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J Burda

Publications and source records attributed to J Burda.

At least 19 recordsLinked to original sources
PubMed2006

Calpain-induced proteolysis after transient global cerebral ischemia and ischemic tolerance in a rat model.

The activation of the [Ca(2+)]-dependent cysteine protease calpain plays an important role in ischemic injury. Here, the levels of two calpain-specific substrates, p35 protein and eukaryotic initiation factor 4G (eIF4G), as well as its physiological regulator calpastatin, were investigated in a rat model of transient global cerebral ischemia with or without ischemic tolerance (IT). Extracts of the cerebral cortex, whole hippocampus and hippocampal subregions after 30 min of ischemia and different reperfusion times (30 min and 4 h) were used. In rats without IT, the p35 levels slightly decreased after ischemia or reperfusion, whereas the levels of p25 (the truncated form of p35) were much higher than those in sham control rats after ischemia and remained elevated during reperfusion. The eIF4G levels deeply diminished after reperfusion and the decrease was significantly greater in CA1 and the rest of the hippocampus than in the cortex. By contrast, the calpastatin levels did not significantly decrease during ischemia or early reperfusion, but were upregulated after 4 h of reperfusion in the cortex. Although IT did not promote significant changes in p35 and p25 levels, it induced a slight increase in calpastatin and eIF4G levels in the hippocampal subregions after 4 h of reperfusion.

Animals↗
PubMed2006

The effects of normovolemic hemodilution on protein synthesis recovery following postischemic reperfusion in the rat brain.

Normovolemic hemodilution is a possible way to improve the brain recovery after ischemia and reperfusion. Therefore we have decided to examine how this process may affect the post-ischemic protein synthesis machinery. We analysed rat brains after 4-vessel-occlusion and different time intervals of reperfusion using normovolemic hemodilution. We achieved an important increase of [4,5-3H]leucine incorporation into polypeptides in vitro in the rat brain neocortex 30 minutes after ischemia, but concurrently there was no significant change in the hippocampus and striatum. By extending the time course of reperfusion we did not observe any important deviation of in vitro [4,5-3H]leucine incorporation in the studied brain areas. Thus, although hemodilution increased protein synthesis in selective vulnerable regions after ischemia, this improvement is not of significant importance.

Animals↗
PubMed2004

Does phosphorylation of eukaryotic elongation factor eEF2 regulate protein synthesis in ischemic preconditioning?

Ischemia/reperfusion-associated translation inhibition in the hippocampus is attenuated significantly at reinitiation and elongation steps by ischemic preconditioning (Burda et al. [2003] Neurochem. Res. 28:1237-1243). To address potential regulation of the elongation step by changes in eukaryotic elongation factor 2 (eEF2) phosphorylation with and without acquired ischemic tolerance (IT), Wistar rats were preconditioned by 5-min sublethal ischemia and 2 days later, 30-min lethal ischemia was induced. Given the important role that oxidative stress plays in the ischemic process, eEF2 phosphorylation was also studied in a model of oxidative stress in vitro. Three blocks of our results support a lack of correlation between eEF2 phosphorylation status and protein synthesis rate. First, eEF2 was dephosphorylated significantly (activated) after transient cerebral ischemia in rats with and without IT or H2O2-treated cells; however, protein synthesis was significantly inhibited under these three conditions. Second, after 30-min reperfusion, the protein synthesis rate was maintained below control levels in cortex and hippocampus of rats without IT. Eukaryotic EF2 phosphorylated levels were notably low only in the cortex, whereas levels in the hippocampus were close to that of sham controls. In rats with IT, protein synthesis was virtually restored in both brain regions, but phosphorylated eEF2 levels were even higher than in rats without IT. Third, after 4-hr reperfusion, the protein synthesis rate in cortex and hippocampus was observed to be below sham control values in rats with and without IT. Conversely, phosphorylated eEF2 levels were below sham control in rats with IT and reached sham control values in rats without IT.

Animals↗
PubMed2004

The effect of preconditioning on the iron deposition after transient forebrain ischemia in rat brain.

In this study we investigated iron deposition in the hippocampus CA1 area and the corpus striatum pars dorsolateralis in a rat model of cerebral ischemia and ischemic tolerance. Forebrain ischemia was induced by four-vessel occlusion for 5-min as ischemic preconditioning. Two days after the preconditioning or the sham operation, a second ischemia was induced for 20-min. With the use of iron histochemistry, regional changes were examined after 2 to 8 weeks of recirculation following the 20-min ischemia with or without preconditioning. Perl's reaction with DAB intensification demonstrated iron deposits in the CA1 area and in the corpus striatum pars dorsolateralis after 2 weeks of recirculation. These iron deposits gradually increased in density and formed clusters by the 8th week. When the rats were exposed to 5-min ischemia 2 days before lethal 20-min ischemia, the deposition of iron in the CA1 region of the hippocampus and also in the corpus striatum pars dorsolateralis was decreased and produced a minimal number of iron-containing cells between the second and the 8th week of recirculation. Preconditioning with sublethal 5-min ischemia followed by 2 days of reperfusion also prevented the neuronal destruction of the hippocampal CA1 region induced by 20-min ischemia.

Animals↗
PubMed2004

The protective effect of aminoguanidine on cerebral ischemic damage in the rat brain.

The NADPH-diaphorase (NADPH-d) histochemical technique is commonly used to localize the nitric oxide (NO) produced by the enzyme nitric oxide synthase (NOS) in neural tissue. The expression of inducible nitric oxide synthase (iNOS) is induced in the late stage of cerebral ischemia, and NO produced by iNOS contributes to the delay in recovery from brain neuronal damage. The present study was performed to investigate whether the increase in nitric oxide production via inducible nitric oxide synthase was suppressed by the administration of aminoguanidine, a selective iNOS inhibitor, as it follows a decrease of NADPH-diaphorase activity (a marker for NOS) after four-vessel occlusion used as an ischemic model. The administration of aminoguanidine (100 mg/kg i.p., twice per day up to 3 days immediately after the ischemic insult) reduced the number of NADPH-diaphorase positive cells to control levels. Our results indicated that aminoguanidine suppressed NADPH-diaphorase activity, and also decreased the number of NADPH-diaphorase positive cells in the CA1 region of the hippocampus following ischemic brain injury.

Animals↗
PubMed2004

Effect of Tanakan on postischemic activity of protein synthesis machinery in the rat brain.

A growing body of evidence supports the role of free radicals in triggering the functional and metabolic disturbances following transient cerebral ischemia. This study was designed to evaluate whether the extent of reperfusion-induced inhibition of protein synthesis initiation as well as tissue injury can be reduced by Tanakan (Ginkgo biloba extract, EGb 761) (Beaufour-Ipsen Industrie). Rats received Tanakan in the dose of 40 mg/kg/day for 7 days before surgical intervention. Transient forebrain ischemia was induced by 4-vessel occlusion. Rats were subjected to 20 min of ischemia followed by 30 min, 4 h or 7 days of reperfusion. Protein synthesis rate, reinitiation ability and neurodegeneration in the frontal cortex and hippocampus were measured by the incorporation of radioactively labelled leucine into polypeptide chains in postmitochondrial supernatants and by Fluoro-Jade B staining. The protective effect was observed, concerning both the protein synthesis and the number of surviving neurons, in the Tanakan-treated groups. Tanakan significantly reduced the ischemia/reperfusion-induced inhibition of translation in the neocortex as well as in the highly sensitive hippocampus. Our results indicate that free radicals play an important role in the development of reperfusion-induced injury, and the treatment of ischemic and reperfused brain with free radical scavengers may reduce the severity of reperfusion damage.

Animals↗
PubMed2001

Possible mechanisms involved in the down-regulation of translation during transient global ischaemia in the rat brain.

The striking correlation between neuronal vulnerability and down-regulation of translation suggests that this cellular process plays a critical part in the cascade of pathogenetic events leading to ischaemic cell death. There is compelling evidence supporting the idea that inhibition of translation is exerted at the polypeptide chain initiation step, and the present study explores the possible mechanism/s implicated. Incomplete forebrain ischaemia (30 min) was induced in rats by using the four-vessel occlusion model. Eukaryotic initiation factor (eIF)2, eIF4E and eIF4E-binding protein (4E-BP1) phosphorylation levels, eIF4F complex formation, as well as eIF2B and ribosomal protein S6 kinase (p70(S6K)) activities, were determined in different subcellular fractions from the cortex and the hippocampus [the CA1-subfield and the remaining hippocampus (RH)], at several post-ischaemic times. Increased phosphorylation of the alpha subunit of eIF2 (eIF2 alpha) and eIF2B inhibition paralleled the inhibition of translation in the hippocampus, but they normalized to control values, including the CA1-subfield, after 4--6 h of reperfusion. eIF4E and 4E-BP1 were significantly dephosphorylated during ischaemia and total eIF4E levels decreased during reperfusion both in the cortex and hippocampus, with values normalizing after 4 h of reperfusion only in the cortex. Conversely, p70(S6K) activity, which was inhibited in both regions during ischaemia, recovered to control values earlier in the hippocampus than in the cortex. eIF4F complex formation diminished both in the cortex and the hippocampus during ischaemia and reperfusion, and it was lower in the CA1-subfield than in the RH, roughly paralleling the observed decrease in eIF4E and eIF4G levels. Our findings are consistent with a potential role for eIF4E, 4E-BP1 and eIF4G in the down-regulation of translation during ischaemia. eIF2 alpha, eIF2B, eIF4G and p70(S6K) are positively implicated in the translational inhibition induced at early reperfusion, whereas eIF4F complex formation is likely to contribute to the persistent inhibition of translation observed at longer reperfusion times.

Animals↗
PubMed2001

Ischemia-induced inhibition of the initiation factor 2alpha phosphatase activity in the rat brain.

Rats were subjected to the standard four-vessel occlusion model of brain transient ischemia for 30 min. Following different recirculation periods, the level of phosphorylation of the initiation factor 2 subunit alpha (eIF2alpha) and the eIF2alpha kinase/s and phosphatase/s activity were determined. eIF2alpha phosphorylation significantly increased very early during reperfusion (10-30 min), recovering at 4 h of reperfusion. Activation of any eIF2alpha kinases studied during ischemia or reperfusion was not noted. Conversely, eIF2alpha phosphatase activity significantly decreased at 10-15 min of reperfusion, reaching values even higher than in controls at 2-4 h of reperfusion. Our results support the hypothesis that the reperfusion-induced phosphorylated eIF2alpha changes are at least a result of the transiently eIF2alpha phosphatase inhibition.

Animals↗
PubMed1998

The intraischemic and early reperfusion changes of protein synthesis in the rat brain. eIF-2 alpha kinase activity and role of initiation factors eIF-2 alpha and eIF-4E.

Rats were subjected to the standard four-vessel occlusion model of transient cerebral ischemia (vertebral and carotid arteries). The effects of normothermic ischemia (37 degrees C) followed or not by 30-minute reperfusion, as well as 30-minute postdecapitative ischemia, on translational rates were examined. Protein synthesis rate, as measured in a cell-free system, was significantly inhibited in ischemic rats, and the extent of inhibition strongly depended on duration and temperature, and less on the model of ischemia used. The ability of reinitiation in vitro (by using aurintricarboxylic acid) decreased after ischemia, suggesting a failure in the synthetic machinery at the initiation level. Eukaryotic initiation factor 2 (eIF-2) presented almost basal activity and levels after 30-minute normothermic ischemia, and the amount of phosphorylated eIF-2 alpha in these samples, as well as in sham-control samples, was undetectable. The decrease in the levels of phosphorylated initiation factor 4E (eIF-4E) after 30-minute ischemia (from 32% to 16%) could explain, at least partially, the impairment of initiation during transient cerebral ischemia. After reperfusion, eIF-4E phosphorylation was almost completely restored to basal levels (29%), whereas the level of phosphorylated eIF-2 alpha was higher (13%) than in controls and ischemic samples (both less than 2%). eIF-2 alpha kinase activity in vitro as measured by phosphorylation of endogenous eIF-2 in the presence of ATP/Mg2+, was higher in ischemic samples (8%) than in controls (4%). It seems probable that the failure of the kinase in phosphorylating eIF-2 in vivo during ischemia is due to the depletion of ATP stores. The levels of the double-stranded activated eIF-2 alpha kinase were slightly higher in ischemic animals than in controls. Our results suggest that the modulation of eIF-4E phosphorylation could be implicated in the regulation of translation during ischemia. On the contrary, phosphorylation of eIF-2 alpha, by an eIF-2 alpha kinase already activated during ischemia, represents a plausible mechanism for explaining the inhibition of translation during reperfusion.

Animals↗
PubMed1996

Recombinant adenovirus deleted of all viral genes for gene therapy of cystic fibrosis.

Recombinant adenoviruses are being developed for gene therapy of inherited disorders such as cystic fibrosis because they efficiently transduce recombinant genes into nondividing cells in vivo. First generation recombinant adenoviruses, rendered defective by deletion of sequences spanning E1a and E1b, express low levels of early and late viral genes that activate destructive cellular immune responses. Current strategies for improving recombinant adenoviruses attempt to inactivate other essential genes through deletion and growth in new packaging cell lines or incorporation of temperature sensitive mutations which allow propagation of the virus in available packaging cell lines at permissive temperatures. We describe in this report a new type of recombinant adenovirus that is deleted of all viral open reading frames. This recombinant (called delta-rAd), which contains only the essential cis elements (i.e., ITRs and contiguous packaging sequence), is propagated in 293 cells in the presence of E1-deleted helper virus. Concatamers of the monomeric vector genome were passaged and capable of transduction. The delta-rAd genome is packaged into virions that sediment at a lower density than the helper virus in cesium gradients forming the basis for a purification protocol. A fully deleted recombinant adenovirus that expresses human cystic fibrosis transmembrane conductance regulator was produced and used to transduce human airway epithelial cells derived from a cystic fibrosis patient. Packaging and propagation of a fully deleted adenovirus is an important step toward the development of a safer vector. Improved production and purification strategies need to be developed before this new vector system can be evaluated in vivo.

Adenoviridae↗
PubMed1994

Phosphorylation of the alpha subunit of initiation factor 2 correlates with the inhibition of translation following transient cerebral ischaemia in the rat.

Rats were subjected to the standard four-vessel occlusion model of cerebral transient ischaemia (vertebral and carotid arteries) for 15 and 30 min. After a 30 min recirculation period, protein synthesis rate, initiation factor 2 (eIF-2) and guanine nucleotide exchange factor (GEF) activities, and the level of phosphorylation of the alpha subunit of eIF-2 (eIF-2 alpha) were determined in the neocortex region of the brain from sham-operated controls and ischaemic animals. Following reversible cerebral ischaemia, the protein synthesis rate, as measured in a cell-free system, was significantly inhibited (70%) in the ischaemic animals. eIF-2 activity, as measured by its ability to form a ternary complex, also decrease parallel to the decrease in protein synthesis. As eIF-2 activity was assayed in the presence of Mg2+ and GTP-regenerating capacity, the decrease in ternary-complex formation indicated the possible impairment of GEF activity. Since phosphorylated eIF-2 [eIF-2(alpha P)] is a powerful inhibitor of GEF, the levels of phosphorylated eIF-2 alpha were determined, and an increase from 7% phosphorylation in sham control rats to 20% phosphorylation in 15 min and 29% phosphorylation in 30 min in ischaemic rats was observed, providing evidence for a tight correlation of phosphorylation of eIF-2 alpha and inhibition of protein synthesis. Moreover, GEF activity measured in the GDP-exchange assay was in fact inhibited in the ischaemic animals, proving that protein synthesis is impaired by the presence of eIF-2(alpha P), which blocks eIF-2 recycling.

Animals↗
PubMed1994

Ischemia-induced delayed-onset paraplegia is accompanied by an unusual form of synaptic degeneration in the lumbosacral segments: an experimental light and electron microscopic study in dogs.

We studied the effect of high thoracic aorta cross-clamping, complete transverse section of the spinal cord at Th6 level, and combined hemisection at Th6 level followed later by high thoracic aorta cross-clamping upon the morphology and number of identified presynaptic knobs in lumbosacral segments in dogs. In animals surviving 48-72 hours after high thoracic aorta cross-clamping the occurrence of an unusual form of boutons accompanied by periboutonal halo in L3-S1 segments was found. According to the bouton size and light as well as electron microscopic appearance, four types, i.e., light giant (T1), dark enlarged (T2), light giant with periboutonal halo (T3), and giant disintegrating (T4) boutons were detected after 48 and 72 hour reperfusion. The appearance of four boutonal types in the lumbosacral segments is caused by spinal cord ischemia secondary to high thoracic aorta cross-clamping followed by 48 or 72 hour reperfusion. At the end of the sixth reperfusion day no signs of enlarged and giant boutons were detected in L3-S1 segments. A statistically significant increase of enlarged and giant boutons was noted at the end of the third reperfusion day in comparison with 48 hour survival. After spinal cord transection at midthoracic (Th6) level, followed by 72 hour survival, no such unusual synaptic knobs could be found in L3-S1 segments. The laminar distribution pattern of T1-T4 types based on light microscopic analysis and confirmed electron microscopically is characteristic and strictly bound to those spinal cord gray matter layers which serve as main termination sites of the descending cortical, brain stem, as well as long propriospinal projections in the lumbosacral segments (laminae V-VII). A statistically significant increase of enlarged and giant boutons was found in the intermediate zone (lamina VII). Hemisection at midthoracic level (Th6) followed later by 30 minute high thoracic aorta cross-clamping and 48 hour reperfusion caused a marked decrease of enlarged and giant boutons in L3-S1 segments on the hemisectioned side in comparison with the contralateral one. Large amounts of irregularly arranged round vesicles and tubular profiles were disclosed in the boutonal matrix of T1, T3, and T4 types in L3-S1 segments of animals subjected to 30 minute high thoracic aorta cross-clamping followed by 72 hour reperfusion. Accumulation of tubular and membranous materials was invariably seen in the bulbous enlargement of the terminal axonal branch.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Concentrations of lead, cadmium, mercury and copper in mushrooms in the vicinity of a lead smelter.

The concentrations of four heavy metals in 149 samples of mushroom fruiting bodies, representing 11 species, mainly all edible, were determined by atomic absorption spectroscopy. The mushrooms were collected up to a distance of 6 km from a lead smelter in central Bohemia (Czechoslovakia) in operation since 1786. Lead was accumulated extensively by Lepiota rhacodes and Lepista nuda. Among other species, significant accumulation was found up to a distance of 1 km from the source. Concentrations of greater than 100 mg kg-1 dry matter were often determined. The safe limit of 5 mg kg-1 dry matter was exceeded in most samples collected at distances of up to 6 km from the source. Concentrations of cadmium in the polluted area were generally significantly higher than in other parts of Bohemia. Cadmium was extensively accumulated by the toxic Amanita muscaria, but also by the edible Boletus edulis and Amanita rubescens, with mean values 28.6, 15.2 and 12.3 mg Cd kg-1 dry matter, respectively. The Czechoslovakian statutory limit is 0.5 mg Cd kg-1 dry matter. Statistically significant linear correlations between lead and cadmium concentrations were found only for Boletus edulis and Paxillus involutus. Mercury was accumulated by Lepista nuda and Lepiota rhacodes; mean values of 11.9 and 6.5 mg Hg kg-1 dry matter, respectively, were found. Concentrations of mercury in most species from the study area were higher than in those from other parts of Bohemia. Lepiota rhacodes and Lepista nuda also accumulated copper extensively with mean values of 280 and 193 mg Cu kg-1 dry matter, respectively.

Basidiomycota↗
PubMed1991

Graded postischemic reoxygenation ameliorates inhibition of cerebral cortical protein synthesis in dogs.

The purpose of this study was to determine the effect of normoxic reperfusion and graded postischemic reoxygenation on cerebral protein synthesis in a cell-free system. Ischemia alone produced a relatively small decrease (15-17%) in activity in all the subcellular systems studied. After a 15-min interval of normoxic reperfusion (75-90 mmHg O2 in arterial blood), a 40% decrease (p less than 0.01) in [14C]leucine incorporation was observed. Reoxygenation with hypoxemic blood containing 37.5 mm Hg O2 at 0-5 min and 56 mm Hg O2 at 6-10 min of recirculation followed by 5 min of normoxic reperfusion resulted in a significant increase (p less than 0.05) of polypeptide chain synthesis in vitro when compared with normoxic reperfusion. The results obtained by this experimental approach tend to show that graded postischemic reoxygenation could be used as a simple and effective neuroprotective tool that substantially diminishes the secondary postischemic damage in nervous tissue, including the newly synthesized proteins.

Animals↗
PubMed1991

Mechanism of protein synthesis inhibition in CNS during postischaemic reperfusion.

We have used brain (dog, rat) and spinal cord (dog, rabbit) cell-free systems to study early postischaemic inhibition of protein synthesis. Ischaemia alone produced a relatively small decrease in activity of all subcellular systems used. When 15 min of normoxic reperfusion was used, more than 30% decrease (p less than 0.01) in [14C]-leucine incorporation was detected. A translational inhibitor that appeared in the postribosomal supernatant fraction at the early stage of reperfusion reduced translational capacity of an initiating cell-free system. It also phosphorylated the small (38 kDa) subunit of eukaryotic initiation factor 2 (eIF-2) in vitro. Effect of the inhibitor can be reversed by addition of partially purified intact eIF-2 and/or high concentration (2 mmol/l) of GTP. A prevention of postischaemic free oxygen radical formation by the reoxygenation with hypoxaemic blood, containing 37.5 mm Hg O2 at 0-5 min and 56 mm Hg O2 at 6-10 min of recirculation, that was followed by 5 min of normoxic reperfusion, resulted in a significant increase (p less than 0.02) of polypeptide chain synthesis in vitro when compared with normoxic reperfusion.

Animals↗
PubMed1991

Effect of ischaemia on protein synthesis in neuron and glia-enriched fractions from the rabbit spinal cord.

The incorporation of 14C-leucine into the post-mitochondrial supernatant and neuron, glia and myelin-enriched fractions isolated from the rabbit spinal cord was studied after ischaemia and subsequent recirculation. In the cell-free system, incorporation decreased to 55% of the control value after 40 min ischaemia, but proteosynthesis returned to the pre-ischaemic value after 3 h recirculation and remained at this level during further recirculation. The incorporation of amino acids into proteins of neurons and neuroglia differed from the cell-free system and from each other. In the enriched neuronal fraction, protein synthesis fall after ischaemia and also during the first hours of recirculation, but during further recirculation it rose to 60% above the control value. In the enriched glial fraction, specific radioactivity of proteins rose abruptly immediately after ischaemia and by the fourth day there was sixfold increase as compared with control values. The results indicate that the ischaemia-induced decrease in protein synthesis is only transient and that a significant increase occurs in the surviving cell populations, especially the neuroglia. The functional changes caused by spinal cord ischaemia are irreversible, however.

Animals↗