Analyse der Redundanz von SHOX und SHOX2 und Identifizierung von Shox2-Zielgenen

Die paralogen Gene SHOX und SHOX2 kodieren für Homöodomänen-Transkriptionsfaktoren mit wichtigen Funktionen innerhalb der Embryonalentwicklung. SHOX wurde als menschliches Wachstumskontrollgen identifiziert. Mutationen dieses Gens führen zu den skelettalen Fehlbildungen, die bei Léri-Weill-, Langer-, und Turner-Syndrom beobachtet werden, und sind zudem mit isoliertem Kleinwuchs assoziiert. Analysen mit Hilfe des Mausmodells zeigten, dass Shox2 ebenfalls eine entscheidende Rolle in der Entwicklung der Extremitäten spielt und darüber hinaus für weitere Prozesse, wie z.B. die Herzentwicklung erforderlich ist. Funktionelle Analysen beider Gene ergaben weitgehende Übereinstimmungen hinsichtlich ihrer Expression, subzellulären Lokalisierung, DNA-Bindung sowie ihrer transaktivierenden Eigenschaften. Diese Befunde sprechen für überlappende Funktionen beider Gene. Ein Ziel dieser Arbeit war es, die Redundanz beider Gene mit Hilfe des Maus- und des Hühnermodells genauer zu untersuchen. Da die Maus kein SHOX-Ortholog besitzt, wurde eine targeting-Strategie entwickelt, um menschliches SHOX unter dem endogenen Shox2-Promotor in die Maus einzubringen und gleichzeitig Shox2 auszuschalten. Dieses Shox2-knockout / SHOX-knockin-Mausmodell würde ein geeignetes Modell darstellen, um die gemeinsamen Funktionen beider Gene zu untersuchen, allerdings war dessen Generierung im Rahmen dieser Arbeit nicht möglich. Daher wurde die Redundanz von Shox und Shox2 mit Hilfe vergleichender Expressionsanalysen in Maus und Huhn untersucht. Es zeigte sich dabei, dass Shox und Shox2 in unterschiedlichen Bereichen des Kiefers und der Extremitäten exprimiert sind. Im Nervensystem stimmen die Expressionsmuster beider Gene weitgehend überein, wobei Shox2 etwas breiter exprimiert ist. Dies spricht für gemeinsame Funktionen beider Gene und könnte erklären, warum der Verlust von SHOX bisher mit keinem menschlichen neuronalen Phänotyp in Verbindung gebracht werden konnte. Möglicherweise kann SHOX2 bei einem Verlust des SHOX-Gens die Funktion in der neuronalen Entwicklung vollständig übernehmen. Im zweiten Teil dieser Arbeit wurde die Rolle von Shox2 in der murinen Gliedmaßenentwicklung näher untersucht. Um Einblicke in Shox2-abhängige Signalwege zu erlangen sollten Zielgene von Shox2 isoliert werden. Mit Hilfe von microarray-Analysen und nachfolgenden qRT-PCR-Analysen wurde Tbx4 als interessantestes Kandidatengen identifiziert. Unter Verwendung des Shox2-knockout-Mausmodells konnte durch whole mount in situ Hybridisierungen und quantitative Analysen gezeigt werden, dass Shox2 die Expression von Tbx4 spezifisch in den Vordergliedmaßen beeinflusst. Nachfolgende Experimente mit Hilfe eines Tbx4-defizienten Mausmodells ergaben wiederum, dass Tbx4 Shox2 in Vorder- und Hinterbeinen positiv reguliert und es sich bei dem zuvor beobachteten Effekt von Shox2 auf Tbx4 wahrscheinlich um einen feedback-Mechanismus handelt. Luziferase-Assays und EMSA-Studien zeigten außerdem, dass es drei TBX4-Bindestellen im genomischen Promotorbereich von SHOX2 gibt und es sich bei dem Einfluss von Tbx4 auf Shox2 um eine direkte Regulation handelt. Damit konnte Tbx4 im Rahmen dieser Arbeit als erster direkter Regulator von Shox2 in der Gliedmaßenentwickung charakterisiert werden, was auf eine mögliche Rolle von Shox2 bei der Initiation des Auswachsens und der Identitätsgebung von Extremitätenknospen hinweist. Weiterhin trägt die Regulation von Shox2 durch Tbx4 zur Aufklärung der Tbx4-involvierten Signalwege in Prozessen wie Muskel- und Skelettentwicklung bei. Erstmalig konnte damit die Beteiligung von Shox2 an einem transkriptionellen feedback-Mechanismus demonstriert werden

Playing Three-Level Games in the Global Economy. Case Studies from the EU. College of Europe EU Diplomacy Paper 4/2008, May 2008

The case studies in this paper are a selection of essays that have been written in the framework of the compulsory first-semester course The EU in a Global Political Economy Context, taught by Professor Sieglinde Gstöhl, in the academic year 2007-2008 in the EU International Relations and Diplomacy Studies programme at the College of Europe. They all address recent cases of two- or three-level games played by the European Union in different policy fields of the global economy (reflecting the state of affairs at the end of 2007)

AN EXPLORATION OF ENROLLMENT AND RETENTION TRENDS OF BEGINNING BAND AND ORCHESTRA STUDENTS IN THE FIRST YEAR OF INSTRUCTION

The purpose of this study was to explore student attitudes towards enrollment and retention in first-year beginning band and orchestra classrooms. A secondary purpose of this study was to investigate if different instrumental ensembles or various school settings demonstrated unique student attitudes regarding enrollment and retention rates in beginning band or orchestra classes. Enrollment and retention rates of participating ensembles were reported to supplement qualitative results. Seven categories of themes influencing enrollment and retention in beginning band and orchestra classes emerged through a constant comparative, grounded theory approach of analysis: (a) family, (b) fun, (c) music, (d) musical history, (e) opportunities, (f) social, and (g) teacher. Results indicated that students enrolled in their first year of beginning band or orchestra because of the encouragement or influence of a parent or trusted adult. All students that elected to continue their enrollment in band or orchestra after the first year of instruction did under perceived support from their parents and/or instrumental music teacher. Ensemble- and location-specific results were found, but were interpreted as circumstantial. Further research is necessary to explore the unique enrollment trends of these groupings. Results were discussed in terms of their value to band and orchestra teachers, their relationship to existing literature, limitations, and suggestions for further research

Association between clinic-level quality of care and patient-level outcomes in multiple sclerosis

Background: Multiple sclerosis (MS) quality of care guidelines are consensus-based. The effectiveness of the recommendations is unknown.// Objective: To determine whether clinic-level quality of care affects clinical and patient-reported outcomes.// Methods: This nationwide observational cohort study included patients with adult-onset MS in the Swedish MS registry with disease onset 2005–2015. Clinic-level quality of care was measured by four indicators: visit density, magnetic resonance imaging (MRI) density, mean time to commencement of disease-modifying therapy, and data completeness. Outcomes were Expanded Disability Status Scale (EDSS) and patient-reported symptoms measured by the Multiple Sclerosis Impact Scale (MSIS-29). Analyses were adjusted for individual patient characteristics and disease-modifying therapy exposure.// Results: In relapsing MS, all quality indicators benefitted EDSS and physical symptoms. Faster treatment, frequent visits, and higher data completeness benefitted psychological symptoms. After controlling for all indicators and individual treatment exposures, faster treatment remained independently associated with lower EDSS (−0.06, 95% confidence interval (CI): −0.01, −0.10) and more frequent visits were associated with milder physical symptoms (MSIS-29 physical score: −16.2%, 95% CI: −1.8%, −29.5%). Clinic-level quality of care did not affect any outcomes in progressive-onset disease.// Conclusion: Certain quality of care indicators correlated to disability and patient-reported outcomes in relapse-onset but not progressive-onset disease. Future guidelines should consider recommendations specific to disease course..

Microtubule depolymerization by the kinesin-8 motor Kip3p: a mathematical model

Proteins from the kinesin-8 family promote microtubule (MT) depolymerization, a process thought to be important for the control of microtubule length in living cells. In addition to this MT shortening activity, kinesin 8s are motors that show plus-end directed motility on MTs. Here we describe a simple model that incorporates directional motion and destabilization of the MT plus end by kinesin 8. Our model quantitatively reproduces the key features of length-vs-time traces for stabilized MTs in the presence of purified kinesin 8, including length-dependent depolymerization. Comparison of model predictions with experiments suggests that kinesin 8 depolymerizes processively, i.e., one motor can remove multiple tubulin dimers from a stabilized MT. Fluctuations in MT length as a function of time are related to depolymerization processivity. We have also determined the parameter regime in which the rate of MT depolymerization is length dependent: length-dependent depolymerization occurs only when MTs are sufficiently short; this crossover is sensitive to the bulk motor concentration.Comment: 34 pages, 11 figure

Identification of novel SHOX target genes in the developing limb using a transgenic mouse model

Deficiency of the human short stature homeobox-containing gene (SHOX) has been identified in several disorders characterized by reduced height and skeletal anomalies such as Turner syndrome, Léri-Weill dyschondrosteosis and Langer mesomelic dysplasia as well as isolated short stature. SHOX acts as a transcription factor during limb development and is expressed in chondrocytes of the growth plates. Although highly conserved in vertebrates, rodents lack a SHOX orthologue. This offers the unique opportunity to analyze the effects of human SHOX expression in transgenic mice. We have generated a mouse expressing the human SHOXa cDNA under the control of a murine Col2a1 promoter and enhancer (Tg(Col2a1-SHOX)). SHOX and marker gene expression as well as skeletal phenotypes were characterized in two transgenic lines. No significant skeletal anomalies were found in transgenic compared to wildtype mice. Quantitative and in situ hybridization analyses revealed that Tg(Col2a1-SHOX), however, affected extracellular matrix gene expression during early limb development, suggesting a role for SHOX in growth plate assembly and extracellular matrix composition during long bone development. For instance, we could show that the connective tissue growth factor gene Ctgf, a gene involved in chondrogenic and angiogenic differentiation, is transcriptionally regulated by SHOX in transgenic mice. This finding was confirmed in human NHDF and U2OS cells and chicken micromass culture, demonstrating the value of the SHOX-transgenic mouse for the characterization of SHOX-dependent genes and pathways in early limb development

PENGGUNAAN MODEL PEMBELAJARAN AKTIF TEAM QUIZ DALAM MENINGKATKAN KEMAMPUAN MEMBACA PEMAHAMAN BAHASA JEPANG

Penelitian ini mengambil judul “penggunaan model pembelajaran aktif team quiz dalam meningkatkan kemampuan membaca pemahaman bahasa Jepang” . membaca pemahaman adalah memahami suatu informasi dari sebuah tulisan. membaca pemahaman merupakan salah satu pembelajaran yang sulit. Disebabkan karena metode yang digunakan oleh pengajar monoton dan kurang memotivasi siswa dalam pembelajaran membaca pemahaman. Penelitian ini bertujuan untuk mengetahui bagaimana penggunaan model pembelajaran aktif team quiz dalam meningkatkan kemampuan membaca pemahaman bahasa Jepang. Penelitian ini menggunakan metode quasi eksperiment dengan one group before-after (pretest dan posttest) design. Sampel dalam penelitian ini adalah siswa kelas XI Bahasa SMA Negeri 1 Parongpong tahun ajaran 2015/2016. Instrument penelitiannya adalah tes dan angket. Berdasarkan analisis data dengan perhitungan statistic yang menggunakan t hitung diketahui bahwa t hitung > t tabel maka Ho ditolak dan Hk diterima yang dapat disimpulkan bahwa penggunaan model pembelajaran aktif team quiz dalam meningkatkan kemampuan membaca pemahaman bahasa jepang efektif. Sedangkan hasil analisis data angket diketahui bahwa penggunaan model pembelajaran aktif team quiz dalam meningkatkan kemampuan membaca pemahaman bahasa Jepang mendapat respon positif dari siswa. ; Reading comprehension is the act of understanding information from a presented text. It is one of difficulties found in learning. It caused by monotonous method conducted by teacher and in turn, students have less motivation on learning reading comprehension. This research aims for how the Team Quiz active learning method can improve reading comprehension in Japanese. The quasi-experiment method with “one group before-after (pretest and posttest)” design is conducted in this research, by taking XI language students from 2015/2016 academic year of SMA Negeri 1 Parongpong as sample. Test and questionnaire is instrument used in this research. According to data analysis with statistical calculation using t count, noting that t count > t table, therefore Ho is rejected and Hk is accepted. To conclude, Team Quiz active learning method is effective in improving reading comprehension in Japanese. Moreover, according to data analysis conducted in questionnaire, the Team Quiz active learning method gains positive responses from students

Symptomatic cerebral oedema during treatment of diabetic ketoacidosis: effect of adjuvant octreotide infusion

<p>Abstract</p> <p>Introduction</p> <p>A potentially lethal complication of diabetic ketoacidosis (DKA) in children is brain oedema, whether caused by DKA itself or by the therapeutic infusion of insulin and fluids.</p> <p>Case presentation</p> <p>A 10-year old previously healthy boy with DKA became unconscious and apnoeic due to cerebral oedema (confirmed by abnormal EEG and CT-scan) during treatment with intravenous fluids (36 ml/h) and insulin (0.1 units/kg/h). He was intubated and artificially ventilated, without impact on EEG and CT-scan. Subsequently, adjuvant infusion of octreotide was applied (3.5 μg/kg/h), suppressing growth hormone (GH) and IGF-1 production and necessitating the insulin dose to be reduced to 0.05 - 0.025 units/kg/h. The brain oedema improved and the boy made a full recovery.</p> <p>Conclusion</p> <p>Co-therapy with octreotide was associated with a favourable outcome in the present patient with DKA and cerebral oedema. Whether this could be ascribed to the effects of octreotide on the insulin requirement or on the GH/IGF-axis remains to be elucidated.</p

Hyperglycaemia induces metabolic dysfunction and glycogen accumulation in pancreatic β-cells

Insulin secretion from pancreatic β-cells is impaired in all forms of diabetes. The resultant hyperglycaemia has deleterious effects on many tissues, including β-cells. Here we use a mouse model of human neonatal diabetes to show that chronic hyperglycemia impairs glucose metabolism and alters expression of metabolic genes in pancreatic islets. This results in marked glycogen accumulation, and increased apoptosis in β-cells. Sulphonylurea therapy rapidly normalizes blood glucose levels, dissipates glycogen stores, increases autophagy, and restores β-cell metabolism. Insulin therapy has the same effect but with slower kinetics. Similar changes are observed in mice expressing an activating glucokinase mutation, in in vitro models of hyperglycaemia, and in islets from type-2 diabetes patients. Altered β-cell metabolism may underlie both the progressive impairment of insulin secretion and reduced β-cell mass in diabetes