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Spezialwissen für SCALE
(2024)
Volker Zickermann und Eric Helfrich sind bei der Exzellenzcluster-Initiative SCALE (Subcellular Architecture of Life) dabei und werden dort ihre Expertise einbringen. Das Spezialgebiet des einen ist ein Proteinkomplex in den Mitochondrien, den Kraftwerken der Zelle. Der andere sucht schwerpunktmäßig bisher unbekannte Naturstoffe, die die Basis für neue Antibiotika sein könnten.
Herzforschung meets KI
(2024)
Moderne Methoden der Künstlichen Intelligenz (KI) spielen in der Wissenschaft eine immer größere Rolle. Wie Forscher des Exzellenzclusters Cardio-Pulmonary Institute (CPI) KI in der Herzbildgebung nutzen, zeigte Professor Eike Nagel im Rahmen der Bürgeruniversität der Goethe-Universität. Er leitet das Institut für experimentelle und translationale kardiovaskuläre Bildgebung am Fachbereich Medizin und forscht an der Entwicklung verbesserter Behandlungsmöglichkeiten für Herz-Kreislauf-Erkrankungen. Mit dem Ziel, seine Forschung für alle Menschen zugänglich und verständlicher zu machen, lud Prof. Nagel interessierte Bürger*innen am 10. Mai in sein Institut ein.
Die vorliegende Arbeit behandelt den Vergleich zweier Geräte - „Endotrust MiFusion TLS 2“ und „Medtronic LigaSure Maryland System“ - zur endoskopischen Entnahme der Arteria radialis (RA) zur Verwendung als Bypass-Gefäß in der Herzchirurgie.
Grundsätzlich kommen in der Bypass-Chirurgie zur Herstellung eines Free-Grafts am Herzen neben der Verwendung der Thoraxarterien die Vena Saphena Magna (VSM) sowie die RA in Frage. In den aktuellen europäischen Leitlinien zur Behandlung von hochgradigen Stenosen wird die Verwendung der RA empfohlen („Class 1 Level B“-Empfehlung).
Die Frage, ob die RA für den Einsatz als Bypass-Gefäß offen oder endoskopisch entnommen werden sollte, ist in der Literatur weiterhin umstritten. In den aktuellen Leitlinien zur Behandlung von koronaren Herzkrankheiten wird aufgrund dieser insoweit uneindeutigen Studienlage keine Empfehlung ausgesprochen. Trotzdem ist die endoskopische Entnahme der RA im klinischen Alltag mittlerweile etabliert. Im Kontext dieser uneindeutigen Studienlage einerseits und der praktischen Bedeutung endoskopischer Entnahme andererseits ist es Zielsetzung der vorliegenden Arbeit, durch einen Gerätevergleich einen Beitrag zur Optimierung der endoskopischen Operationstechnik zu leisten. Es soll zudem aufgezeigt werden, inwieweit die endoskopische Entnahme der RA ein sicheres und effizientes Verfahren darstellt.
In der Literatur zum Vergleich von Operationstechniken zur Entnahme von Bypass-Gefäßen werden häufig histologische Untersuchungen angewendet. Diese ermöglichen eine zeitnahe Beurteilung der Qualität des entnommenen Grafts. Das ist auch in dieser Arbeit der wesentliche Grund dafür, dass die histologische Beurteilung der Qualität der RA als primärer Endpunkt angewendet wird. In Anlehnung an die Literatur wurde die strukturelle Integrität des Endothels und der Elastica interna beurteilt. Die histologische Beurteilung erfolgte nach Immunfluoreszenz-Bearbeitung der Proben.
Im Einklang mit der bestehenden Literatur zur Frage, ob die RA offen oder endoskopisch entnommen werden sollte, wurde in dieser Studie als sekundäre Endpunkte Kriterien bezüglich der Sicherheit und Effizienz der Entnahme verwendet. Dies betrifft das Auftreten von intra- und postoperativen Komplikationen, insbesondere im Hinblick auf neurologische Beeinträchtigungen am operierten Arm, sowie die Entnahmedauer und den operativen Aufwand zur Blutstillung.
Die in dieser Arbeit behandelte Studie wurde als prospektive, 1:1 randomisierte Studie mit zwei Gruppen mit jeweils 50 Patienten durchgeführt. Alle Operationen erfolgten im Zeitraum Januar 2017 bis Juli 2017 im Herzzentrum der Kerckhoff-Klinik Bad Nauheim.
Bezüglich der Resultate des Gerätevergleichs zeigte sich ein eindeutiges Ergebnis. Sämtliche Beurteilungskriterien, bei denen signifikante Unterschiede zwischen den beiden Gruppen aufgetreten sind, u.a. Integrität der Elastica interna, Entnahmedauer, Vorkommen von Residualblutungen, Auftreten von sensorischen Störungen fallen zugunsten des LigaSure-Systems aus.
Dabei ist unserer Meinung nach der wichtigste Einzelaspekt, dass die histologisch ermittelte Integrität der Elastica interna als Indikator für die Qualität des entnommen Grafts in der LigaSure-Gruppe signifikant besser war als in der MiFusion-Gruppe. Dagegen konnten aus der histologischen Untersuchung des Endothels keine klaren Rückschlüsse gezogen werden. Insofern besteht Unsicherheit, ob die Schädigung der Endothelschicht durch die Entnahme selbst oder durch die anschließende Präparierung der entnommenen Proben verursacht wurde.
Bezüglich der sekundären Endpunkte zeigten sich für die Mifusion-Patientengruppe im Vergleich zu anderen Studien zur endoskopischen Entnahme der RA zufriedenstellende bis gute und für die LigaSure Gruppe gute bis sehr gute Ergebnisse. Unabhängig vom verwendeten Gerät kann diese Studie deshalb als eine Bestätigung bisheriger Studien zur Vorteilhaftigkeit der endoskopischen RA-Entnahme angesehen werden. Letztlich fehlt jedoch weiterhin der Nachweis, dass eine endoskopische Entnahme unbedenklich im Hinblick auf den langfristigen kardiologischen Outcome ist. Dies bleibt zukünftiger Forschung vorbehalten.
Darüber hinaus trägt die Studie dazu bei, das klinische Erfahrungswissen über operationstechnische Details bei der Entnahme der Radialarterie zu erweitern und somit die Akzeptanz für die Verwendung der Radialarterie als Bypass-Gefäß in der koronaren Herzchirurgie zu verbessern.
Das Glioblastom ist eine tödliche maligne Erkrankung des zentralen Nervensystems. Etablierte Therapiekonzepte resultieren in einer Fünfjahresüberlebensrate von fünf Prozent. Die derart infauste Prognose wird unter anderem bedingt durch die Heterogenität des Tumors. Insbesondere einer Population stammzellartiger Zellen wird die Verantwortung für Resistenz und Rekurrenz des Glioblastoms zugesprochen. Die genuine Plastizität des Glioblastoms mit entsprechender Fähigkeit zur Änderungen tumorweiter Expressionsprofile und Ausbildung einzigartiger funktioneller Fähigkeiten kann ohne gezielte Beeinträchtigung von stammzellartigen Zellen womöglich nicht ausreichend überwunden werden. Als Urheber kritischer Eigenschaften erscheint die erfolgreiche Elimination dieser Population innerhalb des Glioblastoms notwendig um nachhaltige Therapieerfolge zu erzielen. Mögliche Strategien der Elimination stammzellartiger Zellen setzen an Differenzierung und Ausbeutung stammzelltypischer Signalwege zur Modulation dieser Zellen an. Hierdurch sollen zentrale Fähigkeiten der Population stammzellartiger Zellen, wie Selbsterneuerung, Resistenz gegenüber Strahlen- und Chemotherapie und erneute Formation heterogener Tumore, überwunden werden.
Zentrale zelluläre Prozesse, welche zum Erhalt des stammzellartigen Zustandes dieser Zellen beitragen, sind unter anderem der Hedgehog- und Notch-Signalweg. Einer Beeinträchtigung dieser Signalwege wohnt womöglich die Fähigkeit der effektiven Modulation zentraler Eigenschaften stammzellartiger Zellen inne. Neben diesen Signalwegen gibt es eine Reihe weiterer Prozesse, welchen eine Urheberschaft an der Resistenz der Zellen zugesprochen wird. Hierzu zählt beispielweise der Prozess der Autophagie. Die Autophagie ist ein hochkonservierter zellulärer Mechanismus zur Selbsterneuerung durch Selbstdegradation fehlerhafter zellulärer Komponenten. Gleichzeitig kann die Autophagie durch eine Überaktivität zu einem spezifischen, autophagischen Zelltod beitragen. Die Modulation dieses Dualismus kann in einer Vielzahl von Tumoren, so auch im Glioblastom, das Schicksal einer tumorfördernden Autophagie in eine antitumorale Autophagie ändern.
Im Rahmen dieser Arbeit wurde erstmalig eine Modulation zentraler Eigenschaften stammzellartiger Zellen durch die Beeinflussung ihrer zellulären Prozesse mittels kombinierter Therapie durch Arsentrioxid oder GANT und (-)-Gossypol gezeigt. Arsentrioxid wirkt unspezifisch unter anderem als Inhibitor von Notch- und Hedgehog-Signalweg. Diese Inhibition wurde auch in den untersuchten Zellen nachgewiesen und führte zu einer Reduktion von stammzelltypischen Markerproteinen und Fähigkeiten der Tumorgenese in -vitro und ex -vivo, sowie zur Sensitivierung gegenüber strahleninduzierten Schäden. Gegenüber einer spezifischen Hedgehog-Inhibition durch eine GANT-vermittelte Bindung an Gli-Transkriptionsfaktoren zeigten sich deutliche Vorteile der dualen Inhibition durch Arsentrioxid hinsichtlich der genannten Eigenschaften. Die Kombination der Substanzen mit dem pan-Bcl-Inhibitor (-)-Gossypol führte zu einer synergistischen Steigerung der antitumoralen Effekte. (-)-Gossypol wird in Gliomzellen insbesondere mit der Modulation der autophagischen Maschinerie und Auslösung eines autophagischen Zelltodes in Verbindung gebracht. Die Ergebnisse weisen parallele Signalweginteraktionen mit effektiver Modulation des DNA-Damage-Response-Systems durch die Reduktion des Proteins CHEK als kausalen Mechanismus des Synergismus der Substanzen aus.
Die beobachteten Änderungen der typischen Eigenschaften stammzellartiger Zellen durch die Therapie mit Arsentrioxid und (-)-Gossypol implizieren lohnende Folgeuntersuchungen zur weiteren Evaluation dieser Effekte in -vivo, um zukünftig translationale Ableitungen zu erlauben. Die Heterogenität des Glioblastoms und seine genuine Plastizität lassen sich womöglich erfolgreich durch multiple Eingriffe in unterschiedliche zelluläre Prozesse, hierunter Notch- und Hedgehog-Signaling, modulieren. Hierdurch könnten zentrale Eigenschaften des Glioblastoms eventuell effektiv verändert und Resistenz sowie Rekurrenz überwunden werden.
Motivation DNA CpG methylation (CpGm) has proven to be a crucial epigenetic factor in the gene regulatory system. Assessment of DNA CpG methylation values via whole-genome bisulfite sequencing (WGBS) is, however, computationally extremely demanding.
Results We present FAst MEthylation calling (FAME), the first approach to quantify CpGm values directly from bulk or single-cell WGBS reads without intermediate output files. FAME is very fast but as accurate as standard methods, which first produce BS alignment files before computing CpGm values. We present experiments on bulk and single-cell bisulfite datasets in which we show that data analysis can be significantly sped-up and help addressing the current WGBS analysis bottleneck for large-scale datasets without compromising accuracy.
Availability An implementation of FAME is open source and licensed under GPL-3.0 at https://github.com/FischerJo/FAME.
Complexome profiling (CP) is a powerful tool for systematic investigation of protein interactors that has been primarily applied to study the composition and dynamics of mitochondrial protein complexes. Here, we further optimised this method to extend its application to survey mitochondrial DNA- and RNA-interacting protein complexes. We established that high-resolution clear native gel electrophoresis (hrCNE) is a better alternative to preserve DNA- and RNA-protein interactions that are otherwise disrupted when samples are separated by the widely used blue native gel electrophoresis (BNE). In combination with enzymatic digestion of DNA, our CP approach improved the identification of a wide range of protein interactors of the mitochondrial gene expression system without compromising the detection of other multi-protein complexes. The utility of this approach was particularly demonstrated by analysing the complexome changes in human mitochondria with impaired gene expression after transient, chemically-induced mtDNA depletion. Effects of RNase on mitochondrial protein complexes were also evaluated and discussed. Overall, our adaptations significantly improved the identification of mitochondrial DNA- and RNA-protein interactions by CP, thereby unlocking the comprehensive analysis of a near-complete mitochondrial complexome in a single experiment.
Einleitung: Frailty (engl. für Gebrechlichkeit) bezeichnet eine mit hohem Alter zunehmende Verschlechterung des körperlichen und kognitiven Zustandes von Individuen, woraufhin der Körper nicht mehr in der Lage ist, adäquat auf äußere und innere Stressoren zu reagieren. Frailty ist mit einer erhöhten Morbiditäts- und Mortalitätsrate sowie längerer Krankenhausverweildauer und erhöhter postoperativer Komplikationsrate verbunden und stellt folglich einen chirurgischen Risikofaktor dar.
Problemstellung: Die Relevanz eines strukturierten Frailty Assessments in der präoperativen Risikostratifizierung führte zur Indikationsstellung, diverse validierte Risk Assessment Tools auf ihre prädiktive Vorhersagekraft bezüglich des Auftretens von postoperativen Komplikationen und postoperativer Sterbewahrscheinlichkeit zu untersuchen.
Methoden: In die vorliegende Studie wurden Patienten, die in dem Zeitraum vom 01.09.2018 und 31.01.2019 in der allgemeinchirurgischen Ambulanz vorstellig waren und einen allgemeinchirurgischen Eingriff erhielten, aufgenommen. Mittels Fragebögen wurden die Scores „Risk Analysis Index“, „Edmonton Frail Scale“ sowie „Charlson Comorbidity Index“ präoperativ erhoben und retrospektiv mit Daten aus der digitalen Patientenakte zusammengeführt. Endpunkte waren die 90-Tages- Mortalität sowie das Auftreten von schweren postoperativen Komplikationen ab Clavien Dindo Grad 3b. Die Analyse erfolgte in SPSS mittels Chi-Quadrat Test, t- Test und ROC-Kurven Analysen.
Ergebnisse: Das durchschnittliche Alter der Studienkohorte lag bei 56 ± 15.9 Jahren und der Anteil männlicher Patienten überwog mit 59.2% (n=282).
Die Fragebögen wurden 739 Patienten vorlegt und 476 Patienten konnten in die Datenanalyse eingeschlossen werden. Die 90-Tages-Mortalität lag bei 2.7% (n=13) und 9% (n=43) erlitten schwere postoperative Komplikationen ab Clavien-Dindo Grad IIIb. Die Einteilung nach der ASA-Klassifikation (p=0.024), maligne Diagnosen -7-(p<0.001) und Majorkomplikationen (p<0.001) stellten präoperative Risikofaktoren für postoperative 90-Tage-Mortalität dar. Von den Risk Assessment Scores zeigte lediglich der Risk Analysis Index eine signifikante Korrelation auf (p=0.013). Ein mittels ROC-Analyse ermittelter Cut-Off Wert von 23 klassifizierte 166 (34.9%) Patienten als frail, die mit 69% Sensitivität und 66% Spezifität (AUC=0.735) ein erhöhtes Risiko für postoperatives Versterben innerhalb von 90 Tagen aufwiesen (p=0.008). Risikofaktoren für das Auftreten schwerer postoperativer Komplikationen waren die ASA-Klassifikation (p=0.041), längere Krankenhausverweildauer (p<0.001) und maligne Diagnosen (p<0.001). Der Charlson Comorbidity Index (p=0.031) und RAI-C Werte ≥ 23 (p<0.001) korrelierten signifikant mit Majorkomplikationen. Das Alter ab 65 Jahren stellte mit 77 % Spezifität und 69 % Sensitivität ebenfalls einen prädiktiven Risikofaktor für postoperative Mortalität dar (AUC=0,787).
Schlussfolgerung: Mithilfe validierter Risk Assessment Tools ist es möglich Patienten, die ein erhöhtes Risiko für postoperative negative Ereignisse aufweisen, bereits präoperativ zu erkennen. Dies ermöglicht eine bessere Beurteilung der chirurgischen Indikationsstellung sowie das rechtzeitige Ergreifen von risikominimierenden Maßnahmen. Es ist notwendig die Ergebnisse dieser Arbeit künftig mit risikominimierenden Maßnahmen zu verknüpfen und zu untersuchen, ob die Implementierung der Risk Assessment Tools zu verbesserten postoperativen Ergebnissen führt, wenn modifizierbare Faktoren verbessert werden.
Endothelial tip cells are essential for VEGF-induced angiogenesis, but underlying mechanisms are elusive. Endothelial-specific deletion of EVL, a member of the mammalian Ena/VASP protein family, reduced the expression of the tip cell marker protein endothelial cell specific molecule-1 (Esm1) and compromised the radial sprouting of the vascular plexus in the postnatal mouse retina. The latter effects could at least partly be attributed to reduced VEGF receptor 2 (VEGFR2) internalization and signaling but the underlying mechanisms(s) are not fully understood. In the present study, we revealed that the expression of the long non-coding RNA H19 was significantly reduced in endothelial cells from postnatal EVL-/- mice and in siRNA-transfected human endothelial cells under hypoxic conditions. H19 was recently shown to promote VEGF expression and bioavailability via Esm1 and hypoxia inducible factor 1α (HIF-1α). Similar to EVL-/- mice, the radial outgrowth of the vascular plexus was significantly delayed in the postnatal retina of H19-/- mice. In summary, our data suggests that loss of EVL not only impairs VEGFR2 internalition and downstream signaling, but also impairs VEGF expression and bioavailability in the hypoxic retina via downregulation of lncRNA H19.
Ernsting, Jan ; Holstein, Vincent Leonard ; Winter, Nils Ralf ; Sarink, Kelvin ; Leenings, Ramona ; Gruber, Marius ; Repple, Jonathan ; Risse, Benjamin ; Dannlowski, Udo ; Hahn, Tim
Graph data is an omnipresent way to represent information in machine learning. Especially, in neuroscience research, data from Diffusion-Tensor Imaging (DTI) and functional Magnetic Resonance Imaging (fMRI) is commonly represented as graphs. Exploiting the graph structure of these modalities using graph-specific machine learning applications is currently hampered by the lack of easy-to-use software. PHOTONAI Graph aims to close the gap between domain experts of machine learning, graph experts and neuroscientists. Leveraging the rapid machine learning model development features of the Python machine learning API PHOTONAI, PHOTONAI Graph enables the design, optimization, and evaluation of reliable graph machine learning models for practitioners. As such, it provides easy access to custom graph machine learning pipelines including, hyperparameter optimization and algorithm evaluation ensuring reproducibility and valid performance estimates. Integrating established algorithms such as graph neural networks, graph embeddings and graph kernels, it allows researchers without significant coding experience to build and optimize complex graph machine learning models within a few lines of code. We showcase the versatility of this toolbox by building pipelines for both resting–state fMRI and DTI data in the hope that it will increase the adoption of graph-specific machine learning algorithms in neuroscience research.
Bilateral simultaneous cochlear implantation is a safe method of hearing rehabilitation in adults
(2023)
Purpose: Bilateral cochlear implantation is an effective treatment for patients with bilateral profound hearing loss. In contrast to children, adults mostly choose a sequential surgery. This study addresses whether simultaneous bilateral CI is associated with higher rates of complications compared to sequential implantation.
Methods: 169 bilateral CI surgeries were analyzed retrospectively. 34 of the patients were implanted simultaneously (group 1), whereas 135 patients were implanted sequentially (group 2). The duration of surgery, the incidence of minor and major complications and the duration of hospitalization of both groups were compared.
Results: In group 1, the total operating room time was significantly shorter. The incidences of minor and major surgical complications showed no statistically significant differences. A fatal non-surgical complication in group 1 was particularly extensively reappraised without evidence of a causal relationship to the chosen mode of care. The duration of hospitalization was 0.7 days longer than in unilateral implantation but 2.8 days shorter than the combined two hospital stays in group 2.
Conclusion: In the synopsis of all considered complications and complication-relevant factors, equivalence of simultaneous and sequential cochlear implantation in adults in terms of safety was found. However, potential side effects related to longer surgical time in simultaneous surgery must be considered individually. Careful patient selection with special consideration to existing comorbidities and preoperative anesthesiologic evaluation is essential.
Pérez Berrocal, David Aureliano ; Vishwanatha, Thimmalapura M. ; Horn-Ghetko, Daniel ; Botsch, J. Josephine ; Hehl, Laura A. ; Kostrhon, Sebastian ; Misra, Mohit ; Ðikić, Ivan ; Geurink, Paul P. ; Dam, Hans van ; Schulman, Brenda A. ; Mulder, Monique P. C.
Protein post-translational modification with ubiquitin (Ub) is a versatile signal regulating almost all aspects of cell biology, and an increasing range of diseases is associated with impaired Ub modification. In this light, the Ub system offers an attractive, yet underexplored route to the development of novel targeted treatments. A promising strategy for small molecule intervention is posed by the final components of the enzymatic ubiquitination cascade, E3 ligases, as they determine the specificity of the protein ubiquitination pathway. Here, we present UbSRhodol, an autoimmolative Ub-based probe, which upon E3 processing liberates the pro-fluorescent dye, amenable to profile the E3 transthiolation activity for recombinant and in cell-extract E3 ligases. UbSRhodol enabled detection of changes in transthiolation efficacy evoked by enzyme key point mutations or conformational changes, and offers an excellent assay reagent amenable to a high-throughput screening setup allowing the identification of small molecules modulating E3 activity.
Hintergrund: Epilepsie bezeichnet eine Erkrankung, welche durch eine anhaltende Prädisposition für Symptome, die im Zusammenhang mit einer ungewöhnlich starken oder synchronisierten elektrischen Aktivität des Gehirns auftreten (=epileptischer Anfall) charakterisiert ist. Mutationen in dem Gen DEPDC5 (Dishevelled, Egl-10 and Pleckstrin (DEP) domain-containing protein 5) sind mit fokalen Epilepsien assoziiert und führen in Tiermodellen und humanen Modellen zu einer Überaktivierung des mTOR-Signalweges. Auf neuromorphologischer Ebene zeigt sich die mTOR-Überaktivierung durch eine Vergrößerung des Zelldurchmessers und einer zunehmenden Verästelung der Neuriten. Ziel dieser Studie war es, die morphologischen Auswirkungen der DEPDC5-assoziierten mTOR-Überaktivierung in der SH-SY5Y-Neuroblastomzelllinie zu untersuchen. Dadurch soll eine Einschätzung getroffen werden, ob das im Zusammenhang mit neurodegenerativen Erkrankungen bereits gut etablierte SH-SY5Y-Zellmodell auch bei der Untersuchung epilepsieassoziierter Pathomechanismen zum Einsatz kommen kann.
Methoden: Unter Einsatz der CRISPR/Cas9-Methode wurden Knockout(KO)-Mutationen in Exon 2 und Exon 3 des DEPDC5-Gens erzeugt und diese mittels Sanger-Sequenzierung bestätigt. Danach wurden die Knockouts auf RNA- und Proteinebene, durch Real-time-RT-PCR und Western Blot validiert. Die bestätigten homozygoten DEPDC5-KO-Zelllinen wurden anschließend mittels Western Blot auf eine mTOR-Überaktivierung untersucht. Zuletzt erfolgte die neuromorphologische Validierung des DEPDC5-KO. Die Zellgröße proliferierender SH-SY5Y wurden mittels Durchflusszytometrie (FACS) untersucht. Zudem bestimmten wir die neuronale Architektur differenzierter SH-SY5Y unter Einsatz der Sholl-Analyse.
Ergebnisse: Es konnten vier unabhängige DEPDC5-KO-SH-SY5Y-Zelllinien mit homozygoten Indel-Mutationen und vorzeitigem Stoppcodon in Exon 3 generiert werden. Die erwartete Reduktion an DEPDC5-mRNA konnte mittels Real-time 10 RT-PCR nicht festgestellt werden. Die Abwesenheit des Proteins konnte durch Western Blot aber gezeigt werden. Funktionell konnte für alle Zelllinien eine mTOR-Überaktivierung mittels Western Blot nachgewiesen werden. Dabei konnte phosphoryliertes AKT (AKT serine/threonine kinase 1) als stabilster Marker etabliert werden. Auf neuromorphologischer Ebene ließ sich ein Trend in Richtung vergrößertem Zelldurchmesser bei verlängertem Auswachsen der Neuriten feststellen, wobei sich für das Modell Unterschiede zwischen den einzelnen Klonen ergaben.
Diskussion: In dieser Studie gelang es erstmals, den Zusammenhang zwischen DEPDC5-KO und einer mTOR-Überaktivierung in der onkogenen SH-SY5Y-Zelllinie zu replizieren. Das verlängerte Auswachsen der Neuriten, bei jedoch gleichbleibender Anzahl peripherer Verästelungen, stellt dabei einen neuen Befund dar und könnte durch die frühe neuronale Entwicklungsstufe des SH-SY5Y-Zellmodells erklärt werden. Auf Grundlage der Ergebnisse dieser Arbeit lässt sich sagen, dass das robuste und kostengünstige SH-SY5Y-Zellmodell insbesondere für high-throughput Methoden und Screeningassays ein geeignetes Modell ist. Durch die Kombination mit reiferen Zellmodellen, wie beispielsweise iPSCs (induced pluripotent stem cells), könnte der Phänotyp eines DEPDC5-KO und anderer mTOR-assoziierter Epilepsien, möglichst umfassend in-vitro dargestellt werden.
Kennedy, Eamonn ; Vadlamani, Shashank ; Lindsey, Hannah M. ; Lei, Pui-Wa ; Jo-Pugh, Mary ; Adamson, Maheen ; Alda, Martin ; Alonso-Lana, Silvia ; Ambrogi, Sonia ; Anderson, Tim J. ; Arango, Celso ; Asarnow, Robert F. ; Avram, Mihai ; Ayesa-Arriola, Rosa ; Babikian, Talin ; Banaj, Nerisa ; Bird, Laura J. ; Borgwardt, Stefan ; Brodtmann, Amy ; Brosch, Katharina ; Caeyenberghs, Karen ; Calhoun, Vince D. ; Chiaravalloti, Nancy D. ; Cifu, David X. ; Crespo-Facorro, Benedicto ; Dalrymple-Alford, John C. ; Dams-O’Connor, Kristen ; Dannlowski, Udo ; Darby, David ; Davenport, Nicholas ; DeLuca, John ; Díaz-Caneja, Covadonga M. ; Disner, Seth G. ; Dobryakova, Ekaterina ; Ehrlich, Stefan ; Esopenko, Carrie ; Ferrarelli, Fabio ; Frank, Lea E. ; Franz, Carol ; Fuentes-Claramonte, Paola ; Genova, Helen ; Giza, Christopher C. ; Goltermann, Janik ; Grotegerd, Dominik ; Gruber, Marius ; Gutiérrez-Zotes, Alfonso ; Ha, Minji ; Haavik, Jan ; Hinkin, Charles ; Hoskinson, Kristen R. ; Hubl, Daniela ; Irimia, Andrei ; Jansen, Andreas ; Kaess, Michael ; Kang, Xiaojian ; Kenney, Kimbra ; Keřková, Barbora ; Khlif, Mohamed Salah ; Kim, Minah ; Kindler, Jochen ; Kircher, Tilo ; Knı́žková, Karolina ; Kolskår, Knut K. ; Krch, Denise ; Kremen, William S. ; Kuhn, Taylor ; Kumari, Veena ; Kwon, Jun Soo ; Langella, Roberto ; Laskowitz, Sarah ; Lee, Jungha ; Lengenfelder, Jean ; Liebel, Spencer W. ; Liou-Johnson, Victoria ; Lippa, Sara M. ; Løvstad, Marianne ; Lundervold, Astri ; Marotta, Cassandra ; Marquardt, Craig A. ; Mattos, Paulo ; Mayeli, Ahmad ; McDonald, Carrie R. ; Meinert, Susanne ; Melzer, Tracy R. ; Merchán Naranjo, Jessica ; Michel, Chantal ; Morey, Rajendra A. ; Mwangi, Benson ; Myall, Daniel J. ; Nenadić, Igor ; Newsome, Mary R. ; Nunes, Abraham ; O’Brien, Terence J. ; Oertel, Viola ; Ollinger, John ; Olsen, Alexander ; Ortiz Garcı́a de la Foz, Victor ; Ozmen, Mustafa ; Pardoe, Heath ; Parent, Marise ; Piras, Fabrizio ; Piras, Federica ; Pomarol-Clotet, Edith ; Repple, Jonathan ; Richard, Geneviève ; Rodriguez, Jonathan ; Rodriguez, Mabel ; Rootes-Murdy, Kelly ; Rowland, Jared ; Ryan, Nicholas P. ; Salvador, Raymond ; Sanders, Anne-Marthe ; Schmidt, André ; Soares, Jair C. ; Spalleta, Gianfranco ; Španiel, Filip ; Stasenko, Alena ; Stein, Frederike ; Straube, Benjamin ; Thames, April ; Thomas-Odenthal, Florian ; Thomopoulos, Sophia I. ; Tone, Erin ; Torres, Ivan ; Troyanskaya, Maya ; Turner, Jessica A. ; Ulrichsen, Kristine M. ; Umpierrez, Guillermo ; Vilella, Elisabet ; Vivash, Lucy ; Walker, William C. ; Werden, Emilio ; Westlye, Lars T. ; Wild, Krista ; Wroblewski, Adrian ; Wu, Mon-Ju ; Wylie, Glenn R. ; Yatham, Lakshmi N. ; Zunta-Soares, Giovana B. ; Thompson, Paul M. ; Tate, David F. ; Hillary, Frank G. ; Dennis, Emily L. ; Wilde, Elisabeth A.
Investigators in the cognitive neurosciences have turned to Big Data to address persistent replication and reliability issues by increasing sample sizes, statistical power, and representativeness of data. While there is tremendous potential to advance science through open data sharing, these efforts unveil a host of new questions about how to integrate data arising from distinct sources and instruments. We focus on the most frequently assessed area of cognition - memory testing - and demonstrate a process for reliable data harmonization across three common measures. We aggregated raw data from 53 studies from around the world which measured at least one of three distinct verbal learning tasks, totaling N = 10,505 healthy and brain-injured individuals. A mega analysis was conducted using empirical bayes harmonization to isolate and remove site effects, followed by linear models which adjusted for common covariates. After corrections, a continuous item response theory (IRT) model estimated each individual subject’s latent verbal learning ability while accounting for item difficulties. Harmonization significantly reduced inter-site variance by 37% while preserving covariate effects. The effects of age, sex, and education on scores were found to be highly consistent across memory tests. IRT methods for equating scores across AVLTs agreed with held-out data of dually-administered tests, and these tools are made available for free online. This work demonstrates that large-scale data sharing and harmonization initiatives can offer opportunities to address reproducibility and integration challenges across the behavioral sciences.
Pericytes are capillary-associated mural cells involved in the maintenance and the stability of the vascular network. This thesis aims to investigate the role of pericytes in the heart in the context of ageing and disease. We highlight the malignant effects of the remodelling in the heart and stress the focus on the role of cardiac pericytes in this context. We show that ageing reduces pericyte coverage and that myocardial infarction (MI) causes an activation of these cells. Single-nuclei and single-cell RNA sequencing analysis of murine hearts further revealed that the expression of the Regulator of G-protein signalling 5 (Rgs5) is reduced in cardiac pericytes both in ageing and transiently at day 1 and day 3 after MI. The loss of RGS5 in pericytes drives an entropic state of these mural cells characterized by morphological changes, excessive extracellular deposition and enhanced Gaq mediated GPCR signalling. The deletion of RGS5 in pericytes causes cardiac systolic dysfunction, induces myocardial fibrosis, and drives the activation of cardiac fibroblasts in a TGFb-dependent manner. In conclusion, our results describe the importance of pericytes maintaining cardiac homeostasis, identify RGS5 as a key regulator of this process and propose pericytes as crucial mediators of cardiac fibrosis and possible therapeutic targets to prevent cardiovascular disease.
Das Harnblasenkarzinom ist einer der häufigsten Tumoren weltweit. Insbesondere die muskelinvasiven Tumoren haben eine schlechte Prognose und stellen sich morphologisch sehr unterschiedlich dar. Diese Heterogenität wird bislang bei Therapieentscheidungen nicht beachtet. Um Patienten zukünftig individuell auf den vorliegenden Subtyp des muskelinvasiven Harnblasenkarzinoms (MIBC, muscle invasive bladder cancer) behandeln zu können und dadurch unnötige Belastungen durch Chemotherapien vermeiden zu können, ist eine einfache und kostengünstige Diagnostik erforderlich. Das Ziel der vorliegenden Arbeit war es, den Einsatz bestimmter immunhistochemischer Färbungen als ein mögliches diagnostisches Routineverfahren zur Bestimmung der vorliegenden Subtypen auszutesten. Hierzu wurde die Expression von „luminalen“ und „basalen“ Proteinen mit histologischen Subtypen des MIBCs korreliert. In einem zweiten Schritt wurde der Einfluss auf das Überleben mit und ohne adjuvante Chemotherapie untersucht.
Es wurden insgesamt 181 Tissue-Microarray-Spots analysiert. Alle histologischen Patientenproben sowie klinischen Daten stammten aus dem Universitätsklinikum Frankfurt am Main. Aus den entsprechenden Gewebeblöcken wurden Stanzen zur Erstellung eines Tissue-Microarrays (TMAs) entnommen. Diese wurden konventionell angeschnitten und histologisch mit Hämatoxylin-Eosin (H/E) sowie immunhistochemisch mit Cytokeratin 5/6 (CK5/6), Cytokeratin 20 (CK20), Glutamyl Aminotransferase-Untereinheit A bindendem Protein 3 (GATA3), Tumorsuppressor- protein p53 und Synaptophysin (SYNAPT) gefärbt.
Anhand der H/E-Schnitte wurden die vorliegenden histologischen Subtypen lichtmikroskopisch bestimmt und es folgte eine statistische Auswertung der Färbeergebnisse. Die deskriptive statistische Analyse zeigte insbesondere für die beiden Färbungen CK5/6 und GATA3 signifikante Ergebnisse. Deshalb wurden in einem zweiten Schritt alle Fälle der Studienkohorte den bekannten vier Gruppen: CK5/6 positiv, GATA3 positiv, doppelt positiv und doppelt negativ zugeordnet und näher untersucht. Es folgte eine Überlebensanalyse nach Kaplan Meier Schätzer sowie uni- und multivariate Analysen.
Die Ergebnisse zeigten eine Assoziation der Expression von CK5/6 mit einer squamösen Differenzierung (96%) und eine Assoziation der Expression von GATA3 mit einer mikropapillären Differenzierung (100%). Die adjuvante Chemotherapie ging mit einem Überlebensvorteil (HR 0,15 95%KI 0,1-0,3; p<0,001) der Patienten mit MIBC einher. Immunhistochemisch doppelt negative Patienten mit MIBC wiesen ein verringertes Gesamtüberleben auf (HR 4,96; 95%KI 1,6-15,6; p=0,006); in der Gruppe der doppelt negativen MIBCs fanden sich fünf verschiedene histologische Subtypen. Zusammenfassend lässt sich sagen, dass die immunhistochemische Klassifizierung des MIBCs mit histologischen Subtypen assoziiert ist und dabei helfen kann, Fälle in der pathologischen Routine in „luminal“ und „basal“ zu unterteilen. Jedoch ist ein auf zwei Markern basierendes Klassifizierungssystem nicht ausreichend, um die Heterogenität des MIBCs abzubilden und die Basis für Therapieentscheidungen zu bilden.