Funktionelle Charakterisierung der Gene NPC2 und ADRBK1 im Pankreaskarzinom

Das duktale Adenokarzinom des Pankreas stellt heute immer noch eines der aggressivsten und am schlechtesten behandelbaren Malignome dar. Deshalb ist es umso wichtiger, Genetik und Pathomechanismen dieser Erkrankung im Detail aufzuklären und Marker zur Früh- bzw. Differenzialdiagnose sowie potentielle Ziele für targeted therapies zu finden. Mit Hilfe von enzymatisch generierten, Transkriptom-basierten shRNA-Bibliotheken konnten in Hochdurchsatzanalysen potentielle Onko- bzw. Tumorsuppressorgene in PDAC identifiziert werden. NPC2 wurde als eines dieser potentiellen Onkogene für weitere funktionelle Assays im Bezug auf Zellvitalität, -proliferation und -motilität ausgewählt. Expressionsanalysen auf mRNA-Ebene zeigten eine deutliche Überexpression des Gens sowohl in pankreatischem Tumorgewebe als auch in verschiedenen Pankreaskarzinomzelllinien. Von vier in dieser Arbeit untersuchten Zelllinien zeigte sich nach siRNA-vermitteltem Knockdown jedoch nur in BxPC3-Zellen ein akuter funktioneller Effekt im Sinne einer hoch signifikanten Abnahme der Zellvitalität. Dieser wachstumsfördernde Effekt von NPC2 lässt sich nicht auf das Pankreaskarzinom im Allgemeinen übertragen. Die Zellproliferation und –motilität waren nach transienter Repression von NPC2 im Kurzzeit-Versuch nicht beeinträchtigt. Auch das Gen ADRBK1 wurde in einem Hochdurchsatz-Screening, welches auf Pankreaskarzinom-spezifischen cDNA-Microarrays und dem Prinzip der reversen Transfektion basierte, identifiziert. ADRBK1 zeigte eine zeitabhängige Translokation in den Nukleus und es stellte sich die Frage, ob die Kinase eine direkte Rolle bei der Genregulation spielt. Vorversuche der Arbeitsgruppe zeigten zudem, dass ADRBK1 im humanen PDAC nicht nur signifikant überexprimiert wird, sondern auch pro- proliferative Effekte vermittelt. Rekombinante Überexpression der Kinase beschleunigte das Zellwachstum, während ihr Knockdown dieses signifikant inhibierte. Als Ursache für die beobachtete Wachstumshemmung kamen potentiell die Induktion eines Zellzyklusarrests oder Apoptose in Frage. Die daraufhin von mir im Western-Blot untersuchten Proteinlevel von CDKN1A/p21, CDKN1B/p27, Cyclin A, Cyclin D1, pRB und c-myc waren nach ADRBK1-Repression jedoch unverändert. Eine Spaltung der Apoptosemarker PARP und Caspase 3 war nur in der Zelllinie PaTu-8988t nachweisbar. Somit kann weder die Induktion von Apoptose noch ein Zellzyklusarrest als maßgebliche Ursache der Wachstumsabnahme nach ADRBK-Knockdown gelten. Um die Bedeutung des Gens für PDAC weiter aufzuklären, sollte die nukleäre Translokation und deren potentielle Auswirkungen auf die Genfunktion weiter untersucht werden. Zudem sollten zelluläre Zielproteine von ADRBK1, die Relevanz der Kinase beim Zusammenspiel von Krebszellen mit infiltrierenden Immunzellen und ihre Rolle bezüglich der Zellmotilität identifiziert werden

On the Constructive Content of Proofs

This thesis aims at exploring the scopes and limits of techniques for extracting programs from proofs. We focus on constructive theories of inductive definitions and classical systems allowing choice principles. Special emphasis is put on optimizations that allow for the extraction of realistic programs. Our main field of application is infinitary combinatorics. Higman's Lemma, having an elegant non-constructive proof due to Nash-Williams, constitutes an interesting case for the problem of discovering the constructive content behind a classical proof. We give two distinct solutions to this problem. First, we present a proof of Higman's Lemma for an arbitrary alphabet in a theory of inductive definitions. This proof may be considered as a constructive counterpart to Nash-Williams' minimal-bad-sequence proof. Secondly, using a refined $A$-translation method, we directly transform the classical proof into a constructive one and extract a program. The crucial point in the latter is that we do not need to avoid the axiom of classical dependent choice but directly assign a realizer to its translation. A generalization of Higman's Lemma is Kruskal's Theorem. We present a constructive proof of Kruskal's Theorem that is completely formalized in a theory of inductive definitions. As a practical part, we show that these methods can be carried out in an interactive theorem prover. Both approaches to Higman's Lemma have been implemented in Minlog.Ziel der vorliegenden Arbeit ist es, die Reichweiten und Grenzen von Techniken zur Extraktion von Programmen aus Beweisen zu erforschen. Wir konzentrieren uns dabei auf konstruktive Theorien Induktiver Definitionen und klassische Systeme mit Auswahlprinzipien. Besonderes Gewicht liegt auf Optimierungen, die zur Extraktion von realisischen Programmen f"uhren. Unser Hauptanwendungsgebiet ist die unendliche Kombinatorik. Higmans Lemma, ein Satz mit einem eleganten klassischen, auf Nash-Williams zur"uckgehenden Beweis, ist ein interessantes Fallbeispiel f"ur die Suche nach dem konstruktiven Gehalt in einem klassischen Beweis. Wir zeigen zwei unterschiedliche L"osungen zu dieser Problemstellung auf. Zun"achst pr"asentieren wir einen induktiven Beweis von Higmans Lemma f"ur ein beliebiges Alphabet, der als konstruktives Pendant zum klassischen Beweis angesehen werden kann. Als zweiten Ansatz verwandeln wir mit Hilfe der verfeinerten $A$-"Ubersetzungs-methode den klassischen Beweis in einen konstruktiven und extrahieren ein Programm. Der entscheidende Punkt ist hierbei, dass wir einen direkten Realisierer f"ur das "ubersetzte Auswahlaxiom verwenden. Die Verallgemeinerung von Higmans Lemma f"uhrt zu Kruskals Satz. Wir geben einen konstruktiven Beweis von Kruskals Theorem, der vollst"andig auf den Induktiven Definitionen basiert. Der praktische Teil der Arbeit befasst sich mit der Ausf"uhrbarkeit dieser Methoden und Beweise in dem Beweissystem Minlog

The Chemistry Mechanism in the Community Earth System Model Version 2 (CESM2)

The Community Earth System Model version 2 (CESM2) includes a detailed representation of chemistry throughout the atmosphere in the Community Atmosphere Model with chemistry and Whole Atmosphere Community Climate Model configurations. These model configurations use the Model for Ozone and Related chemical Tracers (MOZART) family of chemical mechanisms, covering the troposphere, stratosphere, mesosphere, and lower thermosphere. The new MOZART tropospheric chemistry scheme (T1) has a number of updates over the previous version (MOZART‐4) in CESM, including improvements to the oxidation of isoprene and terpenes, organic nitrate speciation, and aromatic speciation and oxidation and thus improved representation of ozone and secondary organic aerosol precursors. An evaluation of the present‐day simulations of CESM2 being provided for Climate Model Intercomparison Project round 6 (CMIP6) is presented. These simulations, using the anthropogenic and biomass burning emissions from the inventories specified for CMIP6, as well as online calculation of emissions of biogenic compounds, lightning NO, dust, and sea salt, indicate an underestimate of anthropogenic emissions of a variety of compounds, including carbon monoxide and hydrocarbons. The simulation of surface ozone in the southeast United States is improved over previous model versions, largely due to the improved representation of reactive nitrogen and organic nitrate compounds resulting in a lower ozone production rate than in CESM1 but still overestimates observations in summer. The simulation of tropospheric ozone agrees well with ozonesonde observations in many parts of the globe. The comparison of NOx and PAN to aircraft observations indicates the model simulates the nitrogen budget well

Trends in postoperative radiotherapy delay and the effect on survival in breast cancer patients treated with conservation surgery

The adequate timing of adjuvant radiotherapy (RT) in breast cancer has become a subject of increasing interest in recent years. A population-based study was undertaken to determine the influence of demographic and clinical factors on the postoperative RT delay in patients treated with breast-conserving surgery (BCS) and to assess the impact of delay on survival. In total, 7800 breast cancer patients treated with BCS and adjuvant RT between 1986 and 1998 in Yorkshire were included in the study. The median interval between surgery and the start of RT (S-RT interval) was 8 weeks (7 weeks for chemotherapy negative and 11 for chemotherapy positive patients). This interval increased substantially over time from 5 weeks during 1986-1988, irrespective of patients' chemotherapy status, to 10 and 17 weeks among chemotherapy negative and chemotherapy positive patients, respectively, in 1997-1998. The S-RT interval was also significantly influenced by travel time to RT centre, year and at which RT centre patient had the treatment (

Toxic effects of Pb2+ on the growth and mineral nutrition of signal grass (Brachiaria decumbens) and Rhodes grass (Chloris gayana)

Although grasses are commonly used to revegetate sites contaminated with lead (Pb), little is known regarding the Pb-tolerance of many of these species. Using dilute solution culture to mimic the soil solution, the growth of signal grass (Brachiaria decumbens Stapf cv. Basilisk) and Rhodes grass (Chloris gayana Kunth cv. Pioneer) was related to the mean activity of Pb2+ {Pb2+} in solution. There was a 50% reduction in fresh mass of signal grass shoots at 5 mu M {Pb2+} and at 3 mu M {Pb2+} for the roots. Rhodes grass was considerably more sensitive to Pb in solution, with shoot and root fresh mass being reduced by 50% at 0.5 mu M {Pb2+}. The higher tolerance of signal grass to Pb appeared to result from the internal detoxification of Pb, rather than from the exclusion of Pb from the root. At toxic {Pb2+}, an interveinal chlorosis developed in the shoots of signal grass (possibly a Pb-induced Mn deficiency), whilst in Rhodes grass, Pb2+ caused a bending of the root tips and the formation of a swelling immediately behind some of the root apices. Root hair growth did not appear to be reduced by Pb2+ in solution, being prolific at all {Pb2+} in both species

Characterization of Xenopus Tissue Inhibitor of Metalloproteinases-2: A Role in Regulating Matrix Metalloproteinase Activity during Development

Frog metamorphosis is totally dependent on thyroid hormone (T3) and mimics the postembryonic period around birth in mammals. It is an excellent model to study the molecular basis of postembryonic development in vertebrate. We and others have shown that many, if not all, matrix metalloproteinases (MMPs), which cleave proteins of the extracellular matrix as well as other substrates, are induced by T3 and important for metamorphosis. MMP activity can be inhibited by tissue inhibitors of metalloproteinase (TIMPs). There are 4 TIMPs in vertebrates and their roles in postembryonic development are poorly studied.We analyzed the TIMP2 genes in Xenopus laevis and the highly related species Xenopus tropicalis and discovered that TIMP2 is a single copy gene in Xenopus tropicalis as in mammals but is duplicated in Xenopus laevis. Furthermore, the TIMP2 locus in Xenopus tropicalis genome is different from that in human, suggesting an evolutionary reorganization of the locus. More importantly, we found that the duplicated TIMP2 genes were similarly regulated in the developing limb, remodeling intestine, resorbing tail during metamorphosis. Unexpectedly, like its MMP target genes, the TIMP2 genes were upregulated by T3 during both natural and T3-induced metamorphosis.Our results indicate that TIMP2 is highly conserved among vertebrates and that the TIMP2 locus underwent a chromosomal reorganization during evolution. Furthermore, the unexpected upregulation of TIMP2 genes during metamorphosis suggests that proper balance of MMP activity is important for metamorphosis

Limits on WWZ and WW\gamma couplings from p\bar{p}\to e\nu jj X events at \sqrt{s} = 1.8 TeV

We present limits on anomalous WWZ and WW-gamma couplings from a search for WW and WZ production in p-bar p collisions at sqrt(s)=1.8 TeV. We use p-bar p -> e-nu jjX events recorded with the D0 detector at the Fermilab Tevatron Collider during the 1992-1995 run. The data sample corresponds to an integrated luminosity of 96.0+-5.1 pb^(-1). Assuming identical WWZ and WW-gamma coupling parameters, the 95% CL limits on the CP-conserving couplings are -0.33<lambda<0.36 (Delta-kappa=0) and -0.43<Delta-kappa<0.59 (lambda=0), for a form factor scale Lambda = 2.0 TeV. Limits based on other assumptions are also presented.Comment: 11 pages, 2 figures, 2 table

Improving the Prognostic Ability through Better Use of Standard Clinical Data - The Nottingham Prognostic Index as an Example

Background Prognostic factors and prognostic models play a key role in medical research and patient management. The Nottingham Prognostic Index (NPI) is a well-established prognostic classification scheme for patients with breast cancer. In a very simple way, it combines the information from tumor size, lymph node stage and tumor grade. For the resulting index cutpoints are proposed to classify it into three to six groups with different prognosis. As not all prognostic information from the three and other standard factors is used, we will consider improvement of the prognostic ability using suitable analysis approaches. Methods and Findings Reanalyzing overall survival data of 1560 patients from a clinical database by using multivariable fractional polynomials and further modern statistical methods we illustrate suitable multivariable modelling and methods to derive and assess the prognostic ability of an index. Using a REMARK type profile we summarize relevant steps of the analysis. Adding the information from hormonal receptor status and using the full information from the three NPI components, specifically concerning the number of positive lymph nodes, an extended NPI with improved prognostic ability is derived. Conclusions The prognostic ability of even one of the best established prognostic index in medicine can be improved by using suitable statistical methodology to extract the full information from standard clinical data. This extended version of the NPI can serve as a benchmark to assess the added value of new information, ranging from a new single clinical marker to a derived index from omics data. An established benchmark would also help to harmonize the statistical analyses of such studies and protect against the propagation of many false promises concerning the prognostic value of new measurements. Statistical methods used are generally available and can be used for similar analyses in other diseases

Search For Heavy Pointlike Dirac Monopoles

We have searched for central production of a pair of photons with high transverse energies in $p\bar p$ collisions at $\sqrt{s} = 1.8$ TeV using $70 pb^{-1}$ of data collected with the D\O detector at the Fermilab Tevatron in 1994--1996. If they exist, virtual heavy pointlike Dirac monopoles could rescatter pairs of nearly real photons into this final state via a box diagram. We observe no excess of events above background, and set lower 95% C.L. limits of $610, 870, or 1580 GeV/c^2$ on the mass of a spin 0, 1/2, or 1 Dirac monopole.Comment: 12 pages, 4 figure