Artificial boundary conditions for stationary Navier-Stokes flows past bodies in the half-plane

We discuss artificial boundary conditions for stationary Navier-Stokes flows past bodies in the half-plane, for a range of low Reynolds numbers. When truncating the half-plane to a finite domain for numerical purposes, artificial boundaries appear. We present an explicit Dirichlet condition for the velocity at these boundaries in terms of an asymptotic expansion for the solution to the problem. We show a substantial increase in accuracy of the computed values for drag and lift when compared with results for traditional boundary conditions. We also analyze the qualitative behavior of the solutions in terms of the streamlines of the flow. The new boundary conditions are universal in the sense that they depend on a given body only through one constant, which can be determined in a feed-back loop as part of the solution process

Improved nonviral gene vectors: Efficient and non-toxic polyplexes with enhanced endosomolytic activity

For the development of improved polyethylenimine (PEI) polyplexes towards ‘artificial viruses’ two key issues are i) to improve the toxicity profile of the applied vectors and ii) to enhance endosomal release, one of the major barriers to efficient gene transfer with PEI polyplexes. Nonviral vectors based on PEI usually contain an excess of PEI that is not complexed to DNA. Since free PEI contributes to cellular and systemic toxicity purification of polyplexes from unbound PEI is highly desirable. In this thesis an easy and efficient method based on size exclusion chromatography (SEC) was developed, which for the first time allowed complete removal of PEI from PEI polyplexes. Moreover, purification of polyplexes enabled to clarify the role of free PEI in gene delivery at the cellular level. Most importantly, the removal of unbound PEI significantly reduced toxicity of the applied polyplexes. However, purified polyplexes without free PEI were less efficient in transfection compared to non-purified polyplexes. Mechanistic studies showed that free PEI most likely enhanced endosomal release of polyplexes and therefore contributed to efficient gene transfer with PEI polyplexes. Nevertheless, the availability of a defined, well-tolerated gene transfer formulation is a vital precondition for the further development of nonviral gene therapeutics, and a purification method like the one developed in this thesis will help to fulfill these requirements. To enhance endosomal release of PEI polyplexes, the membrane active peptide melittin was incorporated into the vector particles. It has been shown previously that PEI bound to the N-terminus of natural all-(L)-melittin (all-(L)-N-mel-PEI) enhanced gene delivery with PEI polyplexes. Here, it was demonstrated that transfection efficiency of N-mel-PEI is independent of the enantiomeric configuration of the bound peptide which allowed the use of non-immunogenic all-(D)-melittin for the generation of optimized melittin-PEI conjugates. To determine the optimal site of melittin-linkage to PEI, the polycation PEI was covalently attached to the N-terminus (N-mel-PEI) or the C-terminus of melittin (C-mel-PEI) in all-(D)-configuration. The site of melittin-linkage had a strong impact on the membrane destabilizing activities of the resulting melittin-PEI conjugates. C-mel-PEI was highly lytic at neutral pH and therefore elevated doses of C-mel-PEI polyplexes induced high toxicity. In contrast, N-mel-PEI was less lytic at neutral pH but retained higher lytic activity than C-mel-PEI at endosomal pH 5. This apparently promoted better endosomal release of N-mel-PEI polyplexes resulting in efficient gene delivery in different cell lines. The high potency of C-mel-PEI to destabilize membranes at neutral pH is presumably due to a reported destabilization mechanism proceeding through membrane insertion of the peptide. In contrast, N-mel-PEI is supposed to induce lysis by insertion-independent pore formation according to the toroidal pore model. Since membrane destabilization by membrane insertion requires lower peptide to lipid ratios than destabilization by pore formation, C-mel-PEI was considered as the more potent template to generate improved endosomolytic melittin-PEI conjugates. The new melittin-PEI conjugates should display pronounced lytic activities at endosomal pH 5. Therefore, PEI was attached to the C-terminus of melittin analogs which were modified with acidic residues. The conjugates with the highest lytic activities at endosomal pH 5 were indeed the most efficient in transfection. This apparent correlation of gene transfer efficiency with lytic activity at pH 5 was in excellent agreement with results obtained with unmodified melittin-PEI conjugates and other membrane-active peptides used in gene transfer. The most efficient melittin-PEI conjugates were incorporated into surface-shielded and receptor-targeted PEI polyplexes, and the resulting particles were further purified by SEC to remove unbound toxic polycations. Endosomolytic melittin-PEI conjugates stably incorporated into such purified polyplexes significantly enhanced transfection efficiency in comparison to polyplexes lacking melittin. Most importantly, these polyplexes exposed an improved toxicity profile providing an artificial virus-like vector that is efficient and safe also for potential in vivo administration

A New Cellular Factor Recognizes E2 Binding Sites of Papillomaviruses Which Mediate Transcriptional Repression by E2

AbstractRepression of transcription by the full-length E2 protein of papillomaviruses (PV) seems to occur when the E2 binding sites and those of positively acting cellular factors overlap. Previously, we showed that RUNX1 (formerly called CBF) binds to the repression-mediating E2 binding site P2 of human PV type 8 (HPV8). By a yeast one-hybrid system we could identify an unknown protein binding also to P2, tentatively called PBF (papillomavirus binding factor). PBF recognizes the sequence CCGG, which represents the 3′ half of the E2 binding site just adjacent to the RUNX1 motif. PBF also binds to the repression-mediating E2 BS-1 in BPV1, which is conserved to P2 of HPV8. Point mutations destroying PBF binding to HPV8 P2 and BPV-1 E2 BS-1 in vitro reduce promoter activity in corresponding reporter constructs. Our results suggest that PBF might play a role in transcription of PV genes and in E2-mediated repression

overview and meta-analysis of neuroimaging studies on motor conversion disorder

Background Conversion Disorders (CD) are prevalent functional disorders. Although the pathogenesis is still not completely understood, an interaction of genetic, neurobiological, and psychosocial factors is quite likely. The aim of this study is to provide a systematic overview on imaging studies on CDs and investigate neuronal areas involved in Motor Conversion Disorders (MCD). Methods A systematic literature search was conducted on CD. Subsequently a meta-analysis of functional neuroimaging studies on MCD was implemented using an Activation Likelihood Estimation (ALE). We calculated differences between patients and healthy controls as well as between affected versus unaffected sides in addition to an overall analysis in order to identify neuronal areas related to MCD. Results Patients with MCD differ from healthy controls in the amygdala, superior temporal lobe, retrosplenial area, primary motor cortex, insula, red nucleus, thalamus, anterior as well as dorsolateral prefrontal and frontal cortex. When comparing affected versus unaffected sides, temporal cortex, dorsal anterior cingulate cortex, supramarginal gyrus, dorsal temporal lobe, anterior insula, primary somatosensory cortex, superior frontal gyrus and anterior prefrontal as well as frontal cortex show significant differences. Conclusions Neuronal areas seem to be involved in the pathogenesis, maintenance or as a result of MCD. Areas that are important for motor-planning, motor-selection or autonomic response seem to be especially relevant. Our results support the emotional unawareness theory but also underline the need of more support by conduction imaging studies on both CD and MCD

Raven food calls indicate sender's age and sex.

BACKGROUND: Acoustic parameters of animal signals have been shown to correlate with various phenotypic characteristics of the sender. These acoustic characteristics can be learned and categorized and thus are a basis for perceivers' recognition abilities. One of the most demanding capacities is individual recognition, achievable only after repeated interactions with the same individual. Still, class-level recognition might be potentially important to perceivers who have not previously encountered callers but can classify unknown individuals according to the already learned categories. Especially for species with high fission-fusion dynamics that repeatedly encounter unknown individuals it may be advantageous to develop class-level recognition. We tested whether frequency-, temporal-, and amplitude-related acoustic parameters of vocalizations emitted by ravens, a species showing high fission-fusion dynamics in non-breeder aggregations, are connected to phenotypic characteristics and thus have the potential for class-level recognition. RESULTS: The analysis of 418 food calls revealed that some components summarizing acoustic parameters were differentiated by age-classes and sex. CONCLUSIONS: Together, the results provide evidence for the co-variation of vocal characteristics and respective sex and age categories, a prerequisite for class-level recognition in perceivers. Perceivers that are ignorant of the caller's identity can thus potentially recognize these class-level differences for decision-making processes in feeding contexts