Biomedical subjects

Thomas Arendt

Publications and source records attributed to Thomas Arendt.

At least 19 recordsLinked to original sources

PubMed2007

Expression of embryonic tau protein isoforms persist during adult neurogenesis in the hippocampus.

Tau is a microtubule-associated protein with a developmentally regulated expression of multiple isoforms. The neonatal isoform is devoid of two amino terminal inserts and contains only three instead of four microtubule-binding repeats (0N/3R-tau). We investigated the temporal expression pattern of 0N-tau and 3R-tau in the rat hippocampus. After the decline of 0N- and 3R-tau immunoreactivity during the postnatal development both isoforms remain highly expressed in a few cells residing beneath the granule cell layer. Coexpression of the polysialylated neuronal cell adhesion molecule, doublecortin, and incorporated bromodeoxyuridine showed that these cells are proliferating progenitor cells. In contrast mature granule cells express the adult tau protein isoform containing one aminoterminal insert domain (1N-tau). Therefore a shift in tau isoform expression takes place during adult neurogenesis, which might be related to migration, differentiation, and integration in the granule cell layer. A model for studying shifts in tau isoform expression in a defined subset of neurons might help to understand the etiology of tauopathies, when isoform composition is crucial for neurodegeneration, as in Pick's disease or FTDP-17.

Animals↗

PubMed2006

Effect of pseudophosphorylation and cross-linking by lipid peroxidation and advanced glycation end product precursors on tau aggregation and filament formation.

Accumulation of hyperphosphorylated Tau protein as paired helical filaments in pyramidal neurons is a major hallmark of Alzheimer disease. Besides hyperphosphorylation, other modifications of the Tau protein, such as cross-linking, are likely to contribute to the characteristic features of paired helical filaments, including their insolubility and resistance against proteolytic degradation. In this study, we have investigated whether the four reactive carbonyl compounds acrolein, malondialdehyde, glyoxal, and methylglyoxal accelerate the formation of Tau oligomers, thioflavin T-positive aggregates, and fibrils using wild-type and seven pseudophosphorylated mutant Tau proteins. Acrolein and methylglyoxal were the most reactive compounds followed by glyoxal and malondialdehyde in terms of formation of Tau dimers and higher molecular weight oligomers. Furthermore, acrolein and methylglyoxal induced the formation of thioflavin T-fluorescent aggregates in a triple pseudophosphorylation-mimicking mutant to a slightly higher degree than wild-type Tau. Analysis of the Tau aggregates by electron microscopy study showed that formation of fibrils using wild-type Tau and several Tau mutants could be observed with acrolein and methylglyoxal but not with glyoxal and malondialdehyde. Our results suggest that reactive carbonyl compounds, particularly methylglyoxal and acrolein, could accelerate tangle formation in vivo and that this process could be slightly accelerated, at least in the case of methylglyoxal and acrolein, by hyperphosphorylation. Interference with the formation or the reaction of these reactive carbonyl compounds could be a promising way of inhibiting tangle formation and neuronal dysfunction in Alzheimer disease and other tauopathies.

Acrolein↗

PubMed2006

Constitutive expression of functionally active cyclin-dependent kinases and their binding partners suggests noncanonical functions of cell cycle regulators in differentiated neurons.

Neurodegeneration in Alzheimer's disease and various experimental lesion paradigms are associated with an unscheduled upregulation of cell cycle-related proteins, indicating a link between cell cycle reactivation and neuronal death. Recent evidence, however, suggests that at least some of the canonical cell cycle regulators are constitutively expressed in differentiated neurons of the adult brain. Systematic investigations on the constitutive expression of cell cycle regulators in differentiated neurons in vivo, providing the basis for further insights into their potential role under pathological conditions, however, have not been carried out. Here, we demonstrate a constitutive neuronal expression of Cdks 1, 2, and 4; their activators cyclins D, A, B, and E; and their inhibitors p15(Ink4b), p16(Ink4a), p18(Ink4c), p19(Ink4d), p21(Waf1/Cip1), p27(Kip1), and p57(Kip2) within the neocortex of adult mice by western blot and immunocytochemistry. Expression was verified by single-cell reverse transcriptase-polymerase chain reaction applied to individual microscopically identified neurons captured with laser dissection. Immunoprecipitation and in vitro kinase assays revealed that Cdks 1, 2, and 4 are properly complexed to cyclins and exhibit kinase activity. This physiological expression of positive cell cycle regulators in adult neurons is clearly not related to neuronal proliferation. Taken together, our findings demonstrate a constitutive expression of functionally active cyclin-dependent kinases and their regulators in differentiated neurons suggesting a noncanonical role of cell cycle regulators potentially linked to neuronal plasticity and/or stability.

Animals↗

PubMed2006

Simultaneous detection of tau phospho-epitopes with haptenylated antibodies.

Neurofibrillary tangles as a neuropathological hallmark of Alzheimer's disease are mainly composed of abnormally phosphorylated microtubule-associated protein tau. The present work was primarily focused on the immunohistochemical characterization of recently developed monoclonal antibodies directed against disease-associated epitopes. Anti-phospho-threonine 212/phospho-serine 214 (HPT-1), anti-phospho-threonine 231/phospho-serine 235 (HPT-101) and their biotinylated derivatives were shown to be sensitive markers for the immunohistochemical detection of neuropathological alterations during Alzheimer's disease. Triple carbocyanine immunofluorescence labelling was based on digoxigenylated, fluoresceinated and biotinylated primary antibodies. AT8-immunolabelling of phospho-serine 202 and phospho-threonine 205 combined with HPT-1 and HPT-101-staining revealed similar distribution patterns of the three double-phosphorylated tau epitopes in the neocortex of patients with Alzheimer's disease.

Alzheimer Disease↗

PubMed2006

Different dendrite and dendritic spine alterations in basal and apical arbors in mutant human amyloid precursor protein transgenic mice.

The extracellular deposition of amyloid-beta peptide (Abeta) in brain parenchyma is one of the characteristic features of Alzheimer's disease and is suggested to induce reactive and degenerative changes in neuronal cell bodies, axons and dendritic processes. In particular, within and in close proximity to amyloid plaques, distinctive morphological alterations have been observed, including changes in neurite trajectory and decreases in dendritic diameter and in spine density. Apart from these plaque-associated focal aberrations, little is known regarding modifications of the global dendritic morphology including the detailed and comparative quantitative analysis of apical and basal arbors. The objective of the present study was to investigate the effects of amyloid plaque deposition and elevated soluble Abeta on neuronal morphology in mutant human amyloid precursor protein (hAPP) transgenic mice (line Tg2576; [K. Hsiao, P. Chapman, S. Nilsen, C. Eckman, Y. Harigaya, S. Younkin, F. Yang, G. Cole, Correlative memory deficits, Abeta elevation, and amyloid plaques in transgenic mice, Science 274 (1996) 99-102]). Retrogradelly labeled callosal-projecting pyramidal cells in the primary somatosensory cortex were three-dimensionally analyzed. Although basal dendrites remained unaffected, analysis of apical trees revealed a number of unambiguous morphological changes. Thus, in TG2576 mice, the apical arbors were shortened in total length and less branched. Furthermore, the diameter of proximal dendritic segments was increased whereas that of distal segments was reduced. Analysis of spine numbers and distribution on basal and apical trees demonstrated a significant reduction in spine densities along the whole course of dendrites. The findings suggest that Abeta-related pathology induces morphological aberrations in basal and apical arbors to different degrees which are unrelated to direct plaque-associated changes.

Amyloid beta-Protein Precursor↗

PubMed2006

The unfolded protein response affects neuronal cell cycle protein expression: implications for Alzheimer's disease pathogenesis.

Alzheimer's disease (AD) is characterized by the accumulation and aggregation of misfolded proteins. The presence of misfolded proteins in the endoplasmic reticulum (ER) triggers a cellular stress response called the unfolded protein response (UPR). Previously, we have shown that the UPR is activated in AD neurons. In actively dividing cells, activation of the UPR is accompanied by decreased cell cycle protein expression and an arrest in the G1 phase of the cell cycle. Aberrant expression of cell cycle proteins has been observed in post mitotic neurons in AD and is suggested to be involved in neurodegeneration. In this study we show that the protein levels of BiP/GRP78, an ER-stress marker, is increased in Braak stages B and C for amyloid deposits. This is in contrast to the levels of cell cycle markers cyclin D1, cyclin E and phosphorylated retinoblastoma protein (ppRb) which are decreased in Braak stage C compared to Braak stage A for amyloid deposits. In addition, we report a negative correlation between neuronal expression of ppRb and expression levels of BiP/GRP78 in control and AD cases. Activation of the UPR in neuronal cells induces changes in cell cycle protein expression similar to these observed in AD brain. ER stress inducers tunicamycin and thapsigargin down-regulate cell cycle proteins ppRb and cyclin D1 in differentiated neuroblastoma cells. In contrast, protein levels of p27, a cyclin dependent kinase inhibitor, are increased after induction of ER-stress using tunicamycin. These data suggest that activation of the UPR affects cell cycle protein expression in neurons during neurodegeneration in AD.

Alzheimer Disease↗

PubMed2006

Age- and stage-dependent glyoxalase I expression and its activity in normal and Alzheimer's disease brains.

The reaction of lysine and arginine residues of proteins with 1,2-dicarbonyl compounds result in the formation of advanced glycation end products (AGEs). Accumulation of AGEs is a characteristic feature of the aging brain and contributes to the development of neurodegenerative diseases such as Alzheimer's disease (AD). Therefore, it is of particular interest to study the cellular defense mechanisms against AGE formation and particularly the detoxification of their precursors. AGE precursor compounds such as methylglyoxal and glyoxal were cellulary detoxified by the glyoxalase system, consisting of glyoxalases I and II. Glyoxalase I levels are diminished in old aged brains but elevated in AD brains. However, it is still unknown how glyoxalase I level of AD brains changes in a disease and in an age-dependent manner. Therefore, we investigated the AD stage- and the age-dependent levels of glyoxalase I in the Brodmann area 22 of AD brains (n=25) and healthy controls (n=10). Our results obtained from RT-PCR reveal reducing glyoxalase I RNA levels with advancing stage of AD and with increasing age. Western Blot analysis indicates that in comparison to healthy controls, glyoxalase I protein amounts are 1.5-fold increased in early AD subjects and continuously decrease in middle and late stages of AD. The glyoxalase I protein amounts of AD patients also decrease with age. Results obtained from glyoxalase I activity measurement show 1.05-1.2-fold diminished levels in AD brains compared to healthy controls and no significant decrease neither with the stage of AD nor with age. The immunohistochemical investigations demonstrate an elevated number of glyoxalase I stained neurons in brains of early and middle but not in late AD subjects compared to age-matched healthy controls. In addition, the stage-dependent immunohistochemical investigation demonstrates that with reduced glyoxalase I staining AGE deposits prevail, specifically in late stage of AD. In conclusion, the decrease of glyoxalase I expression with increasing AD stage might be one reason for methylglyoxal-induced neuronal impairment, apoptosis, and AGE formation in plaques and tangles.

Aged↗

PubMed2006

Laser scanning cytometry in human brain slices.

BACKGROUND: The Laser Scanning Cytometry (LSC) offers quantitative fluorescence analysis of cell suspensions and tissue sections. METHODS: We adapted this technique to immunohistochemical labelled human brain slices. RESULTS: We were able to identify neurons according to their labelling and to display morphological structures such as the lamination of the entorhinal cortex. Further, we were able to distinguish between neurons with and without cyclin B1 expression and we could assign the expression of cyclin B1 to the cell islands of layer II and the pyramidal neurons of layer V of the entorhinal cortex in Alzheimer's disease effected brain. In addition, we developed a method depicting the three-dimensional distribution of the cells in intact tissue sections. CONCLUSIONS: In this pilot experiments we could demonstrate the power of the LSC for the analysis of human brain sections.

Alzheimer Disease↗

PubMed2006

Pathological effects of glyoxalase I inhibition in SH-SY5Y neuroblastoma cells.

In Alzheimer's disease (AD), in aging, and under conditions of oxidative stress, the levels of reactive carbonyl compounds continuously increase. Accumulating carbonyl levels might be caused by an impaired enzymatic detoxification system. The major dicarbonyl detoxifying system is the glyoxalase system, which removes methylglyoxal in order to minimize cellular impairment. Although a reduced activity of glyoxalase I was evident in aging brains, it is not known how raising the intracellular methylglyoxal level influences neuronal function and the phosphorylation pattern of tau protein, which is known to be abnormally hyperphosphorylated in AD. To simulate a reduced glyoxalase I activity, we applied an inhibitor of glyoxalase I, p-bromobenzylglutathione cyclopentyl diester (pBrBzGSCp(2)), to SH-SY5Y neuroblastoma cells to induce chronically elevated methylglyoxal concentrations. We have shown that 10 microM pBrBzGSCp(2) leads to a fourfold elevation of the methylglyoxal level after 24 hr. In addition, glyoxalase I inhibition leads to reduced cell viability, strongly retracted neuritis, increase in [Ca(2+)](i), and activation of caspase-3. However, pBrBzGSCp(2) did not lead to tau "hyper"-phosphorylation despite activation of p38 mitogen-activated protein kinase and c-Jun NH(2)-terminal kinase but rather activated protein phosphatases 2 and induced tau dephosphorylation at the Ser(202)/Thr(205) and Ser(396)/Ser(404) epitopes. Preincubation with the carbonyl scavenger aminoguanidine prevented tau dephosphorylation, indicating the specific effect of methylglyoxal. Also, pretreatment with the inhibitor okadaic acid prevented tau dephosphorylation, indicating that methylglyoxal activates PP-2A. In summary, our data suggest that a reduced glyoxalase I activity mimics some changes associated with neurodegeneration, such as neurite retraction and apoptotic cell death.

Alzheimer Disease↗

PubMed2006

Identification of AGE-modified proteins in SH-SY5Y and OLN-93 cells.

The formation of "Advanced Glycation End products" (AGEs) is an inevitable consequence of mammalian glucose metabolism. AGE-mediated protein-protein crosslinks lead to detergent-insoluble and protease-resistant protein aggregates, and in Alzheimer's disease (AD) extra cellular senile plaques (SPs) and intracellular neurofibrillary tangles (NFTs) have been shown to contain AGEs. However, to date little is known concerning the most prevalent protein-targets of AGE modification under normal, non-pathological conditions. Here, a combination of 2D-electrophoresis, Western blotting and mass spectrometry has been used to identify preferentially AGE-modified proteins in oligodendrocyte (OLN-93) and neuroblastoma cell lines (SH-SY5Y) in standard culture. Proteomics analysis identified a total of eight targets with structural, metabolic and regulatory function, three of which (beta-actin, beta-tubulin and eukaryotic Elongation Factor 1-alpha) were common to both cell lines. Based on results from prior studies, modification of these proteins may lead to a loss of function. Consequently, the identification of targets for these proteins is of particular interest for a better understanding of the consequences of AGE-modification in aging, neurodegenerative diseases and diabetes.

Animals↗

PubMed2006

Altered subcellular location of phosphorylated Smads in Alzheimer's disease.

A number of growth factors and cytokines, such as transforming growth factor beta 1 (TGF-beta1), is elevated in Alzheimer's disease (AD), giving rise to activated intracellular mitogenic signaling cascades. Activated mitogenic signaling involving the mitogen-activated protein kinases (MAPKs) and other protein kinases might alter the phosphorylation states of structural proteins such as tau, resulting in hyperphosphorylated deposits. Many intracellular signaling proteins are potential targets of misregulated phosphorylation and dephosphorylation. Recently, a crosstalk between MAPKs and Smad proteins, both involved in mediating TGF-beta1 signaling, has been reported. Although TGF-beta1 has previously been shown to be involved in the pathogenesis of AD, the role of Smad proteins has not been investigated. In this study we thus analysed the subcellular distribution of phosphorylated Smad2 and Smad3 in the hippocampus of both normal and AD brains. Here we report on strong nuclear detection of phosphorylated Smad2 and Smad3 in neurons of control brains. In AD brains these phosphorylated proteins were additionally found in cytoplasmic granules in hippocampal neurons, within amyloid plaques and attached to neurofibrillary tangles. Our data suggest a critical role of Smad proteins in the pathogenesis of AD.

Aged↗

PubMed2005

Systematic analysis of gene expression in human brains before and after death.

BACKGROUND: Numerous studies have employed microarray techniques to study changes in gene expression in connection with human disease, aging and evolution. The vast majority of human samples available for research are obtained from deceased individuals. This raises questions about how well gene expression patterns in such samples reflect those of living individuals. RESULTS: Here, we compare gene expression patterns in two human brain regions in postmortem samples and in material collected during surgical intervention. We find that death induces significant expression changes in more than 10% of all expressed genes. These changes are non-randomly distributed with respect to their function. Moreover, we observe similar expression changes due to death in two distinct brain regions. Consequently, the pattern of gene expression differences between the two brain regions is largely unaffected by death, although the magnitude of differences is reduced by 50% in postmortem samples. Furthermore, death-induced changes do not contribute significantly to gene expression variation among postmortem human brain samples. CONCLUSION: We conclude that postmortem human brain samples are suitable for investigating gene expression patterns in humans, but that caution is warranted in interpreting results for individual genes.

Adolescent↗

PubMed2005

Differentially expressed cortical genes contribute to perivascular deposition in transgenic mice with inducible neuron-specific expression of TGF-beta1.

In the brain the expression of transforming growth factor beta1 (TGF-beta1) is involved both in neuroprotective and neurodegenerative processes. Recently, we have established a transgenic mouse model with inducible neuron-specific expression of TGF-beta1 based on the tetracycline-regulated gene expression system. A long-term expression of TGF-beta1 results in persisting perivascular thioflavin-positive depositions, which did not disappear even though the transgene synthesis was repressed completely by administration of doxycycline. Formation and composition of these depositions are hardly elucidated. The aim of this study was to identify TGF-beta1 responding genes potentially participating in forming these depositions. To address this problem we have compared the cortical mRNA expression pattern of TGF-beta1 expressing mice with mice impeded to express the transgenic protein using oligonucleotide microarray analysis. Differential gene expression was further characterized by quantitative real-time reverse transcription-polymerase chain reaction including animals, where the long-lasting TGF-beta1 expression was repressed. While no change of amyloid precursor protein RNA expression level was detected, various genes strongly involved in calcium homeostasis, tissue mineralization or vascular calcification were identified differentially expressed. It is suggested, that these genes might contribute to the perivascular depositions in the TGF-beta1 expressing mice.

Animals↗

PubMed2005

The expression of wild-type human amyloid precursor protein affects the dendritic phenotype of neocortical pyramidal neurons in transgenic mice.

The current study addresses the morphoregulatory effects of human amyloid precursor protein expression on neocortical pyramidal cells in vivo. For this purpose, a transgenic mouse line was used that expresses wild-type human amyloid precursor protein (APP) at levels similar to endogenous mouse APP. This strain does not develop Alzheimer's disease-related pathology which allowed to study effects of APP or APP cleavage products but excluded the influence of amyloid deposits. Commissural projecting pyramidal neurons of layers II/III within the primary somatosensory cortex were retrogradely labelled by injection of biotinylated dextran amine into the corpus callosum. In transgenic mice, computer-aided morphometric analysis revealed an increase in the surface area of proximal and intermediate basal dendritic segments resulting from an enlarged diameter. On the other hand, the length of the same segments was reduced. Both basal and apical dendrites were characterized by a higher dendritic density within the proximal and intermediate fields. Although the total spatial extension of basal and apical dendrites remained unchanged, a moderate withdrawal of arbors is suggested. The results implicate a physiological function for APP in regulatory mechanisms of neuronal morphogenesis.

Amyloid beta-Protein Precursor↗

PubMed2005

Maximal COX-2 and ppRb expression in neurons occurs during early Braak stages prior to the maximal activation of astrocytes and microglia in Alzheimer's disease.

Neuronal expression of cyclooxygenase-2 (COX-2) and cell cycle proteins is suggested to contribute to neurodegeneration during Alzheimer's disease (AD). The stimulus that induces COX-2 and cell cycle protein expression in AD is still elusive. Activated glia cells are shown to secrete substances that can induce expression of COX-2 and cell cycle proteins in vitro. Using post mortem brain tissue we have investigated whether activation of microglia and astrocytes in AD brain can be correlated with the expression of COX-2 and phosphorylated retinoblastoma protein (ppRb). The highest levels of neuronal COX-2 and ppRb immunoreactivity are observed in the first stages of AD pathology (Braak 0-II, Braak A). No significant difference in COX-2 or ppRb neuronal immunoreactivity is observed between Braak stage 0 and later Braak stages for neurofibrillary changes or amyloid plaques. The mean number of COX-2 or ppRb immunoreactive neurons is significantly decreased in Braak stage C compared to Braak stage A for amyloid deposits. Immunoreactivity for glial markers KP1, CR3/43 and GFAP appears in the later Braak stages and is significantly increased in Braak stage V-VI compared to Braak stage 0 for neurofibrillary changes. In addition, a significant negative correlation is observed between the presence of KP1, CR3/43 and GFAP immunoreactivity and the presence of neuronal immunoreactivity for COX-2 and ppRb. These data show that maximal COX-2 and ppRb immunoreactivity in neurons occurs during early Braak stages prior to the maximal activation of astrocytes and microglia. In contrast to in vitro studies, post mortem data do not support a causal relation between the activation of microglia and astrocytes and the expression of neuronal COX-2 and ppRb in the pathological cascade of AD.

Journal Article↗

PubMed2005

Morphometry of the amygdala in patients with questionable dementia and mild dementia.

The volume of the amygdala is reduced in advanced Alzheimer's disease (AD). However, there is controversy whether amygdala atrophy is present in mild AD and in the transitional phase between health and the onset of dementia. The aim of this prospective longitudinal study was to investigate whether amygdala atrophy is present in subjects with questionable dementia and mild dementia and whether amygdala volume is associated with the future rate of cognitive change, that is the annual change in the Mini Mental State Examination (MMSE). At baseline, volumes of the amygdala were measured in 97 participants aged 70-87 years (40 controls, 33 patients with questionable dementia, 24 patients with mild AD) using magnetic resonance imaging. Eighty-six participants were clinically re-examined after 2.3 years on average. At baseline, significant differences in mean amygdala volume were found between controls and participants with mild AD. There was no significant correlation between the longitudinal annual change in MMSE and the baseline amygdala volume in any of the three groups.

Aged↗

PubMed2005

Age-dependent changes of glyoxalase I expression in human brain.

Increased modification and crosslinking of proteins by advanced glycation end products (AGEs) is a characteristic feature of aging, and contributes to the formation of many of the lesions of neurodegenerative diseases including neurofibrillary tangles and amyloid plaques in Alzheimer's disease. Therefore, defense mechanisms against AGE formation or detoxification of their precursors such as the glyoxalase system are of particular interest in aging research. Thus, we investigated the age-dependent protein expression, the activity as well as the RNA level of glyoxalase I in Brodmann area 22 (auditory association area of superior temporal gyrus) of the human cerebral cortex. Our immunohistochemical results demonstrate the localization of glyoxalase I in neurons, predominantly pyramidal cells, as well as in astroglia, located predominantly in the subpial region. The number of glyoxalase I expressing neurons and astroglia increases with age, with a peak at approximately 55 years, and progressively decreases thereafter. These results were confirmed by biochemical investigations in total brain tissue, where the RNA, the protein level as well as the activity of glyoxalase I enzyme were analyzed in different age groups. In conclusion, the increase in glyoxalase I expression up to the age of 55 may be a compensatory mechanism against high oxoaldyde levels and the accumulation of AGEs. However, the decline of glyoxalase expression and activity in old age, possibly caused by impairment in transcription or/and translation, may subsequently lead to increased levels of reactive carbonyl compounds, followed by protein crosslinking, inflammation, oxidative stress and neuronal degeneration.

Adult↗

PubMed2005

Enhanced Ras activity promotes spine formation in synRas mice neocortex.

The effect of enhanced Ras activity upon dendritic spine formation has been investigated on pyramidal cells in the adult neocortex of synRas mice over-expressing Val12-Ha-Ras specifically in postmitotic neurons. For quantitative analysis, commissural neurons of layers II/III were retrogradely labelled with dextran amine. In synRas mice, spine frequency on several orders of basal dendrites and apical oblique branches was significantly increased. Whereas density on basal dendrites was overall augmented, on apical dendrites, distal regions were most affected. The proportions of different morphological types of spines remained unchanged. It is concluded that Ras might be critically involved in mechanisms that mediate and regulate dendritic spine formation in the adult brain.

Animals↗