be applied immediately after growth to control the number of layers. We use nickel (Ni) foil at high temperature (T = 900 °C) to produce multilayer-AB-stacked-graphene (MLG). The etching process is based on annealing the samples in a hydrogen/argon atmosphere at a relatively low temperature (T = 450 °C) inside the growth chamber. The extent of etching is mainly controlled by the annealing process duration. Using Raman spectroscopy we demonstrate that the number of layers was reduced, changing from MLG to few-layer-AB-stacked-graphene and in some cases to randomly oriented few layer graphene near the substrate. Furthermore, our method offers the significant advantage that it does not introduce defects in the samples, maintaining their original high quality. This fact and the low temperature our method uses make it a good candidate for controlling the layer number of already grown graphene in processes with a low thermal budget.

A new approach to fabricate bioactive silica binary and ternary hybrid microspheres.

Bioactive microspheres represent an extremely developing field in biomedical applications, such as bone tissue engineering and bone pathologies (metabolic bone disease, trauma or bone cancer). Their innate osteogenic properties have turned them to biomaterials with improved added value. The aim of this study was to prepare binary and ternary hybrid silica microspheres with enhanced bioactive properties according to our previous synthetic procedure. In brief, the synthetic approach based on the emulsifier free-emulsion polymerization method, by which polystyrene (PS) microspheres were produced and used as core template for the sol-gel coating method. During the coating reaction an inorganic shell was fabricated by silane and phosphate precursors (tetraethoxysilane, trimethylphosphate). The final microspheres were treated by different catalyst concentrations, during the coating process, which resulted in the formation of diffused voids (a porous-like structure). The in vitro bioactivity of the resultant microspheres was studied by treatment in simulated body fluids (SBF). The bioassay evaluation indicates the deposition of a bone-like apatite layer on microspheres' surface with enhanced bioresorbability, which verifies their bioactivity and permits their application in the treatment of bone pathologies.

A new approach for the one-step synthesis of bioactive PS vs. PMMA silica hybrid microspheres as potential drug delivery systems.

In this work, hybrid microspheres were prepared in a two-step process combining the emulsifier free-emulsion polymerization and the sol-gel coating method. In the first step, polystyrene (St) and poly(methyl methacrylate) (PMMA) microspheres were prepared as sacrificial template and in the second step a silanol shell was fabricated. The functionalized surface of the hybrid microspheres by silane analogs (APTES, TEOS) resulted in enhanced effects. The hollow microspheres were resulted either in an additional step by template dissolution and/or during the coating process. The microspheres' surface interactions and the size distribution were optimized by treatment in simulated body fluids, which resulted in the in vitro prediction of bioactivity. The bioassay test indicated that the induced hydroxyapatite resembled in structure to naturally occurring bone apatite. The drug doxorubicin (DOX) was used as a model entity for the evaluation of drug loading and release. The drug release study was performed in two different pH conditions, at acidic (pH=4.5) close to cancer cell environment and at slightly basic pH (pH=7.4) resembling the orthopedic environment. The results of the present study indicated promising hybrid microspheres for the potential application as drug delivery vehicles, for dual orthopedic functionalities in bone defects, bone inflammation, bone cancer and bone repair.

Nanostructuring the surface of dual responsive hollow polymer microspheres for versatile utilization in nanomedicine-related applications.

The design and fabrication of hollow polymer microspheres responsive to various stimuli comprises a promising approach for the development of multifunctional and efficient systems for various nanomedicine-related applications. In this paper, we present the preparation of poly(methacrylic acid-co-N,N'-methylenebis(acrylamide)-co-poly(ethylene glycol) methyl ether methacrylate-co-N,N'-bis(acryloyl)cystamine) (PMAA(S-S)) hollow microspheres following a two-stage distillation precipitation polymerization procedure. Magnetic and silver nanocrystals were chemically grown on the surface of the hollow polymer microspheres, resulting in a composite system with interesting properties. We evaluated the performance of the composite hollow microspheres as magnetic hyperthermia mediators and their surface-enhanced Raman spectroscopy activity. Assessment of Daunorubicin-loaded PMAA(S-S) hollow microspheres performance as effective drug carriers was carried out through drug release experiments upon application of different pH and reducing conditions. pH and redox responsiveness as well as basic mechanisms of release profiles are discussed. Furthermore, in vitro cytotoxicity of empty and drug-loaded PMAA(S-S) hollow microspheres against MCF-7 cancer cells was investigated in order to evaluate their performance as drug carriers.

Comparative study of LbL and crosslinked pH sensitive PEGylated LbL microspheres: synthesis, characterization and biological evaluation.

In this report, the fabrication of hollow, pH-sensitive microspheres using the layer by layer method is being demonstrated. The process is based on the coating of colloidal silica templates with polyelectrolyte layers, followed by dissolution of the core using a buffer system of hydrofluoric acid and ammonium fluoride. With this buffer system, the template can be dissolved in mild pH conditions, where the polymeric layers are still stable. The resulting microspheres show pH dependent properties, where they swell and dissolve at a certain pH value. In order to improve the nanocontainers properties, such as solubility and cytotoxicity a second specimen of microspheres was fabricated by coating the fabricated microspheres with one layer of poly(acrylic acid) and one layer of methoxy-polyethylene glycol amine, and then these additional layers were crosslinked by carbodiimide chemistry. Loading and release properties of the fabricated microspheres have been studied in order to investigate their behavior as a function of pH and ionic strength. These novel microspheres have potential application in the fields of drug delivery, diagnostics and life sciences.

Microspheres as therapeutic delivery agents: synthesis and biological evaluation of pH responsiveness.

A soft template method was used for the synthesis of pH-responsive microcontainers with an inner cavity. Poly(glycidyl methacrylate) (PGMA) microspheres of narrow size distribution were synthesized by soap-free radical emulsion polymerization and the coating of the microspheres was carried out by the same procedure. The procedure consists of two steps. In the first step the sacrificial template is synthesized and in the second step the shell is formed. Acrylic acid was used as a coating monomer, with the aim of introducing pH sensitivity in the synthesized microcontainers. A loading and release study of the anthracycline drug doxorubicin (DOX) was also carried out. The toxicity evaluation of the drug was carried out using the MTT assay, and the necrotic effect was studied using trypan blue.

Novel PLA modification of organic microcontainers based on ring opening polymerization: synthesis, characterization, biocompatibility and drug loading/release properties.

In the current study, poly lactic acid (PLA) modified hollow crosslinked poly(hydroxyethyl methacrylate) (PHEMA) microspheres have been prepared, in order to obtain a stimulus-responsive, biocompatible carrier with sustained drug release properties. The synthetical process consisted of the preparation of poly(methacrylic acid)@poly(hydroxyethyl methacrylate-co-N,N'-methylene bis(acrylamide)) microspheres by a two stage distillation-precipitation polymerization technique using 2,2'-azobisisobutyronitrile as initiator. Following core removal, a PLA coating of the microspheres was formed, after ring opening polymerization of DL-lactide, attributing the initiator's role to the active hydroxyl groups of PHEMA. The anticancer drug daunorubicin (DNR) was selected for the study of loading and release behavior of the coated microspheres. The loading capacity of the PLA modified microspheres was found to be four times higher than that of the parent ones (16% compared to 4%). This coated microspherical carrier exhibited a moderate pH responsive drug release behavior due to the pH dependent water uptake of PHEMA, and PLA hydrolysis. The in vitro cytotoxicity of both the parent and the DNR-loaded or empty modified hollow microspheres has been also examined on MCF-7 breast cancer cells. The results showed that although the empty microspheres were moderately cytotoxic, the DNR-loaded microspheres had more potent anti-tumor effect than the free drug. Therefore, the prepared coated microspheres are interesting drug delivery systems.

Sacrificial template-directed fabrication of superparamagnetic polymer microcontainers for pH-activated controlled release of Daunorubicin.

Magnetic pH-sensitive microcontainers were produced by a four-step process. The first step involves the synthesis of citrate-modified magnetic nanoparticles via the coprecipitation method. The second step consists of the encapsulation of magnetic nanoparticles in non-cross-linked poly(methacrylic acid) (PMAA) microspheres through distillation precipitation polymerization, resulting in a core/shell structure. The third step concerns the formation of a poly(N,N'-methylenebis(acrylamide)-co-mathacrylic acid) (P(MBAAm-co-MAA)) layer on the surface of magnetic PMAA microspheres by second distillation precipitation polymerization in order to produce a trilayer hybrid microsphere. The last step deals with the removal of PMAA layer in ethanol and formation of a stable P(MBAAm-co-MAA) microcontainer with magnetic nanoparticles entrapped inside the formed cavity. This process is simple and leads to the formation of superparamagnetic pH-sensitive microcontainers. The structure and properties of the magnetic microcontainers were investigated by X-ray diffraction (XRD), transmission electron microscopy (TEM), scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FT-IR), vibrating sample magnetometry (VSM), and dynamic light scattering (DLS) to determine the functionalities of the hybrid structure. The magnetic pH-sensitive microcontainers were loaded with Daunorubicin and tested with respect to release rate at different pH values in order to evaluate their functionality as controlled release system.

Epidemiological and immunological profile of muscle-specific kinase myasthenia gravis in Greece.

OBJECTIVES: The purposes of this study were to determine the epidemiological characteristics of muscle-specific kinase-myasthenia gravis (MuSK-MG) in Greece and the IgG subclass of the anti-MuSK antibodies. METHODS: This population-based study was performed on MuSK-MG patients in Greece between 1 January 1986 and 30 June 2006. Epidemiological and clinical data for 33 patients were collected. In addition, the distribution of anti-MuSK IgG autoantibody subclasses in the sera of 14 patients was determined by immunoprecipitation. RESULTS: The average annual incidence was 0.32 patients/million population/year. On 1st July 2006, there were 33 prevalent cases, giving a point prevalence rate of 2.92/million (women 4.56 and men 1.25). In females, onset of MuSK-MG occurred after the age of 30, whilst, in males, the disease appears in any decade. The female:male incidence ratio was 3.33:1, whilst the prevalence ratio was 3.65:1. Most patients presented with involvement of the facial and bulbar muscles. Amongst about 800 MG patients seropositive for antibodies against either the AChR or MuSK, one patient was found to be seropositive for anti-MuSK antibodies and ambiguous for anti-acetylcholine receptor (anti-AChR) antibodies. The vast majority of anti-MuSK antibodies were IgG4, whilst total IgG4 levels in these patients were similar to those in two healthy controls. CONCLUSIONS: The incidence and prevalence of MuSK-MG in Greece are amongst the highest reported previously for other countries. MuSK-MG in Greece affects both sexes, but mainly females. The main epidemiological indices were calculated. The vast majority of anti-MuSK antibodies were IgG4.

Assessment of liver iron overload by T2-quantitative magnetic resonance imaging: correlation of T2-QMRI measurements with serum ferritin concentration and histologic grading of siderosis.

PURPOSE: To correlate hepatic 1/T2 values obtained by means of a T2-Quantitative MRI (T2-QMRI) technique with three widely applied methods for the evaluation of hemosiderosis, i.e., (a) liver iron concentrations (LFeC) (b) serum ferritin (SF), and (c) histologic grading of siderosis. The impact of coexisting hepatitis was also considered. T2-QMRI measurements were compared with signal intensity (SI) ratio measurements on conventional SE images. MATERIALS AND METHODS: Liver T2 relaxation times were calculated in 40 thalassemic patients, on a 0.5 T magnetic resonance imaging system using a multiple spin-echo sequence with parameters: TR = 2500 ms, TE = 12 ms in 20 symmetrically repeatable echoes. RESULTS: (a) 1/T2 values were well correlated (r = 0.97) with liver iron concentrations, which ranged from 2.32 to 18.0 mg/g dry weight (normal < 1.6 mg/g). (b) 1/T2 values were also correlated with serum ferritin levels (r = 0.84). At various 1/T2 values, serum ferritin levels were higher for the anti-HCV(+) patients than the anti-HCV(-) ones. (c) T2 values corresponding to successive grades of siderosis presented statistically significant differences. (d) SI ratio measurement assigned less statistically significant results, as compared to T2 values. CONCLUSION: T2-QMRI measurement of T2 relaxation time is more accurate than SI ratios in evaluating liver iron overload. It is particularly useful for hemosiderotic patients with coexisting hepatitis since, in this case, serum ferritin is not considered a reliable index of hemosiderosis.