Blickpunkt-adaptive Verschiebung der Stereo-3D-Konvergenzebene

Die Diskrepanz zwischen Akkomodation und Konvergenz beim Betrachten von Stereo-3D-Inhalten führt bei vielen Menschen zu Kopfschmerzen und weiteren Symptomen der Erschöpfung des visuellen Systems. In der Stereo-3D-Produktion werden verschiedene Maßnahmen ergriffen, diesen Effekt möglichst gering zu halten. Dies führt jedoch nicht immer zum gewünschten Erfolg oder bringt Nachteile mit sich. In diesem Dokument wird daher ein Verfahren vorgeschlagen und untersucht, welches den Konflikt zwischen Akkomodation und Konvergenz in eine andere Domäne transformiert und die Diskrepanz auf diesem Wege eliminiert. Das Verfahren funktioniert in der Art, dass die Konvergenzebene immer zu der Tiefe des momentan betrachteten virtuellen Objekts verschoben wird. Dabei ist die Konvergenzebene die virtuelle Tiefenebene, die keine Disparität hat. Das Verfahren basiert auf Blickpunktdetektion und Signalverarbeitung in Kombination mit Disparitätskarten. Dieser Artikel beschreibt die ersten Schritte in diesem Projekt

Multiplex Profiling of Cellular Invasion in 3D Cell Culture Models.

To-date, most invasion or migration assays use a modified Boyden chamber-like design to assess migration as single-cell or scratch assays on coated or uncoated planar plastic surfaces. Here, we describe a 96-well microplate-based, high-content, three-dimensional cell culture assay capable of assessing invasion dynamics and molecular signatures thereof. On applying our invasion assay, we were able to demonstrate significant effects on the invasion capacity of fibroblast cell lines, as well as primary lung fibroblasts. Administration of epidermal growth factor resulted in a substantial increase of cellular invasion, thus making this technique suitable for high-throughput pharmacological screening of novel compounds regulating invasive and migratory pathways of primary cells. Our assay also correlates cellular invasiveness to molecular events. Thus, we argue of having developed a powerful and versatile toolbox for an extensive profiling of invasive cells in a 96-well format. This will have a major impact on research in disease areas like fibrosis, metastatic cancers, or chronic inflammatory states

Gli1 mediates lung cancer cell proliferation and sonic hedgehog-dependent mesenchymal cell activation.

Non-Small-Cell-Lung-Cancer (NSCLC) represents approximately 85% of all lung cancers and remains poorly understood. While signaling pathways operative during organ development, including Sonic Hedgehog (Shh) and associated Gli transcription factors (Gli1-3), have recently been found to be reactivated in NSCLC, their functional role remains unclear. Here, we hypothesized that Shh/Gli1-3 could mediate NSCLC autonomous proliferation and epithelial/stromal signaling in the tumoral tissue. In this context, we have investigated the activity of Shh/Gli1-3 signaling in NSCLC in both, cancer and stromal cells. We report here that inhibition of Shh signaling induces a significant decrease in the proliferation of NSCLC cells. This effect is mediated by Gli1 and Gli2, but not Gli3, through regulation of cyclin D1 and cyclin D2 expression. While exogenous Shh was unable to induce signaling in either A549 lung adenocarcinoma or H520 lung squamous carcinoma cells, both cells were found to secrete Shh ligand, which induced fibroblast proliferation, survival, migration, invasion, and collagen synthesis. Furthermore, Shh secreted by NSCLC mediates the production of proangiogenic and metastatic factors in lung fibroblasts. Our results thus provide evidence that Shh plays an important role in mediating epithelial/mesenchymal crosstalk in NSCLC. While autonomous Gli activity controls NSCLC proliferation, increased Shh expression by NSCLC is associated with fibroblast activation in tumor-associated stroma. Our study highlights the relevance of studying stromal-associated cells in the context of NSCLC regarding new prognosis and therapeutic options

SNAI transcription factors mediate epithelial--mesenchymal transition in lung fibrosis

Background: Idiopathic pulmonary fibrosis (IPF) is a fatal interstitial lung disease characterised by accumulation of activated (myo)fibroblasts and excessive extracellular matrix deposition. The enhanced accumulation of (myo)fibroblasts may be attributed, in part, to the process of transforming growth factor \textgreekb1 (TGF\textgreekb1)-induced epithelial--mesenchymal transition (EMT), the phenotypic switching of epithelial to fibroblast-like cells. Although alveolar epithelial type II (ATII) cells have been shown to undergo EMT, the precise mediators and mechanisms remain to be resolved. The objective of this study is to investigate the role of SNAI transcription factors in the process of EMT and in IPF.Methods: Using quantitative reverse transcription-PCR (RT-PCR), immunofluorescence, immunohistochemistry, western blotting, as well as gain- and loss-of-function studies and functional assays, the role of SNAI1 and SNAI2 in TGF\textgreekb1-induced EMT in ATII cells in vitro was assessed; and the expression of SNAI transcription factors was analysed in experimental and human IPF in vivo.Results: TGF\textgreekb1 treatment increased the expression and nuclear accumulation of SNAI1 and SNAI2, in concert with induction of EMT in ATII cells. SNAI overexpression was sufficient to induce EMT, and small interfering RNA (siRNA)-mediated SNAI depletion attenuated TGF\textgreekb1-induced ATII cell migration and EMT. SNAI expression was elevated in experimental and human IPF and localised to hyperplastic ATII cells in vivo.Conclusions: The results demonstrate that TGF\textgreekb1-induced EMT in ATII cells is essentially controlled by the expression and nuclear translocation of SNAI transcription factors. Increased SNAI1 and SNAI2 expression in experimental and human IPF in vivo suggests that SNAI-mediated EMT may contribute to the fibroblast pool in idiopathic pulmonary fibrosis

Increased expression of 5-hydroxytryptamine(2A/B) receptors in idiopathic pulmonary fibrosis: a rationale for therapeutic intervention

Background Idiopathic pulmonary fibrosis (IPF) has a poor prognosis and limited responsiveness to available treatments. It is characterised by epithelial cell injury, fibroblast activation and proliferation and extracellular matrix deposition. Serotonin (5-hydroxytryptamine; 5-HT) induces fibroblast proliferation via the 5-HTR2A and 5-HTR2B receptors, but its pathophysiological role in IPF remains unclear. A study was undertaken to determine the expression of 5-HT receptors in IPF and experimental lung fibrosis and to investigate the effects of therapeutic inhibition of 5-HTR2A/B signalling on lung fibrosis in vivo and in vitro. Methods and results Quantitative RT-PCR showed that the expression of 5-HTR1A/B and 5-HTR2B was significantly increased in the lungs of patients with IPF (n = 12) and in those with non-specific interstitial pneumonia (NSIP, n = 6) compared with transplant donors (n = 12). The expression of 5-HTR2A was increased specifically in IPF lungs but not in NSIP lungs. While 5-HTR2A protein largely localised to fibroblasts, 5-HTR2B localised to the epithelium. To assess the effects of 5HTR(2A/B) inhibition on fibrogenesis in vivo, mice were subjected to bleomycin-induced lung fibrosis and treated with the 5-HTR2A/B antagonist terguride (or vehicle) in a therapeutic approach (days 14-28 after bleomycin). Terguride-treated mice had significantly improved lung function and histology and decreased collagen content compared with vehicle-treated mice. Functional in vitro studies showed that terguride is a potent inhibitor of transforming growth factor beta(1)- or WNT3a-induced collagen production. Conclusion The studies revealed an increased expression of 5-HTR2A specifically in IPF. Blockade of 5-HTR2A/B signalling by terguride reversed lung fibrosis and is thus a promising therapeutic approach for IPF

Phenotyping of Drosophila Melanogaster-A Nutritional Perspective

The model organism Drosophila melanogaster was increasingly applied in nutrition research in recent years. A range of methods are available for the phenotyping of D. melanogaster, which are outlined in the first part of this review. The methods include determinations of body weight, body composition, food intake, lifespan, locomotor activity, reproductive capacity and stress tolerance. In the second part, the practical application of the phenotyping of flies is demonstrated via a discussion of obese phenotypes in response to high-sugar diet (HSD) and high-fat diet (HFD) feeding. HSD feeding and HFD feeding are dietary interventions that lead to an increase in fat storage and affect carbohydrate-insulin homeostasis, lifespan, locomotor activity, reproductive capacity and stress tolerance. Furthermore, studies regarding the impacts of HSD and HFD on the transcriptome and metabolome of D. melanogaster are important for relating phenotypic changes to underlying molecular mechanisms. Overall, D. melanogaster was demonstrated to be a valuable model organism with which to examine the pathogeneses and underlying molecular mechanisms of common chronic metabolic diseases in a nutritional context

Dynamic horizontal image translation in stereo 3D

Im Bereich Stereo 3D (S3D) bezeichnet „Dynamic Horizontal Image Translation (DHIT)“ das Prinzip, die S3D-Ansichten einer Szene horizontal in entgegengesetzte Richtungen zu verschieben, wodurch die dargestellte Szene in der Tiefe verschoben wird. Dies wird vor allem im Kontext von „Active Depth Cuts“ eingesetzt. Hier werden die S3D-Ansichten vor und nach einem Szenenschnitt so verschoben, dass es nicht zu starken, störenden Tiefensprüngen kommt. Die menschliche Wahrnehmung der DHIT wurde experimentell untersucht. Eine der wichtigsten Erkenntnisse war, dass es starke individuelle Unterschiede in der Empfindlichkeit gegenüber der DHIT gibt. Daher wird empfohlen die Verschiebungsgeschwindigkeit einer S3D-Ansicht nicht höher als 0,10 °/s bis 0,12 °/s zu wählen, sodass Zuschauerinnen und Zuschauer nicht von der DHIT gestört werden. Bei der DHIT kommt es zu einer Verzerrung der dargestellten Szenentiefe. Dies wird bei dem vorgeschlagenen Ansatz „Distortion-Free Dynamic Horizontal Image Translation (DHIT+)“ kompensiert, indem der Abstand zwischen den S3D-Kameras durch Verfahren der Ansichtensynthese angepasst wird. Dieser Ansatz zeigte sich signifikant weniger störend im Vergleich zur DHIT. Die Ansichten konnten ohne Wahrnehmungsbeeinträchtigung etwa 50% schneller verschoben werden. Ein weiteres vorgeschlagenes Verfahren ist „Gaze Adaptive Convergence in Stereo 3D Applications (GACS3D)“. Unter Verwendung eines Eyetrackers wird die Disparität des geschätzten Blickpunkts langsam über die DHIT reduziert. Dies soll die Ermüdung des visuellen Systems mindern, da die Diskrepanz zwischen Akkommodation und Konvergenz reduziert wird. In einem Experiment mit emuliertem Eye-Tracking war GACS3D signifikant weniger störend als eine normale DHIT. Im Vergleich zwischen dem kompletten GACS3D-Prototypen und einer Bildsequenz ohne jegliche Verschiebungen konnte jedoch kein signifikanter Effekt auf den subjektiven Betrachterkomfort registriert werden. Eine Langzeituntersuchung der Ermüdung des visuellen Systems ist nötig, was über den Rahmen dieser Dissertation hinausgeht. Da für GACS3D eine hochgenaue Schätzung der Blickpunktdisparität benötigt wird, wurde die „Probabilistic Visual Focus Disparity Estimation“ entwickelt. Bei diesem Ansatz wird die 3D-Szenenstruktur in Echtzeit geschätzt und dazu verwendet, die Schätzung der Blickpunktdisparität deutlich zu verbessern.Dynamic horizontal image translation (DHIT) denotes the act of dynamically shifting the stereo 3D (S3D) views of a scene in opposite directions so that the portrayed scene is moved along the depth axis. This technique is predominantly used in the context of active depth cuts, where the shifting occurs just before and after a shot cut in order to mitigate depth discontinuities that would otherwise induce visual fatigue. The perception of the DHIT was investigated in an experiment. An important finding was that there are strong individual differences in the sensitivity towards DHIT. It is therefore recommended to keep the shift speed applied to each S3D view in the range of 0.10 °/s to 0.12 °/s so that nobody in the audience gets annoyed by this approach. When a DHIT is performed, the presented scene depth is distorted, i.e., compressed or stretched. A distortion-free dynamic horizontal image translation (DHIT+) is proposed that mitigates these distortions by adjusting the distance between the S3D cameras through depth-image-based rendering techniques. This approach proved to be significantly less annoying. The views could be shifted about 50% faster without perceptual side effects. Another proposed approach is called gaze adaptive convergence in stereo 3D applications (GACS3D). An eye tracker is used to estimate the visual focus whose disparity is then slowly reduced using the DHIT. This is supposed to lessen visual fatigue since the infamous accommodation vergence discrepancy is reduced. GACS3D with emulated eye tracking proved to be significantly less annoying than a regular DHIT. In a comparison between the complete prototype and a static horizontal image translation, no significant effect on subjective visual discomfort could be observed, however. A long-term evaluation of visual fatigue is necessary, which is beyond the scope of this work. In GACS3D, highly accurate visual focus disparity is required. Therefore, the probabilistic visual focus disparity estimation (PVFDE) was developed, which utilizes a real-time estimation of the 3D scene structure to improve the accuracy by orders of magnitude compared to commonly used approaches

Enolase 1 (ENO1) and protein disulfide-isomerase associated 3 (PDIA3) regulate Wnt/beta-catenin-driven trans-differentiation of murine alveolar epithelial cells

The alveolar epithelium represents a major site of tissue destruction during lung injury. It consists of alveolar epithelial type I (ATI) and type II (ATII) cells. ATII cells are capable of self-renewal and exert progenitor function for ATI cells upon alveolar epithelial injury. Cell differentiation pathways enabling this plasticity and allowing for proper repair, however, are poorly understood. Here, we applied proteomics, expression analysis and functional studies in primary murine ATII cells to identify proteins and molecular mechanisms involved in alveolar epithelial plasticity. Mass spectrometry of cultured ATII cells revealed a reduction of carbonyl reductase 2 (CBR2) and an increase in enolase 1 (ENO1) and protein disulfide-isomerase associated 3 (PDIA3) protein expression during ATII-to-ATI cell trans-differentiation. This was accompanied by increased Wnt/beta-catenin signaling, as analyzed by qRT-PCR and immunoblotting. Notably, ENO1 and PDIA3, along with T1 alpha (podoplanin;an ATI cell marker),exhibited decreased protein expression upon pharmacological and molecular Wnt/beta-catenin inhibition in cultured ATII cells, whereas CBR2 levels were stabilized. Moreover, we analyzed primary ATII cells from mice with bleomycin-induced lung injury, a model exhibiting activated Wnt/beta-catenin signaling in vivo. We observed reduced CBR2 significantly correlating with surfactant protein C (SFTPC),whereas ENO1 and PDIA3 along with T1 alpha were increased in injured ATII cells. Finally, siRNA-mediated knockdown of ENO1, as well as PDIA3, in primary ATII cells led to reduced T1 alpha expression, indicating diminished cell trans-differentiation. Our data thus identified proteins involved in ATII-to-ATI cell trans-differentiation and suggest a Wnt/beta-catenin-driven functional role of ENO1 and PDIA3 in alveolar epithelial cell plasticity in lung injury and repair

SFTA2 - a novel secretory peptide highly expressed in the lung - is modulated by lipopolysaccharide but not hyperoxia

Tissue-specific transcripts are likely to be of importance for the corresponding organ. While attempting to define the specific transcriptome of the human lung, we identified the transcript of a yet uncharacterized protein, SFTA2. In silico analyses, biochemical methods, fluorescence imaging and animal challenge experiments were employed to characterize SFTA2. Human SFTA2 is located on Chr. 6p21.33, a disease-susceptibility locus for diffuse panbronchiolitis. RT-PCR verified the abundance of SFTA2-specific transcripts in human and mouse lung. SFTA2 is synthesized as a hydrophilic precursor releasing a 59 amino acid mature peptide after cleavage of an N-terminal secretory signal. SFTA2 has no recognizable homology to other proteins while orthologues are present in all mammals. SFTA2 is a glycosylated protein and specifically expressed in nonciliated bronchiolar epithelium and type II pneumocytes. In accordance with other hydrophilic surfactant proteins, SFTA2 did not colocalize with lamellar bodies but colocalized with golgin97 and clathrin-labelled vesicles, suggesting a classical secretory pathway for its expression and secretion. In the mouse lung, Sfta2 was significantly downregulated after induction of an inflammatory reaction by intratracheal lipopolysaccharides paralleling surfactant proteins B and C but not D. Hyperoxia, however, did not alter SFTA2 mRNA levels. We have characterized SFTA2 and present it as a novel unique secretory peptide highly expressed in the lung