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Biomedical subjects

J Svoboda

Publications and source records attributed to J Svoboda.

At least 37 records · Page 2Linked to original sources
PubMed1999

Proviral load and expression of avian leukosis viruses of subgroup C in long-term persistently infected heterologous hosts (ducks).

Proviral DNA load and expression of avian leukosis viruses of subgroup C (ALV-C) in ducks infected in mid embryogenesis were studied using quantitative PCR, RT-PCR, in situ hybridization employing ALV-specific riboprobe, and immunohistochemistry. A group of long-term surviving, non-reviremic ducks was selected for the study and compared to control reviremic animals in order to obtain information about persisting retroviruses in different duck tissues. A widespread distribution of proviruses in the tested tissues was found, but the proviral load was significantly lower in non-reviremic in comparison to reviremic animals. The only exception were brain and blood cells, in which no significant difference in the quantity of integrated proviruses was found between both categories of ducks, thus indicating an exceptional position of the brain and blood cells among all tested tissues. Contrary to reviremic, the proviruses were not transcribed in non-reviremic ducks, with the exception of brain and thymus. In the majority of non-reviremic ducks viral RNA was revealed in the brain, but no infectious virus could be recovered from this tissue. The opposite situation was observed in the thymus, where infectious virus was recovered but viral RNA remained below the detection limit of the assay. As revealed by in situ analysis, infected cells were either disseminated or focally distributed in tissues. From the long-term follow up of ALV-C in intraembryonally infected ducks we conclude that this model is suitable for the study of retrovirus persistence accompained both by the presence and absence of reviremias. The possible consequences of transmission and long-term persistence of retroviruses in the heterologous host for retroviral evolution are discussed.

Animals↗
PubMed1999

Epidemiological analysis of Pseudomonas aeruginosa strains isolated from a selected patient population in Brno, Czech Republic.

In the 1996/97 period, 1,413 Pseudomonas aeruginosa (PA) strains were isolated from 843 patients of the Brno teaching hospitals of St. Anne and Bohunice together with small groups from other hospitals. In the same period, 203 PA strains, used as controls, were isolated from 187 patients treated outside hospitals. Statistical evaluation was based on 1,023 hospital isolates and 189 control strains. A total of 16 isolates were recovered from the hospital environments and two from therapeutic swimming pools. The epidemiological analysis of these PA strains was based on pyocin typing, serological typing and phage typing. The most frequently occurring pyocin types amongst our strains fell into 8 pyocin-type groups. The prevailing groups differed significantly between the hospital patient and control groups. Similarly, serological typing identified differences in the predominant serotypes between hospital and control patients. The phage typing method revealed that the control PA strains were significantly more sensitive to 21 polyvalent bacteriophages used than the hospital isolates. In relation to pyocin and serological typing, strains isolated from the hospital environment showed characteristics similar to those of the PA strains isolated from hospital patients. Our results indicate that the majority of strain isolated from hospitalised patients had their origin from human or inanimate contacts in the hospitals.

Bacteriophage Typing↗
PubMed1999

[Elastase activity of gram-negative non-fermenting bacteria and two variations of a simple method for their detection].

The ability to produce elastase was examined in 1,168 strains of 11 representative genera of Gram-negative non-fermenting bacteria isolated from weakened patients in the course of eight years. Detection of the elastase activity was performed parallelly using the conventional method and its pulverisation variant. The difference between the results accomplished by the classical method and its pulverisation variant (53.9% versus 60.5% detected elastase-positive strains) was highly significant (p < 0.05). Among the examined strains 500 (42.8%) displayed elastase activity. The highest proportion of the elastase positive strains was found in Pseudomonas aeruginosa (84.5%), P. fluorescens (60.5%), P. stutzeri (56.4%), Burkholderia cepacia (43.8%) and Shewanella putrefaciens (39.7%). Elastase activity detected in Burkholderia cepacia, Pseudomonas alcaligenes, P. mendocina, P. putida, P. stutzeri and in Shewanella putrefaciens is apparently described for the first time.

Bacteriological Techniques↗
PubMed1999

[Substituted benzo[b]thiophenes with anti-leukotriene activity].

The cyclizing reaction of cinnamic acid (Ia) with thionyl chloride was optimized and a series of 3-chloro-6-subst.benzo[b]thiophene-2-carbonyl chlorides Va-Vn was prepared. Chloride Va was transformed into a series of N-aryl-3-subst. (Cl, OCH3, OH) benzo[b]thiophene-2-carboxamides VII, IX, X. The drugs were subjected to an evaluation of selected antileucotriene activities in vitro and of the anti-inflammatory effect in vivo. In agreement with the assumed mechanism, inhibition of the ear inflammation is conditioned by the antileucotriene activity, whereas inhibition of the carrageen oedema is not due to this mechanism alone.

Animals↗
PubMed1999

Residual strain in human atherosclerotic coronary arteries and age related geometrical changes.

The opening angles of 16 rings excised from human coronary arteries with different degrees of atherosclerosis were determined 10 hours after death. Atherosclerosis, as a chronic inflammatory response of arterial endothelium and intima, is defined by three degrees of its development. The opening angle decreases more or less linearly with the distance from the right coronary artery orifice. This is in accordance with the functional requirements posed on blood transport into the coronary arteries. A decrease of the opening angle with age is affected by hardening of the arterial wall, among other factors. This is in accordance with a stochastic model of age related changes in the initial modulus of elasticity of the coronary artery. A part of the free energy of smooth muscle cells, fibroblasts, collagen and elastic fibres is used not for creating residual strain but for remodeling the arterial wall structure. The opening angle is also considerably affected by the degree of atheroclerosis. The dependence on age of the external diameter and the thickness of the intact left and right coronary arteries in the vicinity of the aortic sinus was also analyzed in two female and two male subjects. To ensure the objectivity of the results it is necessary to carry out additional experiments and studies in vivo.

Adolescent↗
PubMed1998

Sp1 binding sites inserted into the rous sarcoma virus long terminal repeat enhance LTR-driven gene expression.

Although the Rous sarcoma virus (RSV) long terminal repeat (LTR) is an efficient promoter of transcription, most RSV proviruses are down-regulated upon retroviral integration in non-permissive mammalian cells. Among other mechanisms, DNA methylation has been shown to be involved in proviral silencing. The presence of Sp1 binding sites has been demonstrated to be essential for protection of a CpG island and also non-island DNA regions from de novo methylation. Also, the presence of these sites in the LTRs correlates with the transcriptional activity of certain proviral structures. Using transient and stable transfection assays, we demonstrate that insertion of Sp1 binding sites into the RSV LTR remarkably increases expression of the LTR-driven genes in permissive and non-permissive cells, despite the reported negative effect of insertion of the non-specific DNA into the LTR promoter/enhancer sequences. Particular arrangement of inserted Sp1 sites was effective even in stably transfected reporter gene constructs into non-permissive mammalian cells, where additional factors exert negative effects on expression.

Animals↗
PubMed1998

Molecular biology of cell nonpermissiveness to retroviruses. Has the time come?

The problem of host cell nonpermissiveness to retrovirus infection is characterized and illustrated on several retroviral models, including the role of viral receptors, cell fusion, and endogenous retroviral genomes as modifiers of the outcome of retroviral infection. Special attention is paid to different barriers against the infection of mammalian cells with avian leukosis/sarcoma viruses (ALV/ASV). Even when avian retroviruses become integrated in mammalian cells, several blocks at the level of provirus expression, processing of viral RNAs, and posttranslational modification prevent virus production in such virogenic cells. The significance of these blocks and new strategies making it possible to overcome some of them are discussed in relation to the development of ALV/ASV-based vectors suitable for gene therapy in mammals.

Animals↗
PubMed1998

Cloning of a novel Y-box homology protein (chkYB-1HP) cDNA lacking the cold-shock domain.

We have isolated the complete cDNA clone of a novel 262 amino acid Chicken YB-1 Homology Protein (chkYB-1HP) by screening a chicken embryo cDNA expression library. While the chkYB-1HP is identical over its carboxyl-terminal 78 amino acids with the Y-box protein YB-1, it differs strikingly from all other Y-box transcription factors by lacking the cold-shock domain (CSD). We propose that proteins like chkYB-1HP that lack the CSD, but retain the hydrophilic carboxyl domain could regulate Y-box proteins through the formation of heterodimeric complexes.

Amino Acid Sequence↗
PubMed1996

Phenotypic changes induced by wild type and variant c-src genes carrying C-terminal sequence alterations.

We previously reported the isolation of PR2257, a novel avian sarcoma retrovirus which transduced the c-src protooncogene. The v-src gene of PR2257 differs from the c-src gene by a sequence change after amino acid 525, resulting in the replacement of tyrosine 527 by a valine, and an extension of the open reading frame into the non coding region of c-src. We investigated the respective roles of Tyr527 mutation and of the C-terminal extension in activating the oncogenic properties of c-src. Therefore we overexpressed the wild type c-src gene and c-src variants, carrying either a substitution of tyrosine 527 or an extension of the C-terminus or both modifications in combination, in chicken embryo fibroblasts and post mitotic neuroretina (NR) cells, using replication defective retroviruses. We also used in vivo inoculation of plasmid DNA to assess the tumorigenicity of the various c-src genes. We report that, in contrast to previous results, overexpression of c-src is sufficient to induce NR cell division. While mutation of tyrosine 527 alone significantly activates c-src transforming and tumorigenic properties, its combination with the C-terminal extension of PR2257 confers to this gene full oncogenic properties and increased metastatic potential as compared to the v-src of Rous sarcoma virus strains.

Animals↗
PubMed1996

[Treatment of duodenal ulcer with pantoprazole. A multicenter study].

BACKGROUND: The inhibition of H+/K(+)-ATPase (proton pump) of gastric parietal cells by substituted benzimidazoles represents a new therapeutic approach in conditions connected with hypersecretion of hydrochloric acid. Pantoprazol is the newest member of this group of drugs. Monotherapy of duodenal ulcer with pantoprazole or ranitidine was evaluated in terms of healing rate, tolerance and compliance. METHODS AND RESULTS: A double-blind, parallel-group comparing study (double dummy technique) of treatment florid duodenal ulcer (diameter 5-20 mm) was performed in 95 subjects (age 18-74 years). The active substances were either pantoprazole (40 mg before breakfast-47 subjects) or ranitidine (300 mg before bedtime-48 subjects). The average diameter of the ulcer and subjective complaints before treatment were comparable in both groups. After 2 weeks of pantoprazole therapy 88.5% of ulcers (a) and 87.2% patients (b) were cured, whereas the corresponding values in the ranitidine series amounted to 66% (a) and 62.5% (b) only (p = 0.006 for both (a) and (b)). The rate of healing in relative values of reducing the ulcer size was significantly higher after 2 weeks of pantoprazole therapy (p = 0.026 (a) and 0.0027 (b)). All ulcers healed after 4 weeks of this regimen. The difference between the pantoprazole and ranitidine series after 4 weeks was closely above the 5% level of significance (p = 0.589 for (a) and 0.0588 for (b) respectively). This was due to the low number of patients in both groups at this time interval, particularly in the pantoprazole group. Pain during day-time and regurgitation were observed significantly more frequently after ranitidine therapy. The compliance was very good and practically no adverse effects of pantoprazole therapy were observed. CONCLUSIONS: The healing rate of duodenal ulcer with pantoprazole monotherapy (40 mg before breakfast) was significantly higher than with ranitidine (300 mg before bedtime). The compliance of patients and the tolerance of pantoprazole were very good and its administration was not associated with any side-effects.

2-Pyridinylmethylsulfinylbenzimidazoles↗
PubMed1996

The LTR, v-src, LTR provirus in H-19 hamster tumor cell line is integrated adjacent to the negative regulatory region.

The tumor hamster cell line H-19 harbors a single copy LTR, v-src, LTR provirus that becomes permanently transcriptionally suppressed in morphological revertants segregating at high rate from this cell line. Our previous data document that the provirus suppression is mediated by epigenetic cell-regulatory mechanisms. In this report, we concentrate on cellular sequences neighboring the integration site. The locus is unique for Syrian hamster and is not detectable in DNA of several animal species. No restriction sites that usually hint at the presence of CpG islands were found in the significantly close vicinity of the provirus. Nevertheless, the chromosomal DNA flanking the provirus is rich in GC content (57.8%). We localized a 0.5-kb region downstream from the provirus that remarkably inhibits transcription in the transient expression assay and is effective both on the homologous RSV LTR promoter/enhancer and heterologous SV40 promoter. We propose that a cellular trans-acting factor is involved in the silencing of the reporter gene. Since this activity is comparable both in transformed and revertant cells, we speculate that this down-regulatory region makes the permissive integration locus prone to provirus silencing initiated by other fluctuating stimuli.

Animals↗
PubMed1996

X-irradiation-induced changes in the diffusion parameters of the developing rat brain.

Three diffusion parameters of brain tissue, extracellular space volume fraction (alpha), tortuosity (lambda) and non-specific uptake (kappa') of tetramethylammonium were studied in the somatosensory neocortex and subcortical white matter of the rat during postnatal development (postnatal days 2-21) after X-irradiation at postnatal days 0-1. The diffusion parameters were determined from extracellular concentration-time profiles of tetramethylammonium. The tetramethylammonium concentration was measured in vivo with ion-selective microelectrodes positioned 130-200 microns from an iontophoretic source. X-irradiation with a single dose of 40 Gy resulted in typical early morphological changes in the tissue, namely cell death, DNA fragmentation, extensive neuronal loss, blood-brain barrier damage, activated macrophages, astrogliosis, increase in extracellular fibronectin and concomitant changes in all three diffusion parameters. The changes were observed as early as 48 h post-irradiation (at postnatal days 2-3) and still persisted at postnatal day 21. On the other hand, X-irradiation with a single dose of 20 Gy resulted in relatively light neuronal damage and loss, while blood-brain barrier damage, astrogliosis and changes in diffusion parameters were not significantly different from those found with 40 Gy. It is known that the volume fraction of the extracellular space in the non-irradiated cortex is large in newborn rats and diminishes with age [Lehmenkühler A. et al. (1993) Neuroscience 55, 339-351]. X-irradiation with a single dose of 40 or 20 Gy blocked the normal pattern of volume fraction decrease during postnatal development, and in fact brought about a significant increase. At postnatal days 4-5, alpha increased to 0.49 +/- 0.036 in layer III, 0.51 +/- 0.042 in layer IV, 0.48 +/- 0.02 in layer V, 0.48 +/- 0.028 in layer VI and 0.48 +/- 0.025 in the white matter. The large increase in alpha persisted three weeks after X-irradiation. Tortuosity and non-specific uptake decreased significantly at postnatal days 2-5; at days 8-9 they were not significantly different from those of control animals, while they increased significantly at days 10-21. Less pronounced but significant changes in all three diffusion parameters were also found in areas in the ipsilateral hemisphere adjacent to directly X-irradiated cortex. Compared to the control animals [Lehmenkühler A. et al. (1993) Neuroscience 55, 339-351], a significant decrease of alpha, lambda and kappa' was found in the contralateral hemisphere 48-72 h after X-irradiation. Later, alpha values were not significantly different from those in control animals. The decrease in lambda persisted at postnatal days 4-5. A significant increase in lambda and kappa' was found at postnatal days 18-21. We conclude that X-irradiation of the brain in the early postnatal period, even when it results in only relatively light damage, still produces changes in all three diffusion parameters, particularly a large increase in extracellular space volume fraction in all cortical layers, and in the subcortical white matter. Such changes in extracellular volume fraction of the brain can contribute to impairment of signal transmission, e.g. by diluting ions and neuroactive substances released from cells, and can play an important role in functional deficits, as well as in the impairment of developmental processes. Moreover, the increase in tortuosity (inferred from the decrease in apparent diffusion coefficient) in the X-irradiated cortex, as well as in the contralateral hemisphere, suggests that, even when extracellular volume is large, the diffusion of the substances is substantially hindered.

Animals↗
PubMed1996

Changes of extracellular space volume and tortuosity in the spinal cord of Lewis rats with experimental autoimmune encephalomyelitis.

Three diffusion parameters of nervous tissue, extracellular space (ECS) volume fraction (alpha), tortuosity (gamma) and non-specific uptake (k') of tetramethylammonium (TMA+), were studied in the spinal cord of rats during experimental autoimmune encephalomyelitis (EAE). The three parameters were determined in vivo from concentration-time profiles of TMA+ using ion-selective microelectrodes. EAE was induced by injection of guinea-pig myelin basic protein (MBP), which resulted in typical morphological changes in the CNS tissue, namely inflammatory reaction, astrogliosis, blood-brain barrier (BBB) damage and paralysis. EAE was accompanied by a statistically significant increase of alpha (mean +/- S.E.M.) in the dorsal horn from 0.21 +/- 0.01 to 0.28 +/- 0.02, in the intermediate region from 0.22 +/- 0.01 to 0.33 +/- 0.02, in the ventral horn from 0.23 +/- 0.01 to 0.47 +/- 0.02 and in white matter from 0.18 +/- 0.03 to 0.30 +/- 0.03. There were significant decreases in tortuosity in the dorsal horn and in the intermediate region and decreases in non-specific uptake in the intermediate region and in the ventral horn. Although the inflammatory reaction and the astrogliosis preceded and greatly outlasted the neurological symptoms, the BBB damage had a similar time course. Moreover, there was a close correlation between the changes in extracellular space diffusion parameters and the manifestation of neurological signs. We suggest that the expansion of the extracellular space alters the diffusion properties in the spinal cord. This may affect synaptic as well as non-synaptic transmission, intercellular communication and recovery from acute EAE, and may contribute to the manifestation of neurological signs in EAE rats.

Animals↗
PubMed1996

Frequent detection of reviraemia in ducks persistently infected with avian leukosis retroviruses.

Ducks intraembryonally infected with avian leukosis viruses of subgroup C (ALV-C) were followed for a long period (up to 6.8 years), and the viraemia and production of virus-neutralizing antibodies were measured. In three independent experiments comprising ducks inoculated with uncloned and/or molecularly cloned ALV-C, we found that after the elimination of primary post-hatching viraemia, reviraemia could be detected in 60-70% of infected animals. Based on the course of viraemia, the individual ducks were assigned to four different groups: Group I (no reviraemia), Group II (one transient reviraemic period), Group III (one persistent reviraemic period), Group IV (fluctuating reviraemia). In comparison to sera from ducks included in Group I and/or II, a significant decrease in neutralizing activity of sera from animals comprised in Group III and/or IV was observed. Two out of four reviraemic viruses were not neutralized by antiserum against ALV-C, instead their infectivity was enhanced. Long-term follow-up of the cell-associated virus revealed that its rescuability by cocultivation with chicken embryo fibroblasts fluctuated in about 50% of animals. In the reviraemic phase of infection, integrated proviruses could be detected by Southern blotting in a majority of tissues examined. Our data document that many features recognized in lentiviruses are valid also for oncoviruses transmitted to heterologous hosts and substantiate further the suitability of ALV-C-infected ducks as a model for studying persistent retroviral infection.

Animals↗