Publications by authors named "Katharina Flach"

7 Publications

  • Page 1 of 1

Non-Phosphorylated Tau as a Potential Biomarker of Alzheimer's Disease: Analytical and Diagnostic Characterization.

J Alzheimers Dis 2017 ;55(1):159-170

Department of Psychiatry and Psychotherapy, Universitätsklinikum Erlangen, and Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.

Background: Virtually nothing is known about a potential diagnostic role of non-phospho-epitopes of Tau (Non-P-Tau) in cerebrospinal fluid (CSF).

Objective: To establish and analytically and clinically characterize the first assay capable to measure concentrations of Non-P-Tau in human CSF.

Methods: An antibody (1G2) was developed that selectively binds to the Tau molecule non-phosphorylated at the positions T175 and T181, and was used in establishing a sandwich ELISA capable to measure Non-P-Tau in human CSF, following analytical and clinical validation of the method.

Results: The 1G2 antibody shows decreasing reactivity to tau peptides containing phosphorylation mainly at positions T175 and T181. Detection limit of the assay is 25 pg/ml; the coefficients of variation (CVs) of the optical densities of the repeated standard curves were between 3.6-15.9%. Median intra-assay imprecision of double measurements was 4.8%; inter-assay imprecision was in the range of 11.2% - 15.3%. Non-P-Tau concentrations are stable in the CSF samples sent to distinct laboratories under ambient temperature; inter-laboratory variation was approximately 30%. The Non-P-Tau CSF concentrations were highly significantly increased in patients with Alzheimer's disease in stage of mild cognitive impairment or dementia (AD/MCI, n = 58, 109.2±32.0 pg/mL) compared to the non-demented Controls (n = 42, 62.1±9.3 pg/mL, p < 0.001). At the cut-off of 78.3 pg/mL, the sensitivity and the specificity were 94.8% and 97.6%, respectively.

Conclusion: For the first time, an assay is reported to reliably measure concentrations of non-phosphorylated Tau in human CSF.
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http://dx.doi.org/10.3233/JAD-160448DOI Listing
February 2018

Hook proteins: association with Alzheimer pathology and regulatory role of hook3 in amyloid beta generation.

PLoS One 2015 23;10(3):e0119423. Epub 2015 Mar 23.

Paul Flechsig Institute of Brain Research, Department of Molecular and Cellular Mechanism of Neurodegeneration, University of Leipzig, Leipzig, Germany.

Defects in intracellular transport are implicated in the pathogenesis of Alzheimer's disease (AD). Hook proteins are a family of cytoplasmic linker proteins that participate in endosomal transport. In this study we show that Hook1 and Hook3 are expressed in neurons while Hook2 is predominantly expressed in astrocytes. Furthermore, Hook proteins are associated with pathological hallmarks in AD; Hook1 and Hook3 are localized to tau aggregates and Hook2 to glial components within amyloid plaques. Additionally, the expression of Hook3 is reduced in AD. Modelling of Hook3 deficiency in cultured cells leads to slowing of endosomal transport and increases β-amyloid production. We propose that Hook3 plays a role in pathogenic events exacerbating AD.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0119423PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4370497PMC
March 2016

Tenascin-R promotes assembly of the extracellular matrix of perineuronal nets via clustering of aggrecan.

Philos Trans R Soc Lond B Biol Sci 2014 Oct;369(1654):20140046

Zentrum für Molekulare Neurobiologie, Universitätsklinikum Hamburg - Eppendorf, Martinistr. 52, 20246 Hamburg, Germany Center for Neuroscience, Shantou University Medical College, 22 Xin Ling Road, Shantou 515041, People's Republic of China Keck Center for Collaborative Neuroscience and Department of Cell Biology, Rutgers University, 604 Allison Road, Piscataway, NJ 08554, USA

Perineuronal nets (PNs) in the brains of tenascin-R-deficient (tn-r(-/-)) mice develop in temporal concordance with those of wild-type (tn-r(+/+)) mice. However, the histological appearance of PNs is abnormal in adult tn-r(-/-) mice. Here, we investigated whether similar defects are also seen in dissociated and organotypic cultures from hippocampus and forebrain of tn-r(-/-) mice and whether the structure of PNs could be normalized. In tn-r(-/-) cultures, accumulations of several extracellular matrix molecules were mostly associated with somata, whereas dendrites were sparsely covered, compared with tn-r(+/+) mice. Experiments to normalize the structure of PNs in tn-r(-/-) organotypic slice cultures by depolarization of neurons, or by co-culturing tn-r(+/+) and tn-r(-/-) brain slices failed to restore a normal PN phenotype. However, formation of dendritic PNs in cultures was improved by the application of tenascin-R protein and rescued by polyclonal antibodies to aggrecan and a bivalent, but not monovalent form of the lectin Wisteria floribunda agglutinin. These results show that tenascin-R and aggrecan are decisive contributors to formation and stabilization of PNs and that tenascin-R may implement these functions by clustering of aggrecan. Proposed approaches for restoration of normal PN structure are noteworthy in the context of PN abnormalities in neurological disorders, such as epilepsy, schizophrenia and addiction.
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http://dx.doi.org/10.1098/rstb.2014.0046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4173296PMC
October 2014

Detection of disease-associated α-synuclein in the cerebrospinal fluid: a feasibility study.

Clin Neuropathol 2014 Sep-Oct;33(5):329-34

Institute of Neurology, Medical University of Vienna, Vienna, Austria, AJ Roboscreen GmbH, Leipzig, Germany, Department of Neurology, present address: Department of Medicine I, Clinical Division of Oncology, Medical University of Vienna, Vienna, Austria, and Neurobiologie, CMRR, Hospices Civils de Lyon, Université Lyon, Lyon, France.

With the aim to evaluate the significance and reliability of detecting disease-specific α-synuclein in the cerebrospinal fluid (CSF) we developed an ELISA and bead-assay. We used a commercial antibody (5G4) that does not bind to the physiological monomeric form of α-synuclein, but is highly specific for the disease-associated forms, including high molecular weight fraction of β-sheet rich oligomers. We applied both tests in CSF from a series of neuropathologically confirmed α-synucleinopathy cases, including Parkinson' disease dementia (PDD) and dementia with Lewy bodies (DLB) (n = 7), as well as Alzheimer' disease (n = 6), and control patients without neurodegenerative pathologies (n = 9). Disease-specific α-synuclein was detectable in the CSF in a subset of patients with α-synuclein pathology in the brain. When combined with the analysis of total α-synuclein, the bead-assay for disease-specific α-synuclein was highly specific for PDD/DLB. Detection of disease-associated αsynuclein combined with the total levels of α-synuclein is a promising tool for the in-vivo diagnosis of α-synucleinopathies, including PDD and LBD.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4151342PMC
http://dx.doi.org/10.5414/np300796DOI Listing
October 2014

Axotrophin/MARCH7 acts as an E3 ubiquitin ligase and ubiquitinates tau protein in vitro impairing microtubule binding.

Biochim Biophys Acta 2014 Sep 4;1842(9):1527-38. Epub 2014 Jun 4.

Paul Flechsig Institute of Brain Research, Department of Molecular and Cellular Mechanisms of Neurodegeneration, University of Leipzig, 04109 Leipzig, Germany. Electronic address:

Tau is the major microtubule-associated protein in neurons involved in microtubule stabilization in the axonal compartment. Changes in tau gene expression, alternative splicing and posttranslational modification regulate tau function and in tauopathies can result in tau mislocalization and dysfunction, causing tau aggregation and cell death. To uncover proteins involved in the development of tauopathies, a yeast two-hybrid system was used to screen for tau-interacting proteins. We show that axotrophin/MARCH7, a RING-variant domain containing protein with similarity to E3 ubiquitin ligases interacts with tau. We defined the tau binding domain to amino acids 552-682 of axotrophin comprising the RING-variant domain. Co-immunoprecipitation and co-localization confirmed the specificity of the interaction. Intracellular localization of axotrophin is determined by an N-terminal nuclear targeting signal and a C-terminal nuclear export signal. In AD brain nuclear localization is lost and axotrophin is rather associated with neurofibrillary tangles. We find here that tau becomes mono-ubiquitinated by recombinant tau-interacting RING-variant domain, which diminishes its microtubule-binding. In vitro ubiquitination of four-repeat tau results in incorporation of up to four ubiquitin molecules compared to two molecules in three-repeat tau. In summary, we present a novel tau modification occurring preferentially on 4-repeat tau protein which modifies microtubule-binding and may impact on the pathogenesis of tauopathies.
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http://dx.doi.org/10.1016/j.bbadis.2014.05.029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4311138PMC
September 2014

Tau oligomers impair artificial membrane integrity and cellular viability.

J Biol Chem 2012 Dec 5;287(52):43223-33. Epub 2012 Nov 5.

Department of Molecular and Cellular Mechanisms of Neurodegeneration, Paul Flechsig Institute of Brain Research, Faculty of Medicine, Jahnallee 59, University of Leipzig, 04109 Leipzig, Germany.

The microtubule-associated protein Tau is mainly expressed in neurons, where it binds and stabilizes microtubules. In Alzheimer disease and other tauopathies, Tau protein has a reduced affinity toward microtubules. As a consequence, Tau protein detaches from microtubules and eventually aggregates into β-sheet-containing filaments. The fibrillization of monomeric Tau to filaments is a multistep process that involves the formation of various aggregates, including spherical and protofibrillar oligomers. Previous concepts, primarily developed for Aβ and α-synuclein, propose these oligomeric intermediates as the primary cytotoxic species mediating their deleterious effects through membrane permeabilization. In the present study, we thus analyzed whether this concept can also be applied to Tau protein. To this end, viability and membrane integrity were assessed on SH-SY5Y neuroblastoma cells and artificial phospholipid vesicles, treated with Tau monomers, Tau aggregation intermediates, or Tau fibrils. Our findings suggest that oligomeric Tau aggregation intermediates are the most toxic compounds of Tau fibrillogenesis, which effectively decrease cell viability and increase phospholipid vesicle leakage. Our data integrate Tau protein into the class of amyloidogenic proteins and enforce the hypothesis of a common toxicity-mediating mechanism for amyloidogenic proteins.
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http://dx.doi.org/10.1074/jbc.M112.396176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3527910PMC
December 2012

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

J Biol Chem 2007 Mar 1;282(10):6984-91. Epub 2006 Nov 1.

Nutritional Physiology Unit "Oskar Kellner," Wilhelm-Stahl-Allee 2, 18196 Dummerstorf, Germany.

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.
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http://dx.doi.org/10.1074/jbc.M609521200DOI Listing
March 2007