Extinction of gene expression in somatic cell hybrids. a reflection of important regulatory mechanisms?

Extinction in somatic cell hybrids is a multifactorial process that leads to loss of cell-type-specific gene expression. The underlying mechanisms are thought to mirror, at least in part, the repertoire of regulatory mechanisms controlling mammalian cell differentiation

Extinction of tyrosine aminotransferase gene activity in somatic cell hybrids involves modification and loss of several essential transcription factors

Extinction is defined as the loss of cell type-specific gene expression that occurs in somatic cell hybrids derived by fusion of cells with dissimilar phenotypes. To explore the basis of this dominant-negative regulation, we have studied the activities of the control elements of the liver-specific gene encoding tyrosine aminotransferase (TAT) in hepatoma/fibroblast hybrid crosses. We show that extinction in complete somatic cell hybrids is accompanied by the loss of activity of all known cell type-specific control elements of the TAT gene. This inactivity is the result of first, lack of expression of genes coding for the transcriptional activators HNF4 and HNF3[~ and HNF33,, which bind to essential elements of the enhancers; and second, loss of in vivo binding and activity of ubiquitous factors to these enhancers, including CREB, which is the target for repression by the tissue-specific extinguisher locus TSE1. Complete extinction of TAT gene activity is therefore a multifactorial process affecting all three enhancers controlling liver-specific and hormone-inducible expression. It results from lack of activation, rather than active repression, and involves both post-translational modification and loss of essential transcriptional activators

Adjoint-Based Error Estimation and Mesh Adaptation for Hybridized Discontinuous Galerkin Methods

We present a robust and efficient target-based mesh adaptation methodology, building on hybridized discontinuous Galerkin schemes for (nonlinear) convection-diffusion problems, including the compressible Euler and Navier-Stokes equations. Hybridization of finite element discretizations has the main advantage, that the resulting set of algebraic equations has globally coupled degrees of freedom only on the skeleton of the computational mesh. Consequently, solving for these degrees of freedom involves the solution of a potentially much smaller system. This not only reduces storage requirements, but also allows for a faster solution with iterative solvers. The mesh adaptation is driven by an error estimate obtained via a discrete adjoint approach. Furthermore, the computed target functional can be corrected with this error estimate to obtain an even more accurate value. The aim of this paper is twofold: Firstly, to show the superiority of adjoint-based mesh adaptation over uniform and residual-based mesh refinement, and secondly to investigate the efficiency of the global error estimate

A Comparison of Hybridized and Standard DG Methods for Target-Based hp-Adaptive Simulation of Compressible Flow

We present a comparison between hybridized and non-hybridized discontinuous Galerkin methods in the context of target-based hp-adaptation for compressible flow problems. The aim is to provide a critical assessment of the computational efficiency of hybridized DG methods. Hybridization of finite element discretizations has the main advantage, that the resulting set of algebraic equations has globally coupled degrees of freedom only on the skeleton of the computational mesh. Consequently, solving for these degrees of freedom involves the solution of a potentially much smaller system. This not only reduces storage requirements, but also allows for a faster solution with iterative solvers. Using a discrete-adjoint approach, sensitivities with respect to output functionals are computed to drive the adaptation. From the error distribution given by the adjoint-based error estimator, h- or p-refinement is chosen based on the smoothness of the solution which can be quantified by properly-chosen smoothness indicators. Numerical results are shown for subsonic, transonic, and supersonic flow around the NACA0012 airfoil. hp-adaptation proves to be superior to pure h-adaptation if discontinuous or singular flow features are involved. In all cases, a higher polynomial degree turns out to be beneficial. We show that for polynomial degree of approximation p=2 and higher, and for a broad range of test cases, HDG performs better than DG in terms of runtime and memory requirements

Role of cyclic AMP in the control of cell-specific gene expression

Genes have to be expressed in specific cell types at appropriate times of development dependent on external signals. cAMP signaling occurs in all cells, thus raising the question of how this signal transduction pattern is integrated into mechanisms determining cell-specific gene expression. We have analyzed expression of the tyrosine aminotransferase gene as a model to study the basis of this cell type specificity of hormone induction. We found that cell-type-specific expression is generated by combined action of cAMP signal-dependent and liver cell-specific transcription factors. The interdependence of the cAMP response element and an element determining liver cell specificity enables a gene to respond to an ubiquitous signal in a cell-specific manner

meta-analytical evaluation of coronary CT angiography studies

Zielsetzung: Ziel der Arbeit ist es, zu bestimmen, ob die Benutzung einer Sechsfeldertafel mit einer Intention-to-Diagnose Herangehensweise sich zur transparenten Darstellung und zum Umgang mit nichtbeurteilbaren Ergebnissen bei der Beurteilung der diagnostischen Genauigkeit eines Testverfahrens besser eignet, als die Benutzung einer "klassischen" Vierfeldertafel. Studiendesign: Basierend auf einer systematischen Suche nach diagnostischen Genauigkeitsstudien zur CT Koronarangiografie wurden Volltexte relevanter Studien dahingehend untersucht, ob alternativ eine Sechsfeldertafel berechnet werden konnte. Um einen Gesamteffekt zu quantifizieren, wurden die diagnostischen Genauigkeitswerte gemäß einer metaanalytischen Methodik statistisch zusammengefasst (gepoolt). Datenquellen: Gesucht wurde in den elektronische Datenbanken Medline (via PubMed), Embase (via Ovid) und ISI Web of Science. Eignungskriterien: Prospektive Studien in Englisch oder Deutsch, die die CT Koronarangiografie mit der konventionellen Koronarangiografie (beide Verfahren bei allen Patienten durchgeführt) verglichen und adäquate Daten für eine Analyse auf Patientenebene lieferten. Ergebnisse: 120 Studien (mit 10.287 Patienten) wurden als relevant identifiziert. Die Studien unterschieden sich stark in ihrer Herangehensweise, mit nichtbeurteilbaren Ergebnissen zu verfahren. Wir fanden 26 Studien (mit 2298 Patienten), für die sowohl eine Vier- als auch eine zusätzliche Sechsfeldertafel berechnet werden konnte. Unter Benutzung eines bivariaten Random Effects Modells verglichen wir die Ergebnisse der Vierfeldertafel mit denen der Sechsfeldertafel und identifizierten statistisch signifikante Unterschiede für die gepoolte Sensitivität ((98,2 (95% Konfidenzintervall (KI) 96,7 bis 99,1) gegenüber 92,7 (88,5 bis 95,3)), die Fläche unter der Kurve (area under the curve) (0,99 (0,98 bis 1,00) gegenüber 0,93 (0,91 bis 0,95)), die positive Likelihood Ratio (9,1 (6,2 bis 13,3) gegenüber 4,4 (3,3 bis 6,0)) und die Negative Likelihood Ratio (0,02 (0,01 bis 0,04) gegenüber 0,09 (0,06 bis 0,15); (p<0,05)). Schlussfolgerung: Die Parameter der diagnostischen Wertigkeit nehmen signifikant ab, wenn nichtbeurteilbare Ergebnisse aus der Sechsfeldertafel mit in die Analyse einfließen (Intention-to-Diagnose Herangehensweise). Diese Herangehensweise liefert ein realistischeres Abbild der klinischen Wertigkeit diagnostischer Tests.Objective: To determine whether a 3×2 table, using an intention to diagnose approach, is better than the “classic” 2×2 table at handling transparent reporting and non-evaluable results, when assessing the accuracy of a diagnostic test. Design: Based on a systematic search for diagnostic accuracy studies of coronary computed tomography (CT) angiography, full texts of relevant studies were evaluated to determine whether they could calculate an alternative 3×2 table. To quantify an overall effect, we pooled diagnostic accuracy values according to a meta-analytical approach. Data sources: Medline (via PubMed), Embase (via Ovid), and ISI Web of Science electronic databases. Eligibility criteria: Prospective English or German language studies comparing coronary CT with conventional coronary angiography in all patients and providing sufficient data for a patient level analysis. Results: 120 studies (10,287 patients) were eligible. Studies varied greatly in their approaches to handling non-evaluable findings. We found 26 studies (including 2298 patients) that allowed us to calculate both 2×2 tables and 3×2 tables. Using a bivariate random effects model, we compared the 2×2 table with the 3×2 table, and found significant differences for pooled sensitivity (98.2 (95% confidence interval 96.7 to 99.1) v 92.7 (88.5 to 95.3)), area under the curve (0.99 (0.98 to 1.00) v 0.93 (0.91 to 0.95)), positive likelihood ratio (9.1 (6.2 to 13.3) v 4.4 (3.3 to 6.0)), and negative likelihood ratio (0.02 (0.01 to 0.04) v 0.09 (0.06 to 0.15); (P<0.05)). Conclusion: Parameters for diagnostic performance significantly decrease if non-evaluable results are included by a 3×2 table for analysis (intention to diagnose approach). This approach provides a more realistic picture of the clinical potential of diagnostic tests

Role of aggressivity on reactivity and craving before and after cue exposure in recently detoxified alcoholics: Results from an experimental study

The role of aggressivity and cue exposure in induction of craving were investigated in a clinical setting. Thirty abstinent alcoholic patients were divided into a low and a high aggressive group based on scores on the physical aggression subscale of the Buss-Durkee Hostility Inventory and exposed to alcohol cues. Craving was measured by means of the Alcohol Craving Questionnaire (ACQ) and Visual Analogue Scales (VAS). Important findings are: (1) main effects of aggressivity on `emotionality', `purposefulness' and `expectancy' of ACQ were very significant; (2) on `drinking intention' and `craving for alcohol' of VAS, aggressivity and cue exposure showed a significant interaction; (3) the main effect of cue exposure on heart rate also reached a significance level of 0.007. The results were discussed in the context of the Classical, Operant Conditioning Theory, the Cognitive Craving Theory of Tiffany, Gilbert's STAR Model, and the Self-Medication Hypothesis Copyright (C) 2001 S. Karger AG, Basel

Effects of irritability on craving before and after cue exposure in abstinent alcoholic inpatients: Experimental data on subjective response and heart rate

Objective: Irritability is often linked with problem drinking. The aim of this study is to examine the possible influence of irritability on craving induced by a cue-exposure paradigm. Methods: 30 male abstinent alcoholic inpatients of the Psychiatric Hospital of Munich University, Germany gave answers to a series of personality questionnaires. Results of this study concerning the impact of aggressivity on craving for alcohol has recently been published. In this study, the subjects were subdivided into a low- and a high-irritable group based on their scores on the irritability subscale of the Buss-Durkee Hostility Inventory and were exposed to alcohol cues. Craving was measured by means of the Alcohol Craving Questionnaire (ACQ) and Visual Analogue Scales (VAS). The heart rate was also assessed throughout the whole process. ANCOVA for repeated measurement was employed to evaluate the data - irritability disposition as the between-subject factor and the experimental manipulation (absence vs. presence of alcohol cues) as the within-subject factor. Results: Major findings are: (1) main effects of irritability on `emotionality', `purposefulness', and `expectancy' of the ACQ as well as on `craving for alcohol' of the VAS were significant; (2) cue exposure also exerted a significant main effect on I craving for alcohol' of the VAS and on the heart rate after the presentation of alcohol cues; (3) on `compulsivity' of the ACQ and `intention to alcohol intake' of the VAS; there was a significant interaction between irritability and cue exposure. The high-irritable alcoholics, compared with their statements in the baseline, tended to report a higher control over alcohol intake and a lower intention to alcohol use after cue exposure. However, after confrontation with alcohol stimuli, their low-irritable counterparts reported a much lower control and a slightly higher intention than they did in the baseline. Conclusions: The results of this study indicate that induced craving in hospitalized alcohol addicts probably varies with the magnitude of their irritability; it might make patients more aware of their vulnerability to alcohol, help them develop more differential coping strategies and improve medical therapy against alcohol craving. Copyright (C) 2002 S. Karger AG, Basel

Novel quorum sensing inhibitors targeting PqsR

Pseudomonas aeruginosa, a ubiquitous Gram-negative bacterium is counted among the most clinical relevant pathogens according to the World Health Organization (WHO). The opportunistic pathogen is able to form biofilms and persister cells, which makes it hard to eradicate and renders common antibiotic treatment ineffective. These factors are under control of the Pseudomonas Quinolone Signal (PQS) quorum sensing (QS) system, which thereofre displays an attractive drug target. This work describes the design and synthesis of novel PqsR inverse agonists based on a previously described hit compound. A highly divergent synthetic route was established and enabled the synthesis of various classes of 2-(trifluoromethyl)pyridin-4-amines providing deep structure-activity relationship (SAR) insights. Lead generation resulted in a highly active optimized compound possessing also good drug metabolism and pharmacokinetic (DMPK) properties. Furthermore, this compound showed in vivo target engagement in a murine lung infection model. Based on this lead compound, a medicinal chemistry-driven multiparameter optimization allowed for deeper insights into structure-activity relationship (SAR) and structure-property relationship (SPR). In the course of this thesis, several compounds were synthesized with improved activity, solubility and metabolic stability. In addition, compounds based on computer-aided drug design were synthesized.Pseudomonas aeruginosa, ein omnipräsentes Gramnegatives Bakterium zählt laut Weltgesundheitsorganisation (WHO) zu den klinisch relevantesten Pathogenen. Dieses opportunistische Pathogen ist in der Lage, Biofilme und Persisterzellen zu bilden, die eine Eradizierung erschweren und übliche Antibiotika Therapien ineffektiv werden lassen. Diese Faktoren unterliegen dem Pseudomonas Quinolone Signal (PQS) Quorum Sensing (QS) System, welches dabei ein attraktives Target zur Behandlung dar. Diese Arbeit beschreibt das Design und die Synthese von neuen PqsR inversen Agonisten, basierend auf einer bereits beschriebenen hit Verbindung. Eine hoch divergente Syntheseroute wurde dabei etabliert und ermöglichte die Synthese verschiedener Klassen von 2-(Trifluoromethyl)pyridin-4-aminen und lieferte tiefe Einblicke in die Struktur-Aktivitätsbeziehungen. Die Leitstruktur Generierung brachte eine hoch-aktive Verbindung hervor, die ebenso über gute pharmakokinetische Eigenschaften verfügte und in einem murinen Lungeninfektionsmodel in vivo target engagement zeigte. Ausgehend von dieser Leitstruktur lieferte eine medizinal-chemische Multiparameter-Optimierung tiefe Einblicke in Struktur-Aktivitätsbeziehungen und Struktur-Eigenschaftsbeziehungen. Im Zuge dieser Leitstruktur Optimierung wurden verschiedene Verbindungen mit verbesserter Aktivität, Löslichkeit und Stabilität synthetisiert. Darüber hinaus wurden Verbindungen basierend auf Computer-gestütztem Wirkstoffdesign synthetisiert.DZIF, HV