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Biomedical subjects

L László

Publications and source records attributed to L László.

At least 19 recordsLinked to original sources
PubMed2005

Increased incidence of genetic human prion disease in Hungary.

The authors performed analysis of the prion protein gene (PRNP) in 27 out of 109 confirmed prion disease patients between 1994 and 2004. E200K mutation was found in 17 cases. Another 10 patients, lacking PRNP analysis, showed positive family history. The mean annual incidence (0.27/million) and proportion (25.6%) of genetic prion disease is unusually high in Hungary and might be related to the migration of ancestors from the Slovakian focus.

Adult↗
PubMed2001

Cytotoxic effect of alcohol-withdrawal on primary cultures of cortical neurones.

Physical dependence on alcohol was observed previously at the cellular level in cultured IM-9 human lymphoblast cells. To answer the question whether physical dependence can also develop in neurones and to investigate the neuronal processes involved in the development of alcohol dependence and withdrawal symptoms, cultures of cortical neurones were adapted to alcohol. Morphological characteristics of neurones were not altered during the chronic (3-day) repeated (once per day) ethanol (50-100 mM) treatment, whereas obvious signs of neuronal damage were seen after the following 24 h of alcohol-withdrawal. The extent of the damage, quantitated by measuring the release of lactate dehydrogenase (LDH) into the culture media, was dependent on the concentration of ethanol in the medium during adaptation. LDH-release induced by alcohol-withdrawal was significantly reduced by re-addition of ethanol, as well as by administration of non-competitive (MK-801) or NR2B selective (threo-ifenprodil) N-methyl-D-aspartate (NMDA) receptor antagonists. The sigma ligand haloperidol and the L-type voltage sensitive calcium channel blocker nimodipine were also effective, whereas the effect of the gamma-aminobutyric acid type A (GABA(A)) receptor agonist muscimol was not significant. Furthermore, chronic ethanol treatment potentiated the NMDA induced neurotoxicity and the ability of acute alcohol to inhibit LDH-release in response to NMDA. According to these results, (i) the phenomenon of alcohol-dependence can be observed at the level of neurones and (ii) NMDA receptors seem to play a central role in the development of ethanol dependence and in neurotoxicity induced by alcohol-withdrawal.

Alcohol-Induced Disorders↗
PubMed2001

Prominent stress response of Purkinje cells in Creutzfeldt-Jakob disease.

To examine the role of stress-related 70-kDa heat shock proteins (Hsp-s) in Creutzfeldt-Jakob disease (CJD), we performed immunocytochemistry to detect Hsp-72 and Hsp-73, together with the abnormal (PrP(Sc)) and the presumed cellular form (PrP(C)) of the prion protein, and TUNEL method to measure cellular vulnerability in different brain regions in CJD and control cases. While Hsp-73 showed uniform distribution in all the examined samples, an increase in the number of Purkinje cells with prominent accumulation of Hsp-72 in the CJD group was observed. These neurons also showed intense PrP(C) staining, but TUNEL-positive nuclei were only detected in the granular (Hsp-72-negative) cell layer. Fewer cells of the inferior olivary nucleus were immunoreactive for Hsp-72 in CJD than in control cases, and regions showing severe spongiform change and gliosis exhibited fewer Hsp-72-immunoreactive neurons. Our results indicate that accumulation of the inducible Hsp-72 in certain cell types may be part of a cytoprotective mechanism, which includes preservation of proteins like PrP(C).

Adult↗
PubMed2001

Inherited prion disease with A117V mutation of the prion protein gene: a novel Hungarian family.

Three members of a family with inherited prion disease are reported. One additional family member had a progressive neurological disease without details. Two developed symptoms of ataxia, dementia, myoclonus, rigidity, and hemiparesis, and one had a different phenotype with the combination of lower motor neuron deficit, parkinsonism, intellectual decline, and ataxia. In this last patient cell loss of the anterior horn motor neurons and chronic neurogenic muscle atrophy was evident. Immunostaining for the prion protein disclosed unicentric and multicentric plaques, and coarse and fine granular positivity. Genetic analysis of the prion protein gene of the propositus showed a 117 codon alanine to valine mutation and homozygous 129 valine/valine genotype.

Adult↗
PubMed1999

Ultrastructural localization of Hsp-72 examined with a new polyclonal antibody raised against the truncated variable domain of the heat shock protein.

In spite of the intensive search for the determination of the continuously widening physiological and pathological roles of different stress proteins, their ultrastructural localization at the electron microscopic (EM) level has hardly been examined. As it becomes increasingly evident that the function and physiological effectiveness of stress proteins are highly dependent on their spatial location and their associations with diverse regulator proteins, the demand for morphological studies which can identify their detailed distribution within the cells is evident. The reason for the practical lack of studies carried out at the EM level, lies in the shortage of reagents with suitable specificity and avidity necessary for this type of examination. To create such a reagent, a polyclonal antibody was raised using a recombinant truncated form of the inducible Hsp-72 protein. The antibody was extensively characterized, using different immunochemical methods to determine and verify its specificity, and then it was tried in ultrastructural examinations. Using the new antibody, it was possible to analyze the intracellular distribution of Hsp-72 with the immunogold technique. The localization of Hsp-72 was demonstrated directly at the ultrastructural level in the cytoplasm (especially at the cisterns of the RER), in the nucleus (mainly around the heterochromatic regions) and at both sides of the nuclear envelope close to the membrane pores. Apart from these localizations, Hsp-72 was found in several membrane bordered intracellular structures, which mainly belong to the endosomal-lysosomal system. We provide the first morphological verification of the appearance of Hsp-72 on the surface of the cells. Also novel is the indication, that the stress protein may recycle from the cell surface using a common route which includes coated pits and the endosomal system.

3T3 Cells↗
PubMed1996

Time course of vinblastine-induced autophagocytosis and changes in the endoplasmic reticulum in murine pancreatic acinar cells: a morphometric and biochemical study.

The time course of the vinblastine(-sulfate; 10 mg/kg body weight, single injection)-induced enlargement and subsequent regression of the autolysosomal compartment was studied by electron microscopic morphometrical and cell biochemical methods in order to gain information concerning some key problems of this major route of intralysosomal degradation of the cell's endogenous macromolecules and structures. Detailed analysis of the dynamics of the total autophagic vacuole (AV) compartment and its different subcompartments (early, advanced, late, and fused AVs), as well as of changes of rough-surfaced endoplasmic reticulum (RER) showed: 1. Pancreatic acinar cells react to vinblastine biphasically, i.e. two expansion phases of the AV compartment, the first in the 0 to 90 min and the second in the 2 to 8 h post-injectional periods, were detected. 2. Fusions of AVs are not inhibited by vinblastine, at least during the second expansion phase when cytoplasmic volume fraction (CVF) of fused AVs steadily increased until the 12th h. Fusion of early, advanced and late AVs or composition of fused complex vacuoles (AVc) are somehow regulated, as the proportion of the three AV stages from the CVF of AVc, was maintained constant throughout the second expansion phase. 3. Stimulation of autophagosome formation and resulting substrate overload seems to be the primary mode of action by which vinblastine causes the enormous expansion of the autolysosomal compartment. 4. Degranulation of the rough-surfaced endoplasmic reticulum (RER) membranes occurs in a biphasic fashion, similarly to the volume and surface changes of the AV compartment, thus supporting our previous hypothesis, that labilization or change of RER may have a role in the formation of autophagosomes. 5. Vinblastine-induced autophagocytosis is a selective process, as mitochondria, Golgi elements and zymogen granules are very much underrepresented, whereas RER is more than twice overrepresented in the volume of early AVs, when compared to their volume fraction in the whole cytoplasm. 6. Immunogold electron microscopy revealed the presence of ubiquitinylated proteins in advanced and late, but not in early AVs.

Animals↗
PubMed1995

Related organelles of the endosome-lysosome system contain a different repertoire of ubiquitinated proteins in Sf9 insect cells.

Two components of the endosomal/lysosomal compartment of Sf9 cells, multivesicular bodies (MVB) and light vacuoles with membrane complexes (LVMC) have been isolated and probed for ubiquitin protein conjugates with a specific antibody. Immunogold electron microscopy indicates that whereas ubiquitin-protein conjugates are localised to electron dense areas of MVB they are associated with the membranes of LVMC. Five ubiquitinated polypeptides are revealed in MVB by immunoblotting while numerous ubiquitinated species forming a smear following electrophoresis are present in LVMC. We suggest two possible routes for entry of ubiquitin-protein conjugates into these organelles, via the cell surface and via primary lysosomes.

Animals↗
PubMed1993

Immunogold localisation of ubiquitin-protein conjugates in Sf9 insect cells. Implications for the biogenesis of lysosome-related organelles.

Immunogold electron microscopy with antibodies which primarily detect ubiquitin-protein conjugates shows conjugate-specific gold particles enriched severalfold in acid phosphatase-positive lysosomes and multivesicular bodies in insect Sf9 cells. The observations demonstrate that ubiquitinated proteins are associated with small acid phosphatase-containing primary lysosomes (transport vesicles) and indicate a pathway in which primary lysosomes fuse with multivesicular bodies to generate mature lysosome-related structures.

Acid Phosphatase↗
PubMed1992

2,5-Hexanedione-induced intermediate filament aggregates contain ubiquitin-protein conjugate immunoreactivity and resemble Rosenthal fibres.

A number of chronic degenerative disorders including cerebellar astrocytomas and Parkinson's disease are characterized by the presence of cytosolic inclusions which contain intermediate filament (IF) aggregates and ubiquitin-protein conjugate immunoreactivity. In cerebellar astrocytomas these inclusions are known as Rosenthal fibres. 2,5-hexanedione (HD) treatment is known to induce IF aggregates in cells in culture. HD-induced aggregates have therefore been studied as a potential model for the clinical inclusions. Exposure of astrocyte cultures to 2 mM HD for 2 or 4 weeks led to the formation of aggregates of the IFs (glial fibrillary acidic protein and vimentin). The aggregates contained ubiquitin-protein conjugates, which, on electron microscopy appeared to be localized in a peripheral shell. In addition, ubiquitin mRNA levels were found to be elevated approximately threefold by HD treatment. HD-induced inclusions and Rosenthal fibres were found to share a number of features. HD administration, therefore, appears to be a suitable model for the production of pathological inclusions.

Animals↗
PubMed1991

Ubiquitin in health and disease.

Studies in recent years have shown that ubiquitin has increasingly important functions in eukaryotic cells; roles which were previously not suspected in healthy and diseased cells. The interplay between molecular pathological and molecular cell biological findings has indicated that ubiquitin may be pivotal in the cell stress response in chronic degenerative and viral diseases. Furthermore, the studies have led to the notion that ubiquitination may not only serve as a signal for nonlysosomal protein degradation but may be a unifying covalent protein modification for the major intracellular protein catabolic systems; these can act to identify proteins for cytosolic proteinases or direct intact and fragmented proteins into the lysosome system for breakdown to amino acids. This unifying role could explain why ubiquitin is restricted to eukaryotic cells, which possess extensive endomembrane systems in addition to a nuclear envelope. Protein ubiquitination is a feature of most filamentous inclusions and certain other intracellular conglomerates that are found in some degenerative and viral diseases. The detection of ubiquitin-protein conjugates is not of great diagnostic importance in these diseases. Protein ubiquitination is not only essential for the normal physiological turnover of proteins but appears to have been adapted as part of an intracellular surveillance system that can be activated by altered, damaged, or foreign proteins and organelles. The purpose of this system is to isolate and eliminate these noxious structures from the cell: as a cytoprotective mechanism this appears to have evolved in the cell akin perhaps to an 'intracellular immune system'. Other heat shock proteins such as hsp 70 may be involved in this process. It is apparent that ubiquitin has a role in embryonic development. Protein ubiquitination is presumably involved in the reorganisation of cytoplasm that accompanies cell differentiation. Ubiquitin is also necessary for the gross intracellular degradative processes which are consequent upon programmed cell death. Cell elimination is of key importance for a number of developmental morphogenetic changes. An understanding of the molecular details of these processes will no doubt provide further insights into the wide ranging roles of ubiquitin in the life process. As it says in the book 'Ubiquitin'; there is no doubt that ubiquitin is a 'lucky' protein. It is lucky in many ways: lucky for scientific progress, lucky for biomedical scientists and lucky for life! If you have not already done so, why don't you get lucky and look for a role for ubiquitin in your experimental system. As Avram Hershko has said "there is plenty to go round"!

Animals↗
PubMed1991

Immunogold localisation of ubiquitin-protein conjugates in primary (azurophilic) granules of polymorphonuclear neutrophils.

Ubiquitin-protein conjugates are found in the primary (azurophilic) lysosome-related granules but not in the secondary (specific) granules in mature polymorphonuclear neutrophils prepared from bone marrow. This is the first reported demonstration of ubiquitin-protein conjugates in lysosome-related membrane-bound vesicles in granulocytes and complements our previous findings of ubiquitinated proteins in lysosomes of fibroblasts. The significance of the selective presence of conjugates in only one of the two main types of neutrophil granules remains to be elucidated but may relate to the presence of the complement of acid hydrolases, including proteases, in the azurophilic granules compared to the specific granules. Ubiquitin-protein conjugates may enter the primary granules during neutrophil maturation by an autophagic process or by a heterophagic process during the fusion of phagosomes with primary granules. Alternatively protein ubiquitination may be involved in granule biogenesis.

Animals↗
PubMed1991

The latent membrane protein-1 in Epstein-Barr virus-transformed lymphoblastoid cells is found with ubiquitin-protein conjugates and heat-shock protein 70 in lysosomes oriented around the microtubule organizing centre.

Immunofluorescence studies on Epstein-Barr virus (EBV)-transformed lymphoblastoid cells have previously shown that the latent membrane transforming protein (LMP-1) is found in patch-like inclusions which also immunostain for vimentin. We now show that EBV transformation causes a major reorganization of intermediate filaments, microtubules, mitochondria, and lysosomal elements, which generally become oriented around the microtubule organizing centre. Immunogold electron microscopy shows that LMP-1 is primarily concentrated in secondary lysosomes together with ubiquitin-protein conjugates and heat-shock protein 70. Intermediate filament inclusion formation with the above characteristics may be a general response triggered by other membrane glycoproteins; as seen, for example, in major human neurodegenerative diseases such as diffuse Lewy body disease.

Antigens, Viral↗
PubMed1991

Ubiquitin and the lysosome system: molecular immunopathology reveals the connection.

Considerable advances in our understanding of the molecular pathology of the major human neurodegenerative diseases have been made by the use of ubiquitin immunocytochemistry. The technique demonstrates that filamentous inclusions and vacuoles contain ubiquitin-protein conjugates. The molecular structure of the filaments and the morphological type of vacuoles is not completely understood but there is evidence that some of the filamentous inclusions contain intermediate filaments and the perinuclear distribution of the vacuoles resemble the distribution of intraneuronal lysosomes. Intermediate filaments and lysosomes are involved in the sequestration and degradation of viral membrane proteins in tissue culture cells. Immunogold electron microscopical and biochemical evidence indicates that ubiquitin-protein conjugates are normally considerably enriched in the lysosomes of fibroblasts relative to all other organelles. Immunogold electron microscopy shows a similar enrichment of ubiquitin-protein conjugates in the dense lysosomes of granulocytic precursor cells in the bone marrow. Filamentous inclusions showing several of the features seen in inclusions in the neurodegenerative diseases are seen in Epstein-Barr virus transformed lymphoblastoid cells. Immunohistochemistry shows that the inclusions contain vimentin intermediate filaments, the latent membrane transforming protein of the virus, ubiquitin-protein conjugates, and a heat-shock protein 70 (hsp 70). Immunohistochemistry and immunogold electron microscopy demonstrate that the latent membrane protein, ubiquitin-protein conjugates and hsp 70 are in lysosomes entwined in an intermediate filament cage centred on the microtubule organising centre. The implications of the combined observations for our understanding of the cell stress response in degenerating neurones and in virally infected cells are discussed.

Heat-Shock Proteins↗
PubMed1991

The role of protein ubiquitination in neurodegenerative disease.

Ubiquitin immunocytochemistry with an antiserum which reacts with ubiquitin-protein conjugates demonstrates the presence of ubiquitinated proteins in filamentous inclusions found in neurones in the major human neurodegenerative diseases, i.e. Alzheimer's disease, diffuse Lewy body disease, motor neurone disease. Ubiquitin immunohistochemistry has revolutionized the neuropathological diagnosis of dementia showing that diffuse Lewy body disease is not, as previously supposed, a rare cause of dementia. The filamentous inclusions in neurones in the human neurodegenerative diseases can be divided into at least two types based on recent immunocytochemical studies. We have shown that a ubiquitin-carboxyl terminal hydrolase is present in Lewy bodies but not in neurofibrillary tangles in Alzheimer's disease. This observation is significant since it indicates that molecular pathological mechanisms in neurones in diffuse Lewy body disease are fundamentally different to Alzheimer's disease. Ubiquitin-protein conjugates are also found in vacuoles in areas of granulovacuolar degeneration in hippocampal neurones in Alzheimer's disease and in granulovacuoles in neurones of scrapie infected mouse brain. These locations suggest that ubiquitinated protein are present in the lysosome-related system of neurones. We have recently shown that ubiquitin-protein conjugates are indeed enriched some 12-fold in the lysosomes of normal fibroblasts and lymphocytes.

Brain Diseases↗
PubMed1991

The degradative fate of ubiquitin-protein conjugates in nucleated and enucleated cells.

Covalent ligation of multiple copies of ubiquitin to proteins is known to target intracellular proteins for degradation by large molecular weight cytosolic proteinase(s). Ubiquitin protein conjugates are found in cytosolic cell compartments suggesting that ubiquitination may have multiple roles. We have detected ubiquitinated proteins in the lysosomal apparatus of normal fibroblasts and fibroblasts treated with lysosomal proteinase inhibitors. In contrast rabbit reticulocytes lack lysosomes. We present here direct evidence for ubiquitination of mitochondrial proteins during rabbit reticulocyte maturation. In addition ubiquitination appears to be associated with the terminal differentiation of human keratinocytes. These results suggest that: 1. ubiquitin-protein conjugates may be degraded lysosomally 2. organellar proteins may be degraded by the ubiquitin system 3. ubiquitination is involved in the programmed elimination of proteins and organelles from several cell types during differentiation.

3T3 Cells↗
PubMed1991

Translational inhibitors cycloheximide, emetine, and puromycin inhibit cellular autophagy in mouse liver parenchymal and pancreatic acinar cells in vivo.

The protein synthesis inhibitor cycloheximide is widely used (in vitro or in vivo) to inhibit the autophagic degradation of endogenous cellular proteins. Circumstantial evidence has been obtained largely from in vitro experiments for a similar effect of other translational inhibitors. In the present study, the in vivo effects of cycloheximide, emetine and puromycin on autophagy in murine exocrine pancreatic and liver cells were tested using electron microscopic morphometry. The experiments were based on the assumption that the autophagic compartment will regress if the formation of the vacuoles is blocked while degradation in the pre-existing vacuoles goes on. To make the measurements easier, autophagic compartment of the cells was greatly enlarged in both cell types by administering vinblastine (10 mg/kg b. wt.) for 2 h when the inhibitors were set on for an additional 30 min. During this half-an-hour, cycloheximide (0.2 mg/g b. wt.), emetine (0.12 mg/g b. wt.) and puromycin (0.2 mg/g b. wt.), respectively caused 58.5, 35.6, and 69.5% regression of the pancreatic and 46.7, 64.2, and 54.2% of the hepatocytic autophagic vacuole compartment. Thus, similarly to cycloheximide, both emetine and puromycin have proved to be inhibitors of autophagy in vivo. The results argue for a possible relationship between the synthesis and degradation of endogenous cellular proteins.

Animals↗
PubMed1991

Mechanism and dynamics of macroautophagy in murine exocrine pancreatic cells. A review of vinblastine-induced changes.

Autophagy is a major pathway of lysosomal degradation of cellular macromolecules. The paper summarizes the results obtained in the studies on macroautophagy using the exocrine pancreatic acinar cell as model system and vinblastine as inducer. Current knowledge about the origin and properties of the limiting membranes of autophagic vacuoles, and the results of quantitative morphological studies into the dynamics and kinetics of vinblastine-induced autophagocytosis, as well as recent achievements in isolation and characterization of subclasses of autophagic vacuoles (autophagosomes and autolysosomes) are reviewed.

Animals↗