Biomedical subjects

G Lubec

Publications and source records attributed to G Lubec.

At least 73 records · Page 4Linked to original sources

PubMed2001

Deficient brain snRNP70K in patients with Down syndrome.

The small nuclear ribonucleoprotein 70K (snRNP 70K; U1-70 kDa) is an integral part of the spliceosome, a large RNA-protein complex catalyzing the removal of introns from nuclear pre-mRNA. snRNP is one of the best-studied essential subunits of snRNPs, is highly conserved and its inactivation was shown to result in complete inhibition of splicing. Applying subtractive hybridization, we found a sequence with 100% identity to snRNP absent in fetal Down syndrome (DS) brain. This observation made us determine snRNP-mRNA steady-state levels and protein levels in brains of adult patients with DS. snRNP-mRNA and protein levels of five individual brain regions of DS and controls each, were determined by blotting techniques. snRNP-mRNA steady state levels were significantly decreased in DS brain. Performing Western blots with monoclonal and human antibodies, snRNP protein levels were decreased in several regions of DS brain, although one monoclonal antibody did not reveal different snRNP-immunoreactivity. Although decreased snRNP-protein could be explained by decreased mRNA-steady state levels, another underlying mechanism might be suggested: snRNP is one of the death substrates rapidly cleaved during apoptosis by interleukin-1-beta-converting enzyme-like (ICE) proteases, which was well-documented by several groups. As apoptosis is unrequivocally taking place in DS brain leading to permanent cell loses, decreased snRNP-protein levels may therefore reflect decreased synthesis and increased apoptosis-related proteolytic cleavage.

Adult↗

PubMed2001

Evidence for the relation of herpes simplex virus type 1 to Down syndrome and Alzheimer's disease.

The peripheral and central nervous system are harbouring herpes simplex virus type 1 (HSV-1) and this virus has been proposed to be implicated in the aetiology of Alzheimer's disease (AD). We tested whether the HSV-1 genome is found indeed in the brain of controls, patients with AD and Down syndrome (DS) and whether HSV-1 infectious proteins in brain were induced. Moreover, we tested whether interleukin (IL)-6, a marker for neuroinflammation, is found in brains of AD and DS. HSV-1 glycoprotein D gene, as well as viral phosphoprotein and glycoprotein were detected in all brain samples. IL-6 was detectable in seven out of the eight AD and all of the eight DS patients, but only three out of ten controls in the frontal cortex. IL-6 in cerebellum was detectable in all AD and DS patients, but only three out of nine controls. In conclusion, we propose that the detection of HSV-1 genome and HSV-1 inducible protein IL-6 not only shows the presence in human brain, but may indicate a role for HSV-1 in the process of neuroinflammation and apoptosis, known to occur in both neurodegenerative disorders, AD and DS.

Alzheimer Disease↗

PubMed2001

Changes in the brain protein levels following administration of kainic acid.

Kainic acid (KA), a potent neurotoxin and excitatory amino acid, leads to derangements and modulation of brain proteins. No global brain protein expression pattern induced by KA-treatment has been reported yet. We therefore studied the effect of systemic KA administration on the levels of brain proteins. Rats were injected placebo or KA intraperitoneally and brain was taken after one week. The mitochondrial and cytosolic fractions of the brain proteins were analyzed by proteomics technologies and the levels of selected proteins were quantified using specific software. Heat shock protein HSP 27 was exclusively detected in brains of animals treated with KA, whereas the glucose regulated protein GRP 78 was downregulated. The levels of neurofilaments and alpha-internexin were significantly decreased and a fragment of tubulin alpha-1 chain was manifold increased in KA-brains. The mitochondrial enzymes dihydrolipoamide dehydrogenase, ATP synthase beta chain and isocitrate dehydrogenase were reduced and pyruvate kinase M1 was increased following KA treatment. We conclude that the concomitant determination of the brain proteins indicates altered regulation of heat shock proteins, neuronal death, cytoskeletal disruption, and mitochondrial derangement by systemic KA administration. This report confirms and extends previous studies on the effect of KA on the expression of brain proteins and suggests that our analytical system can serve as a model for neurotoxicological, neurobiological, and neuropathological proteome studies.

Animals↗

PubMed2001

Postmortem changes in the level of brain proteins.

A systematic study on postmortem changes of brain proteins has not been performed so far and information is limited to basic principles of specific or nonspecific proteolysis or proteolysis of individual proteins. We studied protein level alterations in rat brain of animals kept at 23 degrees C for several postmortem times up to 72 h. Brain tissue protein extracts were analyzed by two-dimensional electrophoresis and the proteins with different levels were identified by matrix-assisted laser desorption ionization mass spectrometry. The changes observed mainly concerned structural proteins and enzymes. The levels of dihydropyrimidinase-related protein-2 decreased within 6 h and two new spots were detected representing shorter forms of the protein. Most of the other alterations appeared about 48 h postmortem. The most significant were reduced levels of neurofilament, alpha-internexin, synaptosomal-associated protein 25, glial fibrillary acidic protein, heat shock proteins, and dynamin-1; increased levels of 14-3-3 proteins and spectrin; and generation of shorter forms of certain proteins, such as tubulins, actin, and serum albumin. The results may be useful in neuropathology and brain protein studies.

Animals↗

PubMed2001

Decreased alpha-endosulfine, an endogenous regulator of ATP-sensitive potassium channels, in brains from adult Down syndrome patients.

Alpha-endosulfine has the ability to block ATP-sensitive potassium (K(ATP)) channels and stimulate insulin release in beta cells like sulfonylurea. Alpha-endosulfine is expressed in a wide range of tissue, including brain and endocrine tissues. Although K(ATP) channels are also present in brain and its regulators have been reported to be involved in the release of neurotransmitters such as acetylcholine that plays an important role in cognitive function, the neurobiological role of alpha-endosulfine has not been studied yet. We examined the expression levels of alpha-endosulfine protein in frontal cortex and cerebellum from patients with Down syndrome (DS) showing Alzheimer's disease (AD) pathology using Western blotting. In frontal cortex, alpha-endosulfine was detected in all of 10 controls, but only 1 (from female) out of 8 DS with weak density. In cerebellum, alpha-endosulfine was also detected in all of 9 controls, but only 1 (from male) out of 6 DS with weak density. The considerably decreased alpha-endosulfine could result in the continuous opening of K(ATP) channels and the subsequent decrease of neurotransmitters release associated with cognition. This study is of significance providing evidence for a biological role of alpha-endosulfine in brain and alpha-endosulfine protein could be a pharmacological target for therapeutic intervention.

Aged↗

PubMed2001

Selective upregulation of the ubiquitin-proteasome proteolytic pathway proteins, proteasome zeta chain and isopeptidase T in fetal Down syndrome.

The ubiquitin-proteasome proteolytic pathway is involved in an important non-lysosomal proteolytic pathway that is responsible for the highly selective turnover of cellular proteins both under basal metabolic conditions as well as stress. Protein degradation by this pathway is attributed to the 20S proteasome that forms the catalytic core of the complex. Recently there has been increasing interest in the proteasome because of its possible role in neuron degeneration and death. Fetal Down syndrome (DS) neurons were demonstrated to degenerate and undergo apoptosis in vitro. We therefore investigated the expression of different proteins involved in this degradative pathway, including subunits of the 20S proteasome, ubiquitinating and deubiquitinating enzymes, and regulatory subunits of the 26S proteasome in control and DS fetal brains by two-dimensional electrophoresis (2-DE). After 2-DE, approximately 389 protein spots were successfully identified by matrix-associated laser desorption ionization mass spectroscopy (MALDI-MS) and this was followed by quantification of twenty three proteins of the pathway. The results indicate that all but two proteins exhibited no apparent alterations in their pattern of expression. Proteasome zeta chain, an alpha subunit of the 20S proteasome (P < 0.05) and ubiquitin carboxy-terminal hydrolase T (Isopeptidase T), a deubiquitinating enzyme (P < 0.001) were significantly increased in fetal DS compared to controls. Whilst the expression of proteasome iota (n = 9, r = -0.9489, P = 0.0004) and proteasome epsilon (n = 9, r = -0.7227, P = 0.0311) chains was decreased with age in fetal DS brain, no significant correlation was obtained in the other proteins with age. The data suggest that such selective upregulation may have relevance to the developmental abnormalities that characterize this disorder.

Brain↗

PubMed2001

Unaltered expression of Fas (CD95/APO-1), caspase-3, Bcl-2 and annexins in brains of fetal Down syndrome: evidence against increased apoptosis.

Apoptosis is the mechanism by which cells are programmed to die under a wide range of physiological and developmental stimuli. Accumulating evidence indicates that enhanced apoptosis (programmed cell death) in Down syndrome (DS) may play a role in mental retardation and precocious neurodegeneration of the Alzheimer-type. In this regard, alteration of several apoptosis related proteins have been reported in adult DS brain. Fetal DS neurons exhibited increased reactive oxygen species leading to early apoptosis, however, expression of apoptosis related proteins in fetal DS, has never been considered. To address this issue, we investigated the expression of proteins involved in apoptosis including Fas (CD95, APO-1), caspase-3, Bcl-2 and annexins in the cerebral cortex of control and DS fetal brain by western blot and two dimensional electrophoresis. Here, we report that no detectable changes were obtained in fetal DS brain in the expression of Fas, caspase-3, Bcl-2 and Annexins (I, II, V, and VI) compared to controls. In parallel experiment, we also examined the expression of neuron specific enolase (NSE), a neuronal marker found to be decreased in adult DS brain, to see if there is any neuronal loss and no difference was observed between the two groups. Protein expression did not correlate with age. The unchanged levels of Fas, Bcl-2 and annexins together with unaltered caspase-3 expression, a predominant caspase that executes apoptosis in the developing nervous system, suggest that enhanced apoptosis may not be apparent in fetal DS brain as demonstrated for adult DS brain.

Annexins↗

PubMed2001

Alteration of caspases and other apoptosis regulatory proteins in Down syndrome.

Apoptosis has been implicated in the selective neuronal loss of Down syndrome (DS). Apoptosis activates a family of cysteine proteases with specificity for aspartic acid residues referred to as caspases that play a key role in dismantling a cell committed to die. Caspase activity is regulated by a variety of proteins that possess a domain resembling the prodomains of caspases. Little is known, however, about the changes of caspases and their regulatory proteins in DS. Here, we investigated levels of nine such different proteins by western blot technique in frontal cortex and cerebellum of control and DS subjects. The protein levels of DFF45 (DNA fragmentation factor 45), and FLIP (FADD like interleukin-1beta-converting enzyme inhibitory proteins) were significantly decreased whereas that of RICK (RIP-like interacting CLARP kinase) increased in both regions of DS. In contrast, cytochrome c, Apaf-1 (apoptosis protease activating factor-1), procaspase-9 and ARC (apoptosis repressor with caspase recruitment domain) were unchanged. Procaspase-3 and -8 were significantly decreased in frontal cortex but no significant change was observed in cerebellum. Regression analysis revealed no correlation between postmortem interval and levels of the investigated proteins. However, inconsistent correlation was found between age and levels of proteins as well as amongst the density of individual proteins. These findings demonstrate that dysregulation of apoptotic proteins does exist in DS brain and may underlie the neuropathology of DS. The study further suggests that apoptosis in DS may occur via the death receptor pathway independent of cytochrome c. Hence, therapeutic strategies that target caspase activation may prove useful in combating neuronal loss in this disorder.

Aged↗

PubMed2001

Expression of apoptosis related proteins: RAIDD, ZIP kinase, Bim/BOD, p21, Bax, Bcl-2 and NF-kappaB in brains of patients with Down syndrome.

Down syndrome (DS) is a genetic disease that exhibits significant neuropathological parallels with Alzheimer's disease (AD). One of the features of DS, neuronal loss, has been hypothesized to occur as a result of apoptosis. An increasing number of proteins are implicated in apoptosis and several of them were shown to be altered in AD, however, the knowledge in DS is far from complete. To further substantiate the hypothesis that apoptosis is the underlying mechanism for neuronal loss and contribute towards the current knowledge of apoptosis in DS, we analyzed the expression of apoptosis related proteins in frontal cortex and cerebellum of DS by western blot and ELISA techniques. Quantitative analysis revealed a significant increase in DS frontal (P < 0.0001) and cerebellar (P < 0.05) Bim/BOD (Bcl-2 interacting mediator of cell death/Bcl-2 related ovarian death gene), cerebellar Bcl-2 (P < 0.01) as well as p21 (P < 0.05) levels compared to controls. No significant change was detected in Bax, RAIDD (receptor interacting protein (RIP)-associated ICH-1/CED-3-homologus protein with death domain), ZIP (Zipper interacting protein) kinase and NF-kappaB p65 levels in both regions, although frontal cortex levels of RAIDD, Bcl-2 and p21 levels tended to increase. In addition, a 45 kDa truncated form of NF-kappaB p65 displayed a significant elevation (P < 0.05) in DS cerebellum. No significant correlation had been obtained between postmortem interval and level of the proteins analyzed. With regard to age, it was only NF-kappaB p65 that showed significant correlation (r = -0.8964, P = 0.0155, n = 9) in frontal cortex of controls. These findings provide further evidence that apoptosis indeed accounts for the neuronal loss in DS but Bax and RAIDD do not appear to take part in this process.

Aged↗

PubMed2001

Increased brain protein levels of carbonyl reductase and alcohol dehydrogenase in Down syndrome and Alzheimer's disease.

Oxidative stress is considered to be crucial in the pathogenesis of Alzheimer's disease-like neurodegeneration. An elevation of carbonyl compounds that are biomarkers of and leading to oxidative stress has been demonstrated in Down Syndrome (DS) and Alzheimer's Disease (AD) brains and seems to be the result of a multifactorial process. Carbonyls, which are cytotoxic metabolic intermediates, are detoxified by either oxidation catalyzed by aldehyde dehydrogenase (ALDH), or by reduction to their corresponding alcohols by carbonyl reductase (CBR) and/or alcohol dehydrogenase (ADH). We quantified protein levels of CBR and its agonist/antagonist ADH using 2D gel electrophoresis with subsequent MALDI-identification and specific software in several brain regions of DS as well as AD patients and compared them to levels of CBR and ADH in brains of controls. Protein levels of both enzymes were increased in several brain regions because of enzyme induction by elevated carbonyls in DS and AD. Increased CBR in DS (trisomy 21) may be due to a gene dosage effect as the gene encoding CBR has been mapped to chromosome 21. These findings may confirm the proposed increase of reactive carbonyls in the brain thus supporting the involvement of oxidative stress and contribute to the understanding of carbonyl handling in brain of both neurodegenerative disorders.

Adult↗

PubMed2001

Carbohydrate handling enzymes in fetal Down syndrome brain.

Impaired glucose metabolism in adult Down Syndrome (DS) has been well-documented in vivo and information on the underlying biochemical defect i.e. aberrant glucose handling enzymes is already available. Nothing is known on carbohydrate handling, however, in early life of DS patients, when no secondary phenomena as e.g. Alzheimer-like neuropathology occur in the brain yet. We therefore determined a series of key enzymes of carbohydrate metabolism in fetal control and DS brain during the early second trimenon. We used two-dimensional electrophoresis with subsequent MALDI characterization and specific software for quantification of protein spots. We observed comparable levels of phosphoglycerate mutase, phosphoglycerate kinase 1; fructose-biphosphate aldolase A, fructose bisphosphate aldolase C; ribose-phosphate pyrophosphokinase 1; D-phosphoglycerate dehydrogenase, 6-phosphogluconolactonase; aflatoxin B1 aldehyde reductase 1, aldose reductase; inosine-5'-monophosphate dehydrogenase 2; galactokinase, in brain of fetal controls and DS. We conclude that our biochemical findings point to the fact that DS patients start early life with unchanged glucose handling, pentose phosphate shunt, glycolysis, sugar aldehyde, guanine nucleotide- and ribonucleoside formation and galactose metabolism.

Brain↗

PubMed2001

Changes in nicotinic acetylcholine receptor subunits expression in brain of patients with Down syndrome and Alzheimer's disease.

Cholinergic deficit associated with loss of nicotinic acetylcholine receptors (nAChRs) has been described in Alzheimer's disease (AD) by receptor binding assays, positron emission tomography and immunoblotting. However, little is known about the alteration of these receptors in a related disease, Down syndrome (DS) which might be of importance for therapeutic strategies. The protein levels of neuronal nAChR alpha and beta subunits in human postmortem brain samples (frontal cortex and cerebellum) of control, adult DS, and AD were investigated by making use of western blot analysis. Two major bands at 26 and 45 kDa for alpha3, one at 50 kDa for alpha4 and beta2, and one at 45 kDa for alpha7 were detected by the respective antibodies. Specific alteration in individual subunits was also apparent in DS and AD. In frontal cortex, the 45kDa alpha3 subunit was significantly increased in DS (121%) (P < 0.05) and AD (93%) (P < 0.05), whereas the 26kDa, an isoform/truncated form of alpha3, displayed a reversed pattern. It was significantly decreased in DS (75%) (P < 0.001) and AD (52.6%) (P < 0.05). Alpha4 was comparable in all groups by contrast, alpha7 was significantly decreased in AD (64%) (P < 0.05). In DS, however, although the levels tended to be lower (17.3%) the reduction was not significant. Beta2 was unchanged in AD but showed a significant increase in DS frontal cortex (98.1%) (P < 0.01). In cerebellum, no significant alteration was observed in any of the subunits except beta2. It exhibited a significant increase (161%) (P < 0.01) in DS. Derangement in expression of nAChRs is apparent in DS, as in AD that may have some relevance to DS neuropathology. Furthermore, the increase in beta2 expression indicate that these subunits may have more than a structural role. Hence, therapeutic strategies tailored towards these end might be of some benefit for cognitive enhancement in these disorders.

Aged↗

PubMed2001

Protein levels of human peroxiredoxin subtypes in brains of patients with Alzheimer's disease and Down syndrome.

Human peroxiredoxin (Prx) play important roles in eliminating hydrogen peroxide generated during cellular mechanisms using electrons from thioredoxin (Trx). Oxidative stress induced by reactive oxygen species (ROS) such as hydrogen peroxide has been implicated in the pathogenesis of several neurodegenerative diseases. We applied the proteomic approach to study protein levels of three subtypes of human Prx in brain regions from patients with Alzheimer's disease (AD) and Down Syndrome (DS). Protein levels of Prx-I and Prx-II were significantly increased in AD and DS. Protein levels of Prx-III, a mitochondrial protein, however, were significantly decreased. We conclude that increased protein levels of Prx-I and Prx-II could provide protection against neuronal cell death induced by hydrogen peroxide. Decreased protein levels of Prx-III could be caused by mitochondrial damage shown in AD and DS. Showing upregulated Prx protein levels provides evidence for the involvement of ROS in the pathogenesis of AD and DS.

Aged↗

PubMed2001

Effects of a single transdermal nicotine dose on cognitive performance in adults with Down syndrome.

Subjects with Down syndrome exhibit various types of cognitive impairment. Neuropathological and neurochemical studies revealed similarities between Down syndrome and Alzheimer's disease, cholinergic deficits being the most consistent findings. To explore the potential for cognitive enhancement utilizing nicotinic stimulation, 8 patients with Down syndrome (aged 18.5-31 years) received placebo and a single dose of transdermal nicotine (5 mg patch) over 2h in a single-blind, within-subjects repeated measures design. Auditory event-related potentials (ERPs) and neuropsychological tests, comprising digit symbol performance subtest from WAIS-R and the Frankfurt Attention Inventory (FAIR) were performed. Effects of nicotine administration in Down syndrome individuals were a decrease of ERP-P3 latency in 7 of 8 subjects (electrode position Cz: 386.9+/-24.0 ms vs. 363.1+/-26.9.2 ms, placebo vs. nicotine, respectively; P = 0.058) and an increase of ERP-P3 amplitude in 6 of 8 subjects (electrode position Cz: 17.4+/-5.5 vs. 18.0+/-4.5 microV, placebo vs. nicotine respectively; P = 0.725). Neuropsychological tests exhibited improvements in digit symbol performance subtest in 4 of 8 subjects and 7 of 8 subjects in the Frankfurt Attention Inventory. These results suggest that stimulating central nicotinic receptors might have an acute cognitive benefit in young adult Down syndrome subjects.

Administration, Cutaneous↗

PubMed2001

The brain in Down syndrome.

Down syndrome (trisomy 21) is a genetic disease with developmental brain abnormalities resulting in early mental retardation and precocious, age dependent Alzheimer-type neurodegeneration. We tried to discuss the role of neurodevelopmental abnormalities in connection with aberrant expression of genes on chromosome 21 including amyloid precursor protein (APP), CuZn superoxide dismutase (SOD1) and glial-derived S100 beta protein for neurodegeneration in DS. In this model, alterations in developmental pathways due to aberrant gene expression can impair cellular homeostasis and predispose to neurodegeneration of certain brain regions and types of nerve cells, involving cholinergic, serotonergic and catecholaminergic transmission, by shifting balance toward a pro-apoptotic state.

Brain↗

PubMed2001

Decreased levels of ARPP-19 and PKA in brains of Down syndrome and Alzheimer's disease.

ARPP-19 (cAMP-regulated phosphoprotein of Mr = 19,000) is a substrate for cAMP-dependent protein kinase (PKA). ARPP-19 is found in all brain regions but the function of ARPP-19 is not fully elucidated yet. We detected a downregulated sequence with 100% homology with ARPP-19 in temporal cortex of patients with Down syndrome (DS) as compared to controls, but not in Alzheimer's disease (AD) using differential displaypolymerase chain reaction (DD-PCR). We subsequently determined protein levels of ARPP-19 in temporal cortex and cerebellum by immunoblotting and observed significant reduction of ARPP-19 in DS (temporal cortex) and AD (cerebellum). We also observed decreased activities of PKA in DS (temporal cortex and cerebellum) and AD (temporal cortex). These findings suggest that decreased ARPP-19 along with decreased activities of PKA is involved in pathomechanisms of both neurodegenerative disorders. Furthermore, these findings provide first evidence for an impaired mechanism of cAMP-related signal transduction and phosphorylation in both dementing disorders.

Alzheimer Disease↗

PubMed2001

Increased protein levels of heterogeneous nuclear ribonucleoprotein A2/B1 in fetal Down syndrome brains.

Heterogeneous nuclear ribonucleoproteins (hnRNPs) are predominantly nuclear RNA-binding proteins that form complexes with RNA polymerase II transcripts. These proteins play pivotal roles in transcription, pre-mRNA processing in the nucleus, cytoplasmic mRNA translation and its turnover. In addition, hnRNPs have been shown to be essential for embryonic development of Drosophila. Here we studied the protein levels of hnRNPs (A2/B1, H and H') in fetal brain with Down syndrome (DS; n = 5) compared to controls (n = 7). We used two-dimensional (2-D) gel electrophoresis, matrix-assisted laser desorption ionization mass spectroscopy (MALDI-MS) and specific software for quantification. hnRNP A2/B1 was significantly increased in fetal DS brain (13.52+/-4.50) compared to controls (9.16+/-1.35), but both hnRNP H and H' were unchanged. Increased hnRNP A2/B1 in fetal DS brain may represent high activity of RNA processing such as RNA trafficking and telomere protection, and/or it could contribute to abnormal development of DS brains. Furthermore, comparable expression of hnRNP H and H' suggest a specific upregulation of hnRNP A2/B.

Brain↗

PubMed2001

Decreased protein levels of stathmin in adult brains with Down syndrome and Alzheimer's disease.

Stathmin, distributed in neurons with high abundance, acts as an intracellular relay, integrating various transduction pathways triggered by extracellular signals and it is involved in physiological regulation of microtubule destabilization. Stathmin has been also shown to be a critical molecule in pathology of neurodegeneration such as Alzheimer's disease (AD), particularly, in neurofibrillary tangle (NFT) formation. Here we evaluated protein levels of stathmin in adult brain from patients with AD and Down syndrome (DS) showing AD-like pathology by applying proteomic technologies with two-dimensional (2-D) gel electrophoresis, matrix-assisted laser desorption ionization mass spectroscopy (MALDI-MS) identification and specific software for quantification of proteins. Significantly decreased protein levels of stathmin were observed in frontal (2.12+/-1.17, n = 6) and temporal (3.05+/-2.81, n = 10) cortices of AD compared to controls (frontal cortex: 4.41+/-1.70, n = 8; temporal cortex: 5.26+/-2.26, n = 13). Stathmin was also significantly decreased in frontal (2.47+/-1.11, n = 7) and temporal (2.02+/-1.18, n = 9) cortices of DS. We also investigated stathmin levels in fetal brain. Stathmin was not significantly changed between fetal DS brain and controls. We suggest that the decreased protein level of stathmin in brains is associated with tangle formation and microtubule instability in DS as well as AD, but stathmin is not involved in the abnormal development of fetal DS brain.

Adult↗