Impact of DICER1 and DROSHA on the Angiogenic Capacity of Human Endothelial Cells

RNAi-mediated knockdown of DICER1 and DROSHA, enzymes critically involved in miRNA biogenesis, has been postulated to affect the homeostasis and the angiogenic capacity of human endothelial cells. To re-evaluate this issue, we reduced the expression of DICER1 or DROSHA by RNAi-mediated knockdown and subsequently investigated the effect of these interventions on the angiogenic capacity of human umbilical vein endothelial cells (HUVEC) in vitro (proliferation, migration, tube formation, endothelial cell spheroid sprouting) and in a HUVEC xenograft assay in immune incompetent NSG^TM mice in vivo. In contrast to previous reports, neither knockdown of DICER1 nor knockdown of DROSHA profoundly affected migration or tube formation of HUVEC or the angiogenic capacity of HUVEC in vivo. Furthermore, knockdown of DICER1 and the combined knockdown of DICER1 and DROSHA tended to increase VEGF-induced BrdU incorporation and induced angiogenic sprouting from HUVEC spheroids. Consistent with these observations, global proteomic analyses showed that knockdown of DICER1 or DROSHA only moderately altered HUVEC protein expression profiles but additively reduced, for example, expression of the angiogenesis inhibitor thrombospondin-1. In conclusion, global reduction of miRNA biogenesis by knockdown of DICER1 or DROSHA does not inhibit the angiogenic capacity of HUVEC. Further studies are therefore needed to elucidate the influence of these enzymes in the context of human endothelial cell-related angiogenesis.     

Medical target prediction from genome sequence: combining different sequence analysis algorithms with expert knowledge and input from artificial intelligence approaches.

By exploiting the rapid increase in available sequence data, the definition of medically relevant protein targets has been improved by a combination of: (i) differential genome analysis (target list): and (ii) analysis of individual proteins (target analysis). Fast sequence comparisons, data mining, and genetic algorithms further promote these procedures. Mycobacterium tuberculosis proteins were chosen as applied examples.     

Lineare versus sprungfixe Erfolgsbeteiligung

Zusammenfassung Der vorliegende Beitrag diskutiert die Gestaltung optimaler Anreizsysteme für zwei Typen von Erfolgsbeteiligungen: die lineare und die sprungfixe Belohnungsfunktion. Im Gegensatz zu der üblichen Annahme der Principal-Agent-Theorie einer einmaligen Realisation eines Projekts betrachten wir die spezielle Situation, daß eine mehrmalige Wiederholung eines bestimmten Projekttyps in gleicher Form möglich ist. Es wird gezeigt, daß für den Fall der Risikoversion des Entscheidungsträgers und Risikoneutralität der Instanz die sprungfixe Belohnungsfunktion gegenüber der linearen Belohnungsfunktion von Vorteil sein kann.Wir danken einem anonymen Gutachter für wertvolle Verbesserungsvorschläge.     

Behavioral Subtyping Relations for Active Objects

In this paper we investigate the object-oriented notion of subtyping in the context of behavioral formalisms. Behavioral formalisms are used to describe the dynamic behavior of active objects, viz. the protocols of interaction that active objects have to obey.Subtyping in OO-formalisms is closely related to the concept of inheritance. The central issue in the choice of subtyping relations among classes is the principle of substitutability: an instance of the subtype should be usable wherever an instance of the supertype was expected. Depending on the interpretation of usable, we obtain a variety of subtyping relations: stronger relations, allowing sharing of subtype instances among different clients, and weaker relations, restricting the possibilities of interference of different clients on the subtype instance. The subtyping relations are taxonomically ordered in a hierarchy. Substitutability is formalized via testing scenarios, which provide alternative characterizations for the subtyping relations.     

Development of a method for the quantification of the molar gold concentration in tumour cells exposed to gold-containing drugs

The knowledge of the cellular molar concentration of a drug is an extremely important parameter for the discussion and interpretation of its efficacy and bioavailability. Concerning metal complexes, electrothermal atomic absorption spectroscopy (ETAAS) offers a valuable analytical tool. However, matrix effects often hamper proper quantification of the metal concentration in biological tissues. This paper describes the development of an ETAAS method for the quantification of the molar gold concentration in HT-29 colon carcinoma cells. ETAAS analytical conditions were optimised and a factor was developed which allows the calculation of the molar cellular gold concentration from the measured gold per cellular biomass value. The method was used to quantify the gold content in HT-29 cells after exposure to the gold drug auranofin. Results indicated a strong cellular uptake of auranofin (compared to other metal anticancer drugs), which significantly correlated with the antiproliferative effects triggered by this agent.     

Evaluation of bacterial RNase P RNA as a drug target

RNA has gained increasing importance as a therapeutic target. However, so far mRNAs rather than stable cellular RNAs have been considered in such studies. In bacteria, the tRNA-processing enzyme RNase P has a catalytic RNA subunit. Fundamental differences in structure and function between bacterial and eukaryotic RNase P, and its indispensability for cell viability make the bacterial enzyme an attractive drug target candidate. Herein we describe two approaches utilized to evaluate whether the catalytic RNA subunit of bacterial RNase P is amenable to inactivation by antisense-based strategies. In the first approach, we rationally designed RNA hairpin oligonucleotides targeted at the tRNA 3'-CCA binding site (P15 loop region) of bacterial RNase P RNA by attempting to include principles derived from the natural CopA-CopT antisense system. Substantial inactivation of RNase P RNA was observed for Type A RNase P RNA (such as that in Escherichia coli) but not for Type B (as in Mycoplasma hyopneumoniae). Moreover, only an RNA oligonucleotide (Eco 3') complementary to the CCA binding site and its 3' flanking sequences was shown to be an efficient inhibitor. Mutation of Eco 3' and analysis of other natural RNase P RNAs with sequence deviations in the P15 loop region showed that inhibition is due to interaction of Eco 3' with this region and occurs in a highly sequence-specific manner. A DNA version of Eco 3' was a less potent inhibitor. The potential of Eco 3' to form an initial kissing complex with the P15 loop did not prove advantageous. In a second approach, we tested a set of oligonucleotides against E. coli RNase P RNA which were designed by algorithms developed for the selection of suitable mRNA targets. This approach identified the P10/11-J11/12 region of bacterial RNase P RNA as another accessible region. In conclusion, both the P15 loop and P10/11-J11/12 regions of Type A RNase P RNAs seem to be promising antisense target sites since they are easily accessible and sufficiently interspersed with nonhelical sequence elements, and oligonucleotide binding directly interferes with substrate docking to these two regions.     

High-Temperature Electrical Properties in the ThO2-Rich Region of ThO2-RE2O3 Systems

In the present study the n -type electronic conduction in terms of the parameter p e, and the phase relations in several ThO₂-RE2O₃ systems were examined. Large fluorite solid solution regions exist at elevated temperatures. It was demonstrated that RE₂O₃-doped thoria compositions feature lower parameters p e, and higher chemical stability than the conventional stabilized ZrO2 electrolytes. The results are given in terms of the characteristic parameter p e, in the temperature range from 1000° to 1600°C. The experimental investigations were made using a new thermodynamic measuring system.     

The Function of the Histamine H4 Receptor in Inflammatory and Inflammation-Associated Diseases of the Gut

Histamine is a pleiotropic mediator involved in a broad spectrum of (patho)-physiological processes, one of which is the regulation of inflammation. Compounds acting on three out of the four known histamine receptors are approved for clinical use. These approved compounds comprise histamine H1-receptor (H₁R) antagonists, which are used to control allergic inflammation, antagonists at H₂R, which therapeutically decrease gastric acid release, and an antagonist at H₃R, which is indicated to treat narcolepsy. Ligands at H₄R are still being tested pre-clinically and in clinical trials of inflammatory diseases, including rheumatoid arthritis, asthma, dermatitis, and psoriasis. These trials, however, documented only moderate beneficial effects of H₄R ligands so far. Nevertheless, pre-clinically, H₄R still is subject of ongoing research, analyzing various inflammatory, allergic, and autoimmune diseases. During inflammatory reactions in gut tissues, histamine concentrations rise in affected areas, indicating its possible biological effect. Indeed, in histamine-deficient mice experimentally induced inflammation of the gut is reduced in comparison to that in histamine-competent mice. However, antagonists at H₁R, H₂R, and H₃R do not provide an effect on inflammation, supporting the idea that H₄R is responsible for the histamine effects. In the present review, we discuss the involvement of histamine and H₄R in inflammatory and inflammation-associated diseases of the gut.