Engel T.,Westphalian Wilhelms University |
Fobker M.,University Hospital Muenster |
Buchmann J.,German Institute of Human Nutrition |
Kannenberg F.,University Hospital Muenster |
And 4 more authors.
Atherosclerosis | Year: 2014
Objective: Oxysterols are oxidized derivatives of sterols that have cytotoxic effects and are potent regulators of diverse cellular functions. Efficient oxysterol removal by the sub-family G member 1 of the ATP-binding cassette transporters (ABCG1) is essential for cell survival and control of cellular processes. However, the specific role of ABCG1 in the transport of various oxysterol species has been not systematically investigated to date. Here, we examined the involvement of ABCG1 in the oxysterol metabolism by studying oxysterol tissue levels in a mouse model of Abcg1-deficiency. Methods and results: Analysis of lung tissue of Abcg1-/- mice on a standard diet revealed that 3β,5α,6β-cholestanetriol (CT) and 25-hydroxycholesterol (HC) accumulated at more than 100-fold higher levels in comparison to wild-type mice. 24S-HC and 27-HC levels were also elevated, but were minor constituents. A radiolabeled assay employing regulable ABCG1-expressing HeLa cell lines revealed that 25-HC export to albumin was dependent on functional ABCG1 expression and could be blocked by an excess of unlabeled 25-HC or 27-HC. In this cell line, 25-HC at low doses triggered mitochondrial membrane potential, and reactive oxygen species production, which are both indirect indicators of cellular energy expenditure. Conclusion: Our results suggest that 25-HC and CT are physiologic substrates for ABCG1. Excessive accumulation of these oxysterols may explain the increased rate of cell death and the inflammatory phenotype observed in Abcg1-deficient animals and cells. © 2014 Elsevier Ireland Ltd.
News Article | January 4, 2016
The earth is 4.57 billion years old – an unimaginable temporal dimension. To understand how the blue planet was first formed long ago, scientists today are analyzing other rock bodies from our solar system, such as fragments of asteroids that have arrived on Earth as meteorites after collisions in space. According to current knowledge, many planetary bodies were formed through the merger of chondrules – which are silicate beads that are about 0.1 to 3 mm wide. How does this cosmic rock formation process work, though? That is what scientists from the Institute of Planetology at the Westphalian Wilhelms University of Münster and the Technical University of Braunschweig are investigating in unique experiments. They are being supported by researchers at the Fraunhofer Institute for Silicate Research ISC in Würzburg. The scientists have developed a special glass for the project and formed tiny beads from it to represent the chondrules as realistically as possible. Previous findings indicate that the original particles had the consistency of hot, liquid glass before they aggregated into larger conglomerates of rock, cooled down and crystallized. "This glass is very different from the material composition of technical glasses with which we are usually working," explains Dr. Martin Kilo, Head of Glass Unit at the ISC. The chemical composition of a glass determines certain physical properties, though, such as the melting and crystallization behavior. Both play a central role in the development process of larger rock bodies. "That's why we have used modeling programs in advance to calculate which melting conditions prevail for the required compositions, how stable the glass particles are, as well as the temperatures and forms at which they crystallize," says Dr. Kilo. Another challenge was to give the glass particles their spherical shape. To do so, the experts use two different procedures. In the first approach, rough glass gravel is prepared, sifted to the right size and then rounded out by thermal treatment. The second solution is to cut glass plates into small cubes and to grind them mechanically – very similar to the marble production. For the experiment, the researchers from Würzburg produced several versions of their beads, each of which differs slightly in material composition. These beads were first heated in special melting units in which the temperature and atmosphere can be adjusted precisely. Those beads which had characteristics closest to the theoretical model after this test melting were selected for the project. The research team from the Universities of Münster and Braunschweig now uses the cosmic glass beads from the ISC in experiments at the Center for Applied Space Technology and Microgravity (ZARM) in Bremen: The drop tower which is operated there surrounds a 120-meter-high steel drop tube, in which a high-vacuum is kept. Through a catapult system, the glass beads are shot in a capsule to the tip of the drop tube. As a result, approximately 9.5 seconds of weightlessness are achieved – the same conditions as in space. During this period, the glass beads are heated up to 1100 °C. During the dropping procedure, the beads can collide and form larger clusters. The experts record the collision behavior with high-speed cameras that colleagues at the TU Braunschweig assess. "Our colleagues from Münster then investigate how the beads merge, whether the clusters are composed of a homogeneous composition or whether the form of the individual beads is still recognizable, and whether and to what extent crystallization results," Dr. Kilo explains. In the next step, the planetologists will compare the results with observations of meteorites to then draw conclusions about the validity of their theoretical models.
Bednarski C.,Albert Ludwigs University of Freiburg |
Tomczak K.,Westphalian Wilhelms University |
Hovel B.V.,Albert Ludwigs University of Freiburg |
Weber W.-M.,Westphalian Wilhelms University |
Cathomen T.,Albert Ludwigs University of Freiburg
PLoS ONE | Year: 2016
In vitro disease models have enabled insights into the pathophysiology of human disease as well as the functional evaluation of new therapies, such as novel genome engineering strategies. In the context of cystic fibrosis (CF), various cellular disease models have been established in recent years, including organoids based on induced pluripotent stem cell technology that allowed for functional readouts of CFTR activity. Yet, many of these in vitro CF models require complex and expensive culturing protocols that are difficult to implement and may not be amenable for high throughput screens. Here, we show that a simple cellular CF disease model based on the bronchial epithelial δF508 cell line CFBE41o-can be used to validate functional CFTR correction. We used an engineered nuclease to target the integration of a super-exon, encompassing the sequences of CFTR exons 11 to 27, into exon 11 and re-Activated endogenous CFTR expression by treating CFBE41o-cells with a demethylating agent. We demonstrate that the integration of this super-exon resulted in expression of a corrected mRNA from the endogenous CFTR promoter and used short-circuit current measurements in Ussing chambers to corroborate restored ion transport of the repaired CFTR channels. In conclusion, this study proves that the targeted integration of a large super-exon in CFTR exon 11 leads to functional correction of CFTR, suggesting that this strategy can be used to functionally correct all CFTR mutations located downstream of the 5' end of exon 11. © 2016 Adel et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Hein S.,Julius Kuhn Institute |
Hein S.,Westphalian Wilhelms University |
Jacob J.,Julius Kuhn Institute
Wildlife Research | Year: 2015
In this review we summarise published knowledge regarding small mammal population recovery following sudden population collapse, regardless as to whether the collapse is caused by natural or man-made events. We determine recovery mechanisms, recovery time and recovery rate, and suggest how to adapt and optimise current methods to regulate small mammal population size, for pest management and/or conservation. It is vital that the principles underlying the recovery mechanisms are known for both pest control and conservation to align management methods to either maintain animal numbers at a permanent minimum level or increase population size. Collapses can be caused naturally, as in the declining phase of multi-annual fluctuations and after natural disasters, or by man-made events, such as pesticide application. In general, there are three ways population recovery can occur: (1) in situ survival and multiplication of a small remaining fraction of the population; (2) immigration; or (3) a combination of the two. The recovery mechanism strongly depends on life history strategy, social behaviour and density-dependent processes in population dynamics of the species in question. In addition, the kind of disturbance, its intensity and spatial scale, as well as environmental circumstances (e.g. the presence and distance of refuge areas) have to be taken into account. Recovery time can vary from a couple of days to several years depending on the reproductive potential of the species and the type of disturbances, regardless of whether the collapse is man made or natural. Ultimately, most populations rebound to levels equal to numbers before the collapse. Based on current knowledge, case-by-case decisions seem appropriate for small-scale conservation. For pest control, a large-scale approach seems necessary. Further investigations are required to make sound, species-specific recommendations. © 2015 CSIRO.
Berger L.,Vienna University of Technology |
Eichler J.,Medical University of Graz |
Ryll E.J.S.,Westphalian Wilhelms University |
Fischerauer S.,Auenbruggerplatz |
And 4 more authors.
Materials Science and Engineering C | Year: 2016
Flexible intramedullary nailing (FIN) is a minimally invasive and widespread standard method for osteosynthesis of pediatric long bone fractures. In the case of unstable fractures of the lower extremity, interlocking systems need to be used to prevent axial shortening and subsequent perforation of the nail at its insertion site. In the present study, four different screw-fixed interlocking systems for FINs (Hofer TwinPlug with two 3-mm titanium interlocking screws, Hofer FixPlug with 3-mm titanium interlocking screw, Hofer Plug with 3.5-mm titanium interlocking screw, and Hofer Plug with 3-mm titanium interlocking screw) in comparison with the commonly used Ender stainless steel nails (locked with 3.5-mm screw) were experimentally investigated in cadaveric lamb tibiae, regarding their load characteristics and failure modes in the case of heavy loading. The specimens were subjected to sequential axial cyclic loading of 5000cycles with stepwise increase of the load amplitude until failure. Migration of locking screws and internal damage of bone tissue was quantified by micro-computed tomography (CT) imaging. Ender nails failed on average at a peak load of 800 N, TwinPlugs at 1367 N, FixPlugs at 1222 N, Plugs 3.5mm at 1225 N and Plugs 3.0mm at 971 N. TwinPlugs, FixPlugs, and Plugs 3.5mm failed in a slow manner over several hundred loading cycles, whereas Ender nails and Plugs 3.0mm exhibited abrupt failure without any prior indication. Our results confirm that axial stability of FIN can be further improved by screw-fixed plugs by simultaneously avoiding shortcomings of an eye-locked system, which the Ender nails are. Considering biomechanical results, plug interlocking systems with 3.5-mm screws should be favored over conventional Ender nails and plugs with 3-mm screws. © 2016 Elsevier B.V. All rights reserved.
Clark K.L.,Glaxosmithkline |
Hughes S.A.,Glaxosmithkline |
Bulsara P.,Glaxosmithkline |
Coates J.,Glaxosmithkline |
And 20 more authors.
Molecular Therapy - Nucleic Acids | Year: 2013
Lung pathology in cystic fibrosis is linked to dehydration of the airways epithelial surface which in part results from inappropriately raised sodium reabsorption through the epithelial sodium channel (ENaC). To identify a small-interfering RNA (siRNA) which selectively inhibits ENaC expression, chemically modified 21-mer siRNAs targeting human ENaCα were designed and screened. GSK2225745, was identified as a potent inhibitor of ENaCα mRNA (EC50 (half maximal effective concentration) = 0.4 nmol/l, maximum knockdown = 85%) and protein levels in A549 cells. Engagement of the RNA interference (RNAi) pathway was confirmed using 5′ RACE. Further profiling was carried out in therapeutically relevant human primary cells. In bronchial epithelial cells, GSK2225745 elicited potent suppression of ENaCα mRNA (EC50 = 1.6 nmol/l, maximum knockdown = 82%). In human nasal epithelial cells, GSK2225745 also produced potent and long-lasting (≥72 hours) suppression of ENaCα mRNA levels which was associated with significant inhibition of ENaC function (69% inhibition of amiloride-sensitive current in cells treated with GSK2225745 at 10 nmol/l). GSK2225745 showed no evidence for potential to stimulate toll-like receptor (TLR)3, 7 or 8. In vivo, topical delivery of GSK2225745 in a lipid nanoparticle formulation to the airways of mice resulted in significant inhibition of the expression of ENaCα in the lungs. In conclusion, GSK2225745 is a potent inhibitor of ENaCα expression and warrants further evaluation as a potential novel inhaled therapeutic for cystic fibrosis. © 2012 American Society of Gene & Cell Therapy All rights reserved.
Leier G.,Westphalian Wilhelms University |
Bangel-Ruland N.,Westphalian Wilhelms University |
Sobczak K.,Westphalian Wilhelms University |
Knieper Y.,Westphalian Wilhelms University |
Weber W.-M.,Westphalian Wilhelms University
Cellular Physiology and Biochemistry | Year: 2012
The phosphodiesterase-5 inhibitor sildenafil is an established and approved drug to treat symptoms of a variety of human diseases. In the context of cystic fibrosis (CF), a genetic disease caused by a defective CFTR gene (e.g. ΔF508-CFTR), it was assumed that sildenafil could be a promising substance to correct impaired protein expression. This study focuses on the molecular mechanisms of sildenafil on CFTR recovery. We used ΔF508-CFTR/wt-CFTR expressing Xenopus laevis oocytes and human bronchial epithelial cell lines (CFBE41o-/16HBE14o-) to investigate the pathways of sildenafil action. Cells were treated with sildenafil and cAMP-mediated current (Im), conductance (Gm), and capacitance (Cm) were determined. Sildenafil increased Im, Gm, and Cm of wt-CFTR and functionally restored ΔF508-CFTR in oocytes. These effects were also seen in CFBE41oand 16HBE14o-cells. Transepithelial measurements revealed that sildenafil mediated increase (wt-CFTR) and restoration (ΔF508-CFTR) of channel activity. cGMP pathway blocker inhibited the activity increase but not CFTR/ΔF508-CFTR exocytosis. From these data we conclude that sildenafil mediates potentiation of CFTR activity by a cGMP-dependent and initiates cGMP-independent functional insertion of CFTR/ΔF508-CFTR molecules into the apical membranes. Thus, sildenafil is a corrector and potentiator of CFTR/ΔF508-CFTR. Yet, the necessary high doses of the drug for CFTR recovery demonstrate that sildenafil might not be suited as a therapeutic drug for CF lung disease. Copyright © 2012 S. Karger AG, Basel.
PubMed | Westphalian Wilhelms University
Type: Journal Article | Journal: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology | Year: 2012
The phosphodiesterase-5 inhibitor sildenafil is an established and approved drug to treat symptoms of a variety of human diseases. In the context of cystic fibrosis (CF), a genetic disease caused by a defective CFTR gene (e.g. F508-CFTR), it was assumed that sildenafil could be a promising substance to correct impaired protein expression. This study focuses on the molecular mechanisms of sildenafil on CFTR recovery. We used F508-CFTR/wt-CFTR expressing Xenopus laevis oocytes and human bronchial epithelial cell lines (CFBE41o(-)/16HBE14o(-)) to investigate the pathways of sildenafil action. Cells were treated with sildenafil and cAMP-mediated current (I(m)), conductance (G(m)), and capacitance (C(m)) were determined. Sildenafil increased I(m), G(m), and C(m) of wt-CFTR and functionally restored F508-CFTR in oocytes. These effects were also seen in CFBE41o(-) and 16HBE14o(-) cells. Transepithelial measurements revealed that sildenafil mediated increase (wt-CFTR) and restoration (F508-CFTR) of channel activity. cGMP pathway blocker inhibited the activity increase but not CFTR/F508-CFTR exocytosis. From these data we conclude that sildenafil mediates potentiation of CFTR activity by a cGMP-dependent and initiates cGMP-independent functional insertion of CFTR/F508-CFTR molecules into the apical membranes. Thus, sildenafil is a corrector and potentiator of CFTR/F508-CFTR. Yet, the necessary high doses of the drug for CFTR recovery demonstrate that sildenafil might not be suited as a therapeutic drug for CF lung disease.