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L Bacáková

Publications and source records attributed to L Bacáková.

At least 19 recordsLinked to original sources
PubMed2004

Cell adhesion on artificial materials for tissue engineering.

Advanced interdisciplinary scientific field of tissue engineering has been developed to meet increasing demand for safe, functional and easy available substitutes of irreversibly damaged tissues and organs. First biomaterials were constructed as "two-dimensional" (allowing cell adhesion only on their surface), and durable (non-biodegradable). In contrast, biomaterials of new generation are characterized by so-called three dimensional porous or scaffold-like architecture promoting attachment, growth and differentiation of cells inside the material, accompanied by its gradual removal and replacement with regenerated fully functional tissue. In order to control these processes, these materials are endowed with a defined spectrum of bioactive molecules, such as ligands for adhesion receptors on cells, functional parts of natural growth factors, hormones and enzymes or synthetic regulators of cell behavior, incorporated in defined concentrations and spatial distribution against a bioinert background resistant to uncontrolled protein adsorption and cell adhesion.

Animals↗
PubMed2003

Cell adhesion on polytetrafluoroethylene modified by UV-irradiation in an ammonia atmosphere.

We report on the modification of polytetrafluoroethylene (PTFE) by exposure to the ultraviolet (UV) light of a Xe(2)*-excimer lamp at a wavelength of 172 nm in an ammonia atmosphere. Typical treatment times were up to 30 min. Subsequently, the samples were grafted with the amino acid alanine from an aqueous solution. The samples were characterized by means of optical transmission spectroscopy, laser-induced fluorescence and contact-angle measurements. We studied the adhesion of rat aortic smooth muscle cells (SMC) and mouse fibroblasts (3T3 cells) to the modified polymer samples using an in vitro technique, where the population density and spread of adhering cells is determined 24 h after seeding by image analysis. For both cell types the exposure of PTFE to UV-light in an ammonia atmosphere resulted in a significant increase in the number of adhering cells and in the size of their spreading area. The grafting with alanine enhanced this effect. Additional experiments with human endothelial cells (HEC) also demonstrated improved adhesion to modified PTFE. Thus, PTFE modified by our method appears to be a promising material for fabrication of artificial vascular prostheses and implants or for cultivation of skin substitutes.

3T3 Cells↗
PubMed2002

Colonization of ion-modified polyethylene with vascular smooth muscle cells in vitro.

Polyethylene (PE) foils were implanted with 40 and 150 keV Ar+ ions to the fluences from 1 x 10(13) to 1 x 10(15) cm(-2). Production of conjugated double bonds, characterizing degradation of the PE surface layer, was studied using UV-VIS spectroscopy. Wettability of the PE surface, determined by conventional goniometric techniques, was shown to be an increasing function of both ion energy and fluence. It was also increased after exposure of PE to serum-supplemented cell culture media. Cell culture experiments showed that the ion irradiation significantly increased the adherence of vascular smooth muscle cells (VSMC) and their subsequent growth on the PE surface. On day 1 after seeding, the number of initially adhered VSMC exhibited two maxima. On day 3 after seeding. these maxima disappeared, which was partially due to a significantly shorter doubling time of VSMC. On the other ion-modified samples. the doubling time did not differ significantly from that on the unmodified PE. Enzyme-linked immunosorbent assay revealed increased concentration of talin, a protein of focal adhesion plaques, and alpha-actin, a marker of VSMC differentiation, in cells on ion-implanted surfaces. It can be concluded that the ion irradiation supports the adhesion and differentiation of VSMC without excessive proliferation of these cells.

Animals↗
PubMed2001

Polishing and coating carbon fiber-reinforced carbon composites with a carbon-titanium layer enhances adhesion and growth of osteoblast-like MG63 cells and vascular smooth muscle cells in vitro.

Carbon fiber-reinforced carbon composites (CFRC) are considered to be promising materials for orthopedic and dental surgery. Their mechanical properties can be tailored to be similar to those of bone, and their chemical composition (close to pure carbon) promises that they will be tolerated well by the surrounding tissue. In this study, CFRC composites were fabricated from phenolic resin and unidirectionally oriented Torayca carbon fibers by carbonization (1000 degrees C) and graphitization (2500 degrees C). The material then was cut with a diamond saw into sheets of 8 x 10 x 3 mm, and the upper surface was polished by colloidal SiO2 and/or covered with a carbon-titanium (C:Ti) layer (3.3 microm) using the plasma-enhanced physical vapor deposition method. Three different kinds of modified samples were prepared: polished only, covered only, and polished + covered. Untreated samples served as a control. The surface roughness of these samples, measured by a Talysurf profilometer, decreased significantly after polishing but usually did not decrease after coating with a C:Ti layer. On all three modified surfaces, human osteoblast-like cells of the MG63 line and rat vascular smooth muscle cells (both cultured in a Dulbecco's minimum essential medium with 10% fetal bovine serum) adhered at higher numbers (by 21-87% on day 1 after seeding) and exhibited a shorter population doubling time (by 13-40%). On day 4 after seeding, these cells attained higher population densities (by 61-378%), volume (by 18-37%), and protein content (by 16-120%). These results were more pronounced in VSMC than in MG63 cells and in both groups of C:Ti-covered samples than in the polished only samples. The release of carbon particles from the CFRC composites was significantly decreased--by 8 times in the polished only, 24 times in the covered only, and 42 times in the polished + covered samples. These results show that both polishing and carbon-titanium covering significantly improve the biocompatibility of CFRC composites in vitro, especially when these two modifications are combined.

Animals↗
PubMed2001

Adhesion and proliferation of rat vascular smooth muscle cells (VSMC) on polyethylene implanted with O+ and C+ ions.

Polyethylene was implanted with 30-keV oxygen (PE/O+) or 23-keV carbon ions (PE/C+) at 10(13) to 5 x 10(15) ions cm(-2) doses in order to improve the adhesion of vascular smooth muscle cell (VSMC) to the polymer surface in vitro because of its oxidation and carbon-enrichment. The concentration of -CO- groups in the PE/O+ and PE/C+ samples increased only up to doses of 3 x 10(14) and 10(15) ions cm(-2), respectively, and then declined. At the same time, the concentration of these groups, measured at a dose of 3 x 10(14) ions cm(-2), was higher in PE/O+ than in PE/C+ samples. Similarly, the number of initially-adhering rat VSMC (24 h after seeding) increased only up to a dose of 3 x 10(13) and 10(15) ions cm(-2) on PE/O+ and PE/C+ samples, respectively. In addition, between doses of 10(13) and 10(14) ions cm(-2), this number was about two to three times higher on PE/O+ samples. On the other hand, the surface wettability increased proportionally to the implanted ion dose, especially above a dose of 10(14) ions cm(-2). Thus, the number of initially-adhered cells appeared to be positively correlated with the amount of the oxygen group present at the polymer surface rather than with the surface wettability. The higher cell adhesion was accompanied by adsorption of fluorescent dye-conjugated collagen IV in larger amounts. The highest numbers of initially-adhered cells were usually associated with the lowest rates of subsequent proliferation (measured by the doubling time, BrdU labelling and M

Animals↗
PubMed2001

A sex-related difference in the hypertrophic versus hyperplastic response of vascular smooth muscle cells to repeated passaging in culture.

Activation of growth of vascular smooth muscle cells (VSMC) in adults participates in pathogenesis of dysplastic diseases of the vascular system. In this study, we examined the impact of gender of rat donors on the degree of hyperplastic and hypertrophic responses of VSMC in cultures subjected to repeated passaging. The cells were derived from the outgrowth zone of explants of the thoracic aorta and were studied up to passage 45. Under these conditions, the cells undergo repeated growth stimulation by the serum growth factors mimicking some pathological situations in vivo. At lower passages (5-7), the cells from both sex donors did not differ significantly in their doubling time, maximum population density, protein content and ploidy. At higher passages (40-45), we found that the hyperplastic response, monitored by doubling time and BrdU-revealed DNA synthesis, was more intense in VSMC of male origin. In contrast, female-derived cells reacted by more prominent hypertrophic changes. The latter included a relatively higher increase in the volume and protein content of cells. As indicated by the DNA content histograms and chromosome numbers, these cells also showed a higher degree of passage-dependent polyploidization. In addition, the female-derived VSMC were found to be more effective in adhesion to the growth support evidenced by wider spreading and higher resistance of these cells to trypsin-mediated detachment as well as higher expression of some integrin and cytoskeletal molecules. These features could partly account for the slower proliferation and polyploidization of these cells. The results suggest that rat VSMC populations of male and female origin contain cells which are intrinsically different with respect to their capability of reacting to growth stimuli. The lower responsiveness of female-derived cells to growth stimuli may contribute to less frequent formation of hyperplastic vascular lesions in female organisms.

Actins↗
PubMed2000

Fluorine ion-implanted polystyrene improves growth and viability of vascular smooth muscle cells in culture.

Vascular smooth muscle cells derived from the rat aorta were cultured on unmodified or F(+) ion-implanted polystyrene (5 x 10(12) or 5 x 10(14) ions/cm(2), energy 150 keV). In 1-day-old cultures, the cells adhered to the modified polystyrene in higher numbers and over larger contact areas. Increased resistance of the cells to trypsin-mediated detachment from the growth support indicated an improved adhesion of cells to the modified polymer at later culture intervals. The cells cultured on ion-modified polymers also were larger and had a higher total protein content. By use of immunocytochemistry, several specific protein species were increased, including the cytoskeletal alpha-actin and vimentin and the plasma membrane-associated vinculin, talin, alpha-v integrins, ICAM-1, and VCAM-1, which account for stronger cell-cell and cell-extracellular matrix adhesion. The lower number of cells found floating in the medium suggests that the spontaneous detachment of cells from the modified polystyrene was lower and that the viability of the adhered cell population was higher. As was shown by the two-parameter flow-cytometric measurements of BrdU incorporation and DNA content, as well as by (3)H-thymidine autoradiography, the cell proliferation on samples modified by the dose of 5 x 10(12) ions/cm(2) was similar to that in controls; and at the dose of 5 x 10(14) ions/cm(2), it tended to be even lower. The cells grown on the polymer implanted with the dose of 5 x 10(12) ions/cm(2) responded to a new artificially created cell-free area in a confluent cell layer by more intense migration whereas at the dose of 5 x 10(14) ions/cm(2), the migration ability of cells was similar to that on the unmodified polymer. The data revealed a higher biocompatibility of ion-implanted polystyrene with vascular smooth muscle cells in culture. There was better adhesion, differentiation, and survival, and there was neither excessive migration nor proliferation.

Animals↗
PubMed2000

Molecular mechanisms of improved adhesion and growth of an endothelial cell line cultured on polystyrene implanted with fluorine ions.

Endothelial cells derived from the bovine pulmonary artery (line CPAE, CCL 209, American Tissue Culture Collection, Rockville, MD, USA) were cultured on pristine or fluorine ion-irradiated polystyrene (5 x 10(12) or 5 x 10(14) F ions/cm2, 150 keV). At 24-h post-seeding interval, the number of cells which adhered to the ion-modified polystyrene was significantly higher than on the unmodified material (+20 and +58% in cultures with the polystyrene irradiated by lower and higher ion doses, respectively). On day 7, the populations cultured on the irradiated substrates grew to higher densities, exceeding the controls at the lower and higher ion doses by 69 and 180%, respectively. The cells on ion-implanted samples were also larger (+70-95% and +90-99% at the lower and higher ion doses, respectively) and contained more protein (+16% at both ion doses). As was shown by ELISA, the polystyrene irradiated by the higher ion dose enhanced the expression of a cytoskeletal protein, vimentin (+65%) and protein of focal adhesion plaques, talin (+15%). The content of integrin alpha5beta1 (VLA-5), receptor for fibronectin, was increased at both lower and higher ion doses (+22 and +57%). In contrast to this, the content of ICAM-1 and vinculin was similar in cells grown on both pristine and ion-irradiated growth substrates. Moreover, the expression of VCAM-1 and ELAM-1 was lower by 11-14% in both ion dose groups. The present study has shown that ion implantation of polymers improves the adhesion and growth of endothelial cells without elevating the expression of immunoglobulin and selectin types of adhesion molecules. This surface modification should promote colonization of an artificial vascular prosthesis by endothelial cells and make it less vulnerable by immune system cells of the recipient.

Animals↗
PubMed2000

Gender differences in growth of vascular smooth muscle cells isolated from hypertensive and normotensive rats.

Higher male sensitivity to atherosclerotic and hypertensive events was a reason to study sex differences in migration and proliferation of vascular smooth muscle cells (VSMC) isolated from male and female spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) controls. Outgrowth of cells from explants, doubling time, curves of cumulative labeling and the length of cell cycle were measured in aortic VSMC. Systolic and mean arterial pressures were higher in males than in females of the two strains. The migration of cells from male explants was significantly faster than those from female aortas in both strains. The doubling time was always shorter in male VSMC than in those from females and this was more apparent in the late exponential phase of growth. The thymidine incorporation into newly synthesized DNA, which was enhanced in SHR compared to WKY cells, was also higher in male cells compared to female ones. Cell cycle was always shorter in male than in female VSMC due to the shorter G1 phase. In contrast, shorter S phase caused shorter cell cycle in SHR compared to WKY VSMC. Consequently, the shortest cell cycle was found in VSMC from SHR males with the highest blood pressure. It can be concluded that gender and genotype are two independent factors participating in the control of migration and proliferation of VSMC.

Aging↗
PubMed1999

Gender-related differences in adhesion, growth and differentiation of vascular smooth muscle cells are enhanced in serum-deprived cultures.

In 1-day cultures with 10% serum, the number of rat aortic smooth muscle cells (VSMC) adhering to the growth support was similar in cells from both sexes, whereas in 1% serum, the number of VSMC from male donors was lower. In 10% serum medium, the doubling time was significantly shorter and the number of [3H]thymidine-labelled nuclei was higher in cells of high passage from male rats. In serum-free medium, these differences increased and were also seen in cells of low passage number. Morphologically, the cells in male-derived cultures at higher passage number were mainly spindle-shaped, formed well-developed 'hills and valleys' and possessed longitudinally oriented bundles of alpha-actin-containing microfilaments. Most cells from female rats were flat, polygonal, the multilayered 'hills' were less prominent, with alpha-actin microfilaments forming a mesh-like network.

Animals↗
PubMed1999

Influence of macrophages and macrophage-modified collagen I on the adhesion and proliferation of vascular smooth muscle cells in culture.

The adhesion, proliferation and morphology of rat vascular smooth muscle cells (VSMC) in cocultures with macrophages or in cultures on type I collagen modified by activated macrophages were evaluated. In the first set of experiments, rat alveolar macrophages were added to 24-hour-old VSMC cultures. Between days 2 and 5 after VSMC seeding, the population densities and doubling times of cells were similar in both VSMC-macrophage and pure VSMC cultures. However, from day 5, the cocultures proliferated about two times more rapidly and on day 7, they reached higher cell population density by 40%. The pure macrophage cultures did not proliferate. In the second set of experiments, rat alveolar macrophages were activated by non-toxic TiO2 dust to produce reactive oxygen species and incubated for 120 min with collagen I. The collagen was then adsorbed on plastic culture dishes and seeded with VSMC. The collagen exposed for 10 min only, the unmodified collagen and pure culture dishes were used as control growth supports. On all four tested substrates, the number of initially adhered cells was similar, but on the collagen modified for 120 min, the cells were less spread. Moreover, on day 2 to 3 after seeding, some cells on this collagen became vacuolated and detached spontaneously from the growth support. The remaining VSMC, however, rapidly proliferated, so that on day 9, the cell population density on 120-min-modified collagen was similar as on both control collagens and significantly higher compared to that on uncoated dishes. Our results suggest that 1. The delayed growth-stimulating effect of macrophages on VSMC-macrophage mixed population is probably due to autocrine production of mitogens by both cell types rather than due to an acute effect of short-living oxygen radicals released from macrophages immediately after adding to VSMC cultures. 2. The effect of collagen I exposed to activated macrophages for 120 min is slightly cytotoxic, which could, however, stimulate a release of mitogens from damaged as well as surviving VSMC.

Animals↗
PubMed1999

Growth and differentiation of the vascular smooth muscle and endothelial cells cultured on fluorine ion-implanted polystyrene.

The rat vascular (SMCs) and bovine endothelial cells (BECs) were cultured on conventional or fluorine ion-implanted polystyrene (5 x 10(12) and 5 x 10(14) fluorine ions/cm2). The cells grown on the implanted growth supports showed better adherence, higher volume and higher total protein content. The immunocytochemical analysis revealed that SMCs contained more of the cytoskeletal vimentin and the vascular SMC-specific alpha-actin as well as several cell adhesion-mediating molecules (vinculin, talin, alpha(v)-integrin and ICAM-1). In BECs, only the content of vimentin and talin increased, while expression of ICAM-1 was unchanged. The data suggest that cells on the ion implanted polymers could be more viable and that increased expression of some adhesion molecules mediating interactions with the host immune system is cell type-dependent.

Actins↗
PubMed1999

Different responsiveness of male and female rat aortic smooth muscle cells (SMCs) to repeated passaging in culture.

The smooth muscle cell (SMC) cultures were prepared from the aorta of male and female 8-week-old rats and used at passage 5-7 or 40-45. On day 1, low-passaged cells of both sex groups adhered to growth supports at similar numbers while after repeated passaging the adherence of female-derived cells was higher. These cells had also higher total protein content and contained more of the SMC specific alpha-actin, vimentin and alpha(v) integrins. Compared to the male type of cultures, the high passaged cells of female origin cycled at a slower rate and were undergoing massive polyploidization. Male-derived cells remained of the same morphology, ploidy and the differentiation status at all passages. Their passage response consisted mainly in faster cycling and growth to higher population densities. The data could be of importance for explanation of different incidence of hyperplastic vascular diseases in males and females.

Actins↗
PubMed1998

The influence of angiotensin II on sex-dependent proliferation of aortic VSMC isolated from SHR.

The growth response to angiotensin II (Ang II) was studied using cultured vascular smooth muscle cells (VSMC) isolated from the aortae of male and female spontaneously hypertensive rats (SHR). Systolic and mean arterial blood pressure of 10-week-old males was significantly higher when compared to age-matched females. The specific growth rate of male VSMC was significantly higher on the third and sixth day after synchronisation. Angiotensin II in concentration 10(-7) M stimulated the specific growth rate only in male VSMC during the exponential phase of growth. Moreover, doubling time was 3 hours shorter in male VSMC in comparison with the females. Our results suggest that both the increased specific growth rate and augmented growth-response of male VSMC to Ang II may explain the higher sensitivity of males to hypertensive stimuli.

Angiotensin II↗
PubMed1997

Oxidized collagen stimulates proliferation of vascular smooth muscle cells.

We hypothesize that the vascular smooth muscle proliferation after lung injury results from oxidative damage to the matrix proteins in the walls of pulmonary blood vessels. The smooth muscle cells (SMC) isolated from rat aorta were cultured on the surface coated with oxidized and nonoxidized (control) collagen of type I. Oxidation of collagen was induced by UV irradiation and characterized by fluorescence tridimensional spectral arrays and by gel electrophoresis. From day 1 to 6 of the experiment, SMC proliferated more rapidly on the oxidized collagen than on the control surface. At high SMC population densities (day 9 of experiment) the difference disappeared. After 10 min of trypsinization the cells growing on oxidized collagen rounded and detached completely from the growth surface. The control cells on nonoxidized collagen detached only after 30 min of trypsinization. We conclude that oxidation of collagen of vascular wall matrix may participate in stimulation of SMC proliferation after oxidant tissue injury.

Animals↗
PubMed1997

Positive cooperativity of acetylcholine and other agonists with allosteric ligands on muscarinic acetylcholine receptors.

It is well known that allosteric modulators of muscarinic acetylcholine receptors can both diminish and increase the affinity of receptors for their antagonists. We investigated whether the allosteric modulators can also increase the affinity of receptors for their agonists. Twelve agonists and five allosteric modulators were tested in experiments on membranes of CHO cells that had been stably transfected with genes for the M1-M4 receptor subtypes. Allosterically induced changes in the affinities for agonists were computed from changes in the ability of a fixed concentration of each agonist to compete with [3H]N-methylscopolamine for the binding to the receptors in the absence and the presence of varying concentrations of allosteric modulators. The effects of allosteric modulators varied greatly depending on the agonists and the subtypes of receptors. The affinity for acetylcholine was augmented by (-)-eburnamonine on the M2 and M4 receptors and by brucine on the M1 and M3 receptors. Brucine also enhanced the affinities for carbachol, bethanechol, furmethide, methylfurmethide, pilocarpine, 3-(3-pentylthio-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahydro-1- methylpyridine (pentylthio-TZTP), oxotremorine-M, and McN-A-343 on the M1, M3, and M4 receptors, for pentylthio-TZTP on the M2 receptors, and for arecoline on the M3 receptors. (-)-Eburnamonine enhanced the affinities for carbachol, bethanechol, furmethide, methylfurmethide, pentylthio-TZTP, pilocarpine, oxotremorine and oxotremorine-M on the M2 receptors and for pilocarpine on the M4 receptors. Vincamine, strychnine, and alcuronium displayed fewer positive allosteric interactions with the agonists, but each allosteric modulator displayed positive cooperativity with at least one agonist on at least one muscarinic receptor subtype. The highest degrees of positive cooperativity were observed between (-)-eburnamonine and pilocarpine and (-)-eburnamonine and oxotremorine-M on the M2 receptors (25- and 7-fold increases in affinity, respectively) and between brucine and pentylthio-TZTP on the M2 and brucine and carbachol on the M1 receptors (8-fold increases in affinity). The discovery that it is possible to increase the affinity of muscarinic receptors for their agonists by allosteric modulators offers a new way to subtype-specific pharmacological enhancement of transmission at cholinergic (muscarinic) synapses.

Acetylcholine↗
PubMed1997

Monitoring of the boron neutron-capture reaction by a simple animal model.

Sodium borocaptate (BSH, Na2Bl2HllSH), a slow neutron-capture compound, was injected into the left forebrain ventricle of 1-week-old rats (150 microg BSH/3 microl phosphate buffered saline). After 90 min, the animals were irradiated by epithermal neutrons (LVR-15 nuclear reactor in Rez near Prague, flux density 8.8 x 10(7) neutrons cm-2 s-1, 8 MW reactor power, 8.2 cGy/min) for 5, 10 or 20 min. The brains were examined histologically 8 h after irradiation. In animals irradiated for 5 to 10 min (41 and 82 cGy-Eq, respectively) lethal damage of cells was found in the external granular layer of the cerebellum and the subependymal layer of the forebrain. Irradiation for 20 min (164 cGy-Eq) caused more extensive destruction of cell populations in these regions and, in addition, dead cells appeared also in the more differentiated postmitotic compartments, namely the deeper layers of the cerebellum, layers II/III of the cerebral cortex and corpus callosum. In the forebrain periventricular layer, the extent of cell damage was declining towards the olfactory bulbs. In intact animals, as well as in those injected only with the 150 microl phosphate buffered saline, the radiation damage was low and limited only to the most sensitive dividing populations of the cerebellum and the forebrain. The study demonstrates a differentiation-dependent damage of the rat brain cells by alpha particles and presents a simple model for evaluation of the biological effectiveness of slow neutron beams constructed for neutron-capture therapy of tumors.

Animals↗