Structure assembly regularities in vapour-deposited gold-fullerene mixture films.

Self-assembly is an attractive phenomenon that, with proper handling, can enable the production of sophisticated hybrid nanostructures with sub-nm-scale precision. The importance of this phenomenon is particularly notable in the fabrication of metal-organic nanomaterials as promising substances for spintronic devices. The exploitation of self-assembly in nanofabrication requires a comprehension of atomic processes creating hybrid nanostructures. Here, we focus on the self-assembly processes in the vapour-deposited Au x C60 mixture films, revealing the exciting quantum plasmon effects. Through a systematic characterization of the Au x C60 films carried out using structure-sensitive techniques, we have established correlations between the film nanostructure and the Au concentration, x. The analysis of these correlations designates the Au intercalation into the C60 lattice and the Au clustering as the basic processes of the nanostructure self-assembly in the mixture films, the efficiency of which strongly depends on x. The evaluation of this dependence for the Au x C60 composite nanostructures formed in a certain composition interval allows us to control the size of the Au clusters and the intercluster spacing by adjusting the Au concentration only. This study represents the self-assembled Au x C60 mixtures as quantum materials with electronic functions tuneable by the Au concentration in the depositing mixture.

High resolution imaging of 2D distribution of lithium in thin samples measured with multipixel detectors in sandwich geometry.

A method that enables visualization of lateral distribution of Li in thin films is described. The method is based on the simultaneous detection of the reaction products of the 6Li(n,α)t nuclear reaction with thermal neutrons measured with two multipixel detectors in a sandwich geometry with a sample. Here, the principle and basic methodological parameters of the method, including tests with thin polymers with known Li microstructure, are discussed.

Position sensitive detection of neutrons in high radiation background field.

We present the development of a high-resolution position sensitive device for detection of slow neutrons in the environment of extremely high γ and e(-) radiation background. We make use of a planar silicon pixelated (pixel size: 55 × 55 µm(2)) spectroscopic Timepix detector adapted for neutron detection utilizing very thin (10)B converter placed onto detector surface. We demonstrate that electromagnetic radiation background can be discriminated from the neutron signal utilizing the fact that each particle type produces characteristic ionization tracks in the pixelated detector. Particular tracks can be distinguished by their 2D shape (in the detector plane) and spectroscopic response using single event analysis. A Cd sheet served as thermal neutron stopper as well as intensive source of gamma rays and energetic electrons. Highly efficient discrimination was successful even at very low neutron to electromagnetic background ratio about 10(-4).

Deconvolution of charged particle spectra from neutron depth profiling using Simplex method.

Neutron depth profiling (NDP), based on neutron induced nuclear reactions, is a well known, nondestructive technique for the determination of the concentration depth profiles of some isotopes in the surface layers of solids. The profile determination consists of deconvolution of a relevant part of the energy spectra of the charged reaction products. Several solutions have been suggested for this problem. In this work, an alternative computer code (LIBOR), which makes use of the Simplex minimization technique for the deconvolution of the NDP spectra, is described and its performance is documented on several examples.

Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part 4: growth of rat bone marrow stromal cells in three-dimensional hydrogels with positive and negative surface charges and in polyelectrolyte complexes.

The growth of bone marrow stromal cells was assessed in vitro in macroporous hydrogels based on 2-hydro- xyethyl methacrylate (HEMA) copolymers with different electric charges. Copolymers of HEMA with sodium methacrylate (MA(-)) carried a negative electric charge, copolymers of HEMA with [2-(methacryloyloxy)ethyl] trimethylammonium chloride (MOETA(-)) carried a positive electric charge and terpolymers of HEMA, MA(-) and MOETA(+) carried both, positive and negative electric charges. The charges in the polyelectrolyte complexes were shielded by counter-ions. The hydrogels had similar porosities, based on a comparison of their diffusion parameters for small cations as measured by the real-time tetramethylammonium iontophoretic method of diffusion analysis. The cell growth was studied in the peripheral and central regions of the hydrogels at 2 hours and 2, 7, 14 and 28 days after cell seeding. Image analysis revealed the highest cellular density in the HEMA-MOETA(+) copolymers; most of the cells were present in the peripheral region of the hydrogels. A lower density of cells but no difference between the peripheral and central regions was observed in the HEMA-MA(-) copolymers and in polyelectrolyte complexes. This study showed that positively charged functional groups promote the adhesion of cells.

Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part II. Copolymers with positive and negative charges, polyelectrolyte complexes.

Crosslinked macroporous hydrogels based on 2-hydroxyethyl methacrylate (HEMA)-[2-(methacryloyloxy)ethyl]trimethylammonium chloride (MOETACl) copolymer, HEMA-MOETACl-methacrylic acid (MA) terpolymer, and on a polyelectrolyte complex of HEMA-MA copolymer with poly(MOETACl) were prepared. All the hydrogels were prepared in the presence of fractionated sodium chloride particles. The hydrogels were characterized by the number of pores and the total volume of all pores in unit volume, the average volume and the average diameter of single pore. Morphology of the hydrogels was investigated by confocal and scanning electron microscopy. The hydrogels based on polyelectrolyte complexes were also characterized by chemical composition. Homogeneous (non-porous) hydrogels with the same composition as macroporous hydrogels were prepared and characterized by their biocompatibility.

Detection of cell type and marker specificity of nuclear binding sites for anionic carbohydrate ligands.

The emerging functionality of glycosaminoglycan chains engenders interest in localizing specific binding sites using cytochemical tools. We investigated nuclear binding of labeled heparin, heparan sulfate, a sulfated fucan, chondroitin sulfate, and hyaluronic acid in epidermal keratinocytes, bone marrow stromal cells, 3T3 fibroblasts and glioma cells using chemically prepared biotinylated probes. Binding of the markers was cell-type specific and influenced by extraction of histones, but was not markedly affected by degree of proliferation, differentiation or malignancy. Cell uptake of labeled heparin and other selected probes and their transport into the nucleus also was monitored. Differences between keratinocytes and bone marrow stromal cells were found. Preincubation of permeabilized bone marrow stromal cells with label-free heparin reduced the binding of carrier-immobilized hydrocortisone to its nuclear receptors. Thus, these tools enabled binding sites for glycosaminoglycans to be monitored in routine assays.

Development of hydrogel implants for urinary incontinence treatment.

Hydrogel implants for urinary incontinence treatment based on HEMA supplemented with 10% methacrylic acid have been developed. The swelling properties of implants were tested in vitro and in vivo after implantation to laboratory mice. Biocompatibility has been determined by incubation of implants in tissue culture, by histological examination of adjacent tissues after subcutaneous application of implants to laboratory mouse in a long-term experiment, and by flow cytometry examination of blood cells. The swelling of hydrogel implants was completed in 6-24 h. There was no effect on in vitro growth of cells incubated with implants. In mice, implants were well tolerated without any sign of inflammatory reaction. The material allows an elastic compression of urethra compensating a damaged sphincter after trans-urethral sub-mucosal implantation of hydrogel cylinders.

Polymer hydrogels usable for nervous tissue repair.

The implantation of non-resorbable biocompatible polymer hydrogels into defects in the central nervous system can reduce glial scar formation, bridge the lesion and lead to tissue regeneration within the hydrogel. We implanted hydrogels based on crosslinked poly hydroxyethyl-methacrylate (pHEMA) and poly N-(2-hydroxypropyl)-methacrylamide (pHPMA) into the rat cortex and evaluated the cellular invasion into the hydrogels by means of immunohistochemical methods and tetramethylammonium diffusion measurements. Astrocytes and NF160-positive axons grew similarly into both types of hydrogels. We found no cell types other than astrocytes in the pHEMA hydrogels. In the pHPMA hydrogels, we found a massive ingrowth of connective tissue elements. These changes were accompanied by corresponding changes in the extracellular space volume fraction and tortuosity of the hydrogels.

Biological properties of copolymer of 2-hydroxyethyl methacrylate with sulfopropyl methacrylate.

Interaction of organism with non-toxic implanted polymers depends on the physicochemical properties of the implant surface, which influence the adsorption of bioactive proteins and subsequently adhesion and growth of cells. The synthetic hydrogels are known as poorly adhesive surfaces. In this study we demonstrated the adsorption of albumin, fibrinogen, fibronectin, basic fibroblast growth factor, heparin-binding epidermal growth factor-like growth factor and epidermal growth factor to poly(2-hydroxyethyl methacrylate) (pHEMA) and copolymer of 2-hydroxyethyl methacrylate (HEMA) and potassium salt of 3-sulfopropyl methacrylate (SPMAK). The adhesion and growth of 3T3 cells and human keratinocytes on surface of these polymers was tested without and with pretreatment of polymers with heparin-binding epidermal growth factor-like growth factor. The adhesion of mixture of human granulocytes and monocytes to these surfaces was also tested. The strips of both polymers were subcutaneously and intracerebrally implanted into the rat and the extent of foreign body reaction and brain biocompatibility was evaluated. The results showed the extensive adsorption of basic fibroblast growth factor and heparin-binding epidermal growth factor-like growth factor to copolymer containing SPMAK. However the adhesion (and growth) of cells to this type of copolymers was very low. Preadsorption of human plasma to pHEMA clearly stimulated the leukocyte adhesion in contrary to copolymer containing SPMAK. The extent of foreign-body reaction was significantly higher against the pHEMA compared to tested copolymer p(HEMA-co-SPMAK). In conclusion, the tested copolymer was a poorly adhesive substrate that is only poorly recognized by the non-specific immunity, although the adsorption of basic growth factors to this substrate is highly significant. Both polymers were well tolerated by the brain tissue. The phenotype of surrounding neurons was more close to the control neurons in the brain tissue surrounding the p(HEMA-co-SPMAK) implants.

Development of fixable stent for long-term derivation of lower urinary tract.

The object of the envisaged project is the development of easily fixable non-irritant stent of the lower urinary tract. The project has the aim to remove the obstruction and make passage-way through urethra which, after repeated operations, remains narrowed and requires repeated dilatations or, in the patients who have a chronic epicystostomy due to the non-passable urethra. The prosthesis must be self fixable, non-irritant, easily introduceable and removable by endoscopy, enabling normal urination with urinary continence and, last but not least, at a reasonable price.

Synthetic hydrogel capacity to induce formation of foreign-body giant multinucleate cells differs in vivo and in vitro.

The granulomatous reaction accompanied with MGC formation represents the most striking feature of the non-favourable biological tolerance of implanted devices. We compared MGC formation in the course of the granulomatous reaction in vitro and in vivo employing three types of hydrogels whose biocompatibility had been well studied earlier. The efficiency of the in vitro assay for the granulomatous reaction, including MGC formation, was verified employing the nematode Nippostrongylus brasiliensis, a well-known inductor of MGC formation in vitro. The in vitro results demonstrated a very low level of MGC formation in reaction against all three types of hydrogels without polymer-specific differences in comparison with the nematode experiment characterized by a high extent of MGC formation. On the other hand, the extent of MGC formation was implant type-specific in vivo: pHEMA-co-DMAEMA > pHEMA > pHEMA-co-NaMA. These results indicate that in the in vitro assay it was not possible to discriminate among the types of polymers used in the experiment in comparison with the animal experiment. They also indicate potential differences between granuloma formation induced by parasites and by foreign bodies.

[Surgical treatment of recurrent urethral strictures in males after unsuccessful operations]. / Nové moznosti chirurgické lécby muzu po neúspesných operacích recidivujících striktur mocové trubice.

BACKGROUND: Subvesical obstructions of any origin represent a frequent and serious disorder occurring predominantly in males. Often it brings incontinence and/or erectility dysfunction, infection of urinary tract. Relapses of the acute pyelonephritis can turn into chronic tubulointersticial one and terminate in the renal insufficiency. To treat strictures, dilation, intermittent catheterization and recently stent introduction were used. Most suitable appears a stent from composite polymers. The aim of our work was to test properties of stents developed in the Institute of Macromolecular Chemistry ASCR. METHODS AND RESULTS: Stents from composite polymers, which are non-toxic, not-irritable can swell in body fluids and have mechanical properties similar to that of silicone rubber. Properties of the material are functionally graded and the casting or repoussé from the material can subsequently change its shape. Ten patients (males, aged 25 to 78 years) with long urethral strictures in its bulbocavernous part (50%) were treated with this method. Strictures were caused by pelvical fractures (4 times), prostate hypertrophy surgery (4 times), prolonged catheterizations (2 times). All patients were followed for 16 to 26 month and had no severities. CONCLUSION: Our results indicate that stent from composite polymers and silicone may have long-acting effects without irritation or crust formation and beneficially effected healing of the spongio-fibrous process.

Cell-specific nuclear import of plasmid DNA.

One factor limiting the success of non-viral gene therapy vectors is the relative inability to target genes specifically to a desired cell type. To address this limitation, we have begun to develop cell-specific vectors whose specificity is at the level of the nuclear import of the plasmid DNA. We have recently shown that nuclear import of plasmid DNA is a sequence-specific event, requiring the SV40 enhancer, a region known to bind to a number of general transcription factors (Dean DA, Exp Cell Res 1997; 230: 293). From these studies we developed a model whereby transcription factor(s) bind to the DNA in the cytoplasm to create a protein-DNA complex that can enter the nucleus using the protein import machinery. Our model predicts that by using DNA elements containing binding sites for transcription factors expressed in unique cell types, we should be able to create plasmids that target to the nucleus in a cell-specific manner. Using the promoter from the smooth muscle gamma actin (SMGA) gene whose expression is limited to smooth muscle cells, we have created a series of reporter plasmids that are expressed selectively in smooth muscle cells. Moreover, when injected into the cytoplasm, plasmids containing portions of the SMGA promoter localize to the nucleus of smooth muscle cells, but remain cytoplasmic in fibroblasts and CV1 cells. In contrast, a similar plasmid carrying the SV40 enhancer is transported into the nuclei of all cell types tested. Nuclear import of the SMGA promoter-containing plasmids could be achieved when the smooth muscle specific transcription factor SRF was expressed in stably transfected CV1 cells, supporting our model for the nuclear import of plasmids. Finally, these nuclear targeting sequences were also able to promote increased gene expression in liposome- and polycation-transfected non-dividing cells in a cell-specific manner, similar to their nuclear import activity. These results provide proof of principle for the development of cell-specific non-viral vectors for any desired cell type.

Brain lesion-induced alteration of selected phenotypic properties of spleen macrophages and their partial restoration in the course of foreign body reaction against intraperitoneally implanted polymers.

A lesion in the dorsoposterior part of the rat brain septum is known to exert an inhibitory effect on the delayed skin hypersensitivity and incorporation of radiolabeled thymidine into the lymphoid organs. To determine whether distinct properties of macrophages will also be modulated by this type of injury, we have focused upon the monitoring of expression of sugar receptors (lectins). In this study we show a reduction in the number of macrophages expressing carbohydrate-binding sites for asialoglycoproteins (beta-D-galactoside), alpha-D-mannoside and alpha-D-mannoside-6-phosphate in spleen macrophages after the lesion of the dorsoposterior septum of the brain in the rat. The number of ED-1+ macrophages was not influenced. The intraperitoneal injection of beads prepared from the copolymer of 2-hydroxyethyl methacrylate with dimethyl aminoethyl methacrylate (30 wt %) elevated significantly the number of ED-1+ spleen macrophages and number of macrophages with binding site(s) recognizing asialoglycoproteins and alpha-D-mannoside-6-phosphate, respectively. These results indicate that a foreign-body reaction appears to be able to mediate a phenotypic restoration of lectin expression by spleen macrophages altered by the brain lesion. It can be suggested that, for example, a probable production of cytokines by the inflammatory cells colonizing the implanted beads plays a role in this process.

Cultivation and grafting of human keratinocytes on a poly(hydroxyethyl methacrylate) support to the wound bed: a clinical study.

Cultured epithelial sheets on a textile support are used for the treatment of seriously burned patients. In this study we demonstrate a new procedure for the grafting of keratinocytes directly on a polymer cultivation support. This procedure is much easier in comparison with classical techniques, and encouraging results of clinical trials demonstrate the improved healing of the wound bed after the use of this procedure. There is no difference in the cytokeratine pattern (LP-34, cytokeratin-10) of the reconstructed epidermis and normal human skin.

[The histopathologic picture of experimental poisoning with ethylmethacrylates]. / Histopatologický obraz experimentální intoxikace ethylmethakryláty.

Toxic effect of hydroxyethylmethacrylate, acetoxyethylmethacrylate and diethylenglycomethacrylate were studied in rats surviving as long as 1 to 20 days after intramuscular administration. Conspicuous lesion were found only in calf muscles at the site of application. Muscle fibre necroses with inflammatory reaction occurred repeatedly in animals surviving 1-2 days. Newly formed connective tissue replacing impaired muscle fibres was found in rats surviving 5 days. Intoxication related other lesion in rat organs were not identified.

Reactive oxygen species (ROS) generated by xanthine oxidase in the corneal epithelium and their potential participation in the damage of the corneal epithelium after prolonged use of contact lenses in rabbits.

Prolonged use of contact lenses (for 14 days) evoked an imbalance between the activity of xanthine oxidase (an enzyme belonging to reactive oxygen species-generating oxidases) and catalase (an enzyme belonging to reactive oxygen species-scavenging oxidases) in the corneal epithelium of rabbits. The activity of catalase decreased, while xanthine oxidase activity was very high. Of other enzymes studied in the corneal epithelium, the activities of xanthine oxidoreductase, glucoso-6-phosphate dehydrogenase and succinate dehydrogenase were decreased. In contrast, the activities of lactate dehydrogenase and lysosomal hydrolases (acid beta-galactosidase, dipeptidyl peptidase II) were increased and appeared in animals sacrificed immediately after contact lens removal. In rabbits sacrificed later (after 1 h), an additional increase of lactate dehydrogenase and lysosomal hydrolase activities developed in the superficial layers of the corneal epithelium. Catalase supplementation during use of contact lenses prevented both the significant decrease of catalase activity in the corneal epithelium and the development of additional epithelial damage. In contrast, topical treatment with 3-aminotriazole (an inhibitor of catalase) resulted in the nearly complete loss of catalase activity in the corneal epithelium and the appearance of more serious epithelial damage. We conclude that ROS generated by xanthine oxidase induce additional damage of the corneal epithelium related to the use of contact lenses.

Effect of chemical structure of hydrogels on the adhesion and phenotypic characteristics of human monocytes such as expression of galectins and other carbohydrate-binding sites.

The reactivity of diverse immune aspects to the presence of synthetic polymers represents one of the most important aspects of implantable device biocompatibility. In this report, we show the effect of the chemical structure of a synthetic polymer support on monocyte adhesion and selected phenotypic characteristics in vitro as a model for the initial steps of non-self-recognition of an implant. The extent of monocyte adhesion was significantly influenced by the support chemistry. The highest level of monocyte adhesion was observed on a surface copolymer of 2-hydroxyethyl methacrylate with dimethyl aminoethyl methacrylate relative to results of experiments in which poly(2-hydroxyethyl methacrylate) or the copolymer of 2-hydroxyethyl methacrylate with the sodium salt of methacrylic acid was used. Cell adhesion to the polymers tested and to glass was accompanied by enhanced expression of the carbohydrate-binding sites tested for asialoglycoprotein beta-galactosides such as galectins, beta-N-acetylgalactosamine, alpha-mannoside, specific lectin for heparin as well as the lymphokine-macrophage migration inhibitory factor in the monocytes tested. These results suggest the importance of monocyte adhesion to the biomaterial surface for their development into macrophages and further non-self-recognition of the implanted device.