Manuel Gunkel

Universität Hamburg

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Co-Authors

Holger Erfle

Heidelberg University

Michael Hausmann

Ben-Gurion University of the Negev

Karsten Rippe

Heidelberg University

Christoph Cremer

Heidelberg University

Rainer Kaufmann

Centre for Structural Systems Biology

Paul Lemmer

Heidelberg University

Karl Rohr

Heidelberg University

Vytaute Starkuviene

Heidelberg University

Patrick Müller

Stuttgart University of Applied Sciences

Jürgen Reymann

Heidelberg University

Cooperative Institutions

Heidelberg University

118

University Hospital Heidelberg

German Cancer Research Center

Heidelberg University

DKFZ-ZMBH Alliance

European Molecular Biology Laboratory

Kirchhoff (Germany)

Universität Hamburg

University Medical Center Hamburg-Eppendorf

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Tissue-Mimicking Geometrical Constraints Stimulate Tissue-Like Constitution and Activity of Mouse Neonatal and Human-Induced Pluripotent Stem Cell-Derived Cardiac Myocytes

Journal of Functional Biomaterials (2016)

Götz Pilarczyk Alexandra Raulf Manuel Gunkel Bernd K. Fleischmann Robert Lemor

The present work addresses the question of to what extent a geometrical support acts as a physiological determining template in the setup of artificial cardiac tissue. Surface patterns with alternating concave to convex transitions of cell size dimensions were used to organize and orientate human-induced pluripotent stem cell (hIPSC)-derived cardiac myocytes and mouse neonatal cardiac myocytes. The shape of the cells, as well as the organization of the contractile apparatus recapitulates the anisotropic line pattern geometry being derived from tissue geometry motives. The intracellular organization of the contractile apparatus and the cell coupling via gap junctions of cell assemblies growing in a random or organized pattern were examined. Cell spatial and temporal coordinated excitation and contraction has been compared on plain and patterned substrates. While the α-actinin cytoskeletal organization is comparable to terminally-developed native ventricular tissue, connexin-43 expression does not recapitulate gap junction distribution of heart muscle tissue. However, coordinated contractions could be observed. The results of tissue-like cell ensemble organization open new insights into geometry-dependent cell organization, the cultivation of artificial heart tissue from stem cells and the anisotropy-dependent activity of therapeutic compounds.

Cell type

10.3390/jfb7010001

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Citations (15)

An RNA Interference/Adeno‐Associated Virus Vector–Based Combinatorial Gene Therapy Approach Against Hepatitis E Virus

Hepatology Communications (2021)

Cindy Zhang Andrew Freistaedter Carolin Schmelas Manuel Gunkel Viet Loan Dao Thi

Hepatitis E virus (HEV) is a major public health problem with limited therapeutic options. Here, we engineered adeno‐associated viral vectors of serotype 6 (AAV6) to express short hairpin RNAs (shRNAs) against HEV transcripts with the prospect of down‐regulating HEV replication in vivo . We designed 20 different shRNAs, targeting the genome of the HEV genotype 3 (GT3) Kernow‐C1 p6 strain, for delivery upon AAV6 transduction. Using an original selectable HEV GT3 reporter replicon, we identified three shRNAs that efficiently down‐regulated HEV replication. We further confirmed their inhibitory potency with full‐length HEV infection. Seventy‐two hours following transduction, HEV replication in both systems decreased by up to 95%. The three most potent inhibitory shRNAs identified were directed against the methyltransferase domain, the junction region between the open reading frames (ORFs), and the 3´ end of ORF2. Targeting all three regions by multiplexing the shRNAs further enhanced their inhibitory potency over a prolonged period of up to 21 days following transduction. Conclusion: Combining RNA interference and AAV vector–based gene therapy has great potential for suppressing HEV replication. Our strategy to target the viral RNA with multiplexed shRNAs should help to counteract viral escape through mutations. Considering the widely documented safety of AAV vector–based gene therapies, our approach is, in principle, amenable to clinical translation.

Transduction (biophysics)

Replicon

Adeno-associated virus

10.1002/hep4.1842

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Citations (14)

Microscope Image Analysis Course Sept 2109 -- Images siRNA Screen

Zenodo (CERN European Organization for Nuclear Research) (2019)

Manuel Gunkel

Contains 380 Fluorescence images of DAPI stained HeLa nuclei, acquired in 42 wells of a 384 well plate. individual wells were treated with siRNA. Images were acquired with an Olympus ScanR system at 10x magnification.

Course (navigation)

10.5281/zenodo.3358912

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Citations (0)

Quantification of telomere features in tumor tissue sections by an automated 3D imaging-based workflow

bioRxiv (Cold Spring Harbor Laboratory) (2016)

Manuel Gunkel Inn Chung Stefan Wörz Katharina I. Deeg Ronald Simon

Abstract The microscopic analysis of telomere features provides a wealth of information on the mechanism by which tumor cells maintain their unlimited proliferative potential. Accordingly, the analysis of telomeres in tissue sections of patient tumor samples provides can be exploited to obtain diagnostic information and to define tumor subgroups. In many instances, however, analysis of the image data is conducted by manual inspection of 2D images at relatively low resolution for only a small part of the sample. As the telomere feature signal distribution is frequently heterogeneous, this approach is prone to a biased selection of the information present in the image and lacks subcellular details. Here we address these issues by using an automated high-resolution imaging and analysis workflow that quantifies individual telomere features on tissue sections for a large number of cells. The approach is particularly suited to assess telomere heterogeneity and low abundant cellular sub-populations with distinct telomere characteristics in a reproducible manner. It comprises the integration of multi-color fluorescence in situ hybridization, immunofluorescence and DNA staining with targeted automated 3D fluorescence microscopy and image analysis. We apply our method to telomeres in glioblastoma and prostate cancer samples, and describe how the imaging data can be used to derive statistically reliable information on telomere length distribution or colocalization with PML nuclear bodies. We anticipate that relating this approach to clinical outcome data will prove to be valuable for pretherapeutic patient stratification. Abbreviations 3D-TIM 3D targeted imaging ALT alternative lengthening of telomeres APB ALT-associated PML-NB CLSM confocal laser scanning fluorescence microscopy ECTR extrachromosomal telomeric repeat FFPE formalin-fixed, paraffin-embedded FISH fluorescence in situ hybridization IF Immunofluorescence pedGBM pediatric glioblastoma PML promyelocytic leukemia PML-NB PML nuclear body PNA peptide nucleic acid ROI region of interest TMA tissue microarray TMM telomere maintenance mechanism SMLM single molecule localization microscopy

Colocalization

10.1101/053132

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Citations (1)

Correlative Light Microscopy for High-Content Screening

BioTechniques (2013)

Benjamin Flottmann Manuel Gunkel Tautvydas Lisauskas Mike Heilemann Vytaute Starkuviene

High-throughput microscopy is an effective tool for rapidly collecting data on a large scale. However, high throughput comes at the cost of low spatial resolution. Here we introduce correlative light microscopy by combining fast automated widefield imaging, confocal microscopy and super-resolution microscopy. We demonstrate the potential of this approach for scalable experiments. The workflow consists of a robust approach for selecting cells of interest on a wide-field screening microscope at low resolution and subsequently re-localizing those cells with micrometer precision for confocal and super-resolution imaging. As a case study, we visualized and quantified cis- and trans-Golgi markers at increasing resolution.

Correlative

10.2144/000114099

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Citations (23)

Microscope Image Analysis Course Sept 2109 -- Images siRNA Screen

Zenodo (CERN European Organization for Nuclear Research) (2019)

Manuel Gunkel

10.5281/zenodo.3358873

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Citations (0)

Black-Box Hyperparameter Optimization for Nuclei Segmentation in Prostate Tissue Images

Informatik aktuell (2019)

Thomas Wollmann Patrick Bernhard Manuel Gunkel Delia M. Braun Jan Meiners

Hyperparameter

Hyperparameter Optimization

Black box

10.1007/978-3-658-25326-4_75

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Citations (3)

Functional Nuclear Organization of Transcription and DNA Replication: A Topographical Marriage between Chromatin Domains and the Interchromatin Compartment

Cold Spring Harbor Symposia on Quantitative Biology (2010)

Yolanda Markaki Manuel Gunkel Lothar Schermelleh Sven Beichmanis Jürgen Neumann

Y. Markaki1, M. Gunkel2, L. Schermelleh3, S. Beichmanis2, J. Neumann3, M. Heidemann4, H. Leonhardt3,5, D. Eick4,5, C. Cremer2 and T. Cremer⇓1,5 LMU Biocenter, Department of Biology II, Anthropology and Human Genetics, Ludwig Maximilians University (LMU), Martinsried D-82152, Germany; University Heidelberg, Kirchhoff-Institute for Physics and BioQuant Center, Heidelberg D-69120, Germany; LMU Biocenter, Department of Biology II, Epigenetics, Ludwig Maximilian University of Munich (LMU), Martinsried D-82152, Germany; Helmholtz Center Munich, Department of Molecular Epigenetics, Munich D-81377, Germany; Center for Integrated Protein Science, Martinsried D-82152, Germany Correspondence: Thomas.Cremer{at}lrz.uni-muenchen.de

Transcription

RNA polymerase II

Transcription factories

10.1101/sqb.2010.75.042

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Citations (133)

Lokalisationsmikroskopie mit mehreren Farben und ihre Anwendung in biologischen Präparaten

Manuel Gunkel

Fluorescence microscopy is an important method for studying biological structures. By intrinsic properties of the fluorophores it is possible to separate and individually localize their signals. These quantitative position information enable an improved structural resolution and a statistic analysis of the data. In this thesis, localization microscopy was applied for multiple spectral markers within the observed biological samples. Several algorithms to compensate mechanical and chromatic shift and to correct and analyze the position data were developed. For example, the relative distribution of a structural protein of the DNA (H2A) and a DNA remodeler protein (SNF2h) was examined. The experimental results were compared with random distributed signal positions within the same global structures. It could be shown that the real distributions are significantly different from the random ones. It is also possible to check the data for signal clusters and determine their properties. This was first tested on simulated distributions and later applied in the analysis of signal accumulations in biological structures like centromere protein clusters (CENP- A, CENP-B and CENP-C) within the human kinetochore. In addition, the axial position of the localization data within the structure can be determined with a precision of about 50nm. A model for the distribution of the fluorophores in tobacco mosaic virus structures was developed and fitted to the data. The width of these structures determined by electron microscopy to be 18nm could be confirmed. The mean localization accuracy in this case was 8nm. A new microscope setup was build to split the fluorescence signal emitted by the sample due to its different properties like direction of polarization or spectral range. Both parts of the signal could be imaged simultaneously on the same detector. This results in a shorter acquisition time for a two color measurement. Additionally, no mechanical drift between two measurements is apparent.

Position (finance)

SIGNAL (programming language)

Source

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Citations (1)

Hyperparameter optimization for image analysis: application to prostate tissue images and live cell data of virus-infected cells

International Journal of Computer Assisted Radiology and Surgery (2019)

Christian Ritter Thomas Wollmann Patrick Bernhard Manuel Gunkel Delia M. Braun

Hyperparameter

Hyperparameter Optimization

Bayesian Optimization

10.1007/s11548-019-02010-3

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Citations (9)