160 research outputs found
Kombination von Diiminplatin-Fragmenten mit Catecholato-, Semichinonato- und Olefin-Liganden sowie 4-Hydroxyphenyl-, Ferrocenyl- und Bis(terpyridin)ruthenium–Bausteinen : Festphasensynthese, elektronische Strukturen, Redoxchemie und Photochemie
Im Rahmen dieser Arbeit wurden heteroleptische Diiminplatinkomplexe synthetisiert und charakterisiert. Anhand eines Dichloroplatin(II)-Komplexes wurde eine Festphasensynthese (auf der Basis eines Polystyrol-/Divinylbenzol-Copolymers, mit durch Fluoridionen spaltbarem Silyletherlinker) entwickelt und optimiert. Diese Synthesestrategie wurde anschließend auf die Synthese von ein- und mehrkernigen Catecholatoplatin(II)-Komplexen übertragen. Die Catecholatokomplexe wurden durch Oxidation in die entsprechenden Semichinonatokomplexe überführt, welche durch ESR- und UV/Vis/NIR-Spektroskopie charakterisiert wurden. Sowohl der Dichlorokomplex, als auch die Catecholatokomplexe zeigten in Lösung bei Raumtemperatur keine Lumineszenz. Deprotonierung der Komplexe führte bei den Catecholatokomplexen zu Lumineszenz, nicht jedoch beim Dichlorokomplex. Dieser Befund wurde mit Hilfe von DFT-Rechnungen erklärt, da eine Deprotonierung nur bei den Catecholatokomplexen zu einer Versteifung des Ligandensystems führt und somit möglicherweise eine strahlungslose Relaxation über eine Torsionsmode verringert wird. Durch die Kombination des Diiminfragments mit einem Ferrocenylsubstituenten und anschließender Koordination von Platinfragmenten wurden ein Dichloroplatin(II)-, ein Catecholatoplatin(II)- und ein Olefinplatin(0)-Komplex synthetisiert. Die Redoxeigenschaften dieser Komplexe wurden untersucht und aufgeklärt. Oxidation des Dichlorokomplexes mit [N(p-C6H4Br)3][SbCl6] („Magic Blue“) führte zu dem entsprechenden Ferroceniumkomplex. Einelektronenoxidation des Catecholatokomplexes führte zum Semichinonatokomplex. Ebenso wie beim Olefinkomplex führte eine Dreielektronenoxidation zur Dissoziation der Liganden und dem Entstehen des Dichlorokomplexes. Im ersten Fall wirkte „Magic Blue“ als unschuldiges Oxidationsmittel, im zweiten Fall wurde es jedoch nicht-unschuldig. Dies wurde durch NMR- und UV/Vis/NIR-Spektroskopie belegt. Das Dichlorodiiminplatin(II)-Fragment wurde mit einem photoaktiven Bis(terpyridin)ruthenium(II)-Baustein verknüpft, so dass ein potentieller Photokatalysator erhalten wurde. Tatsächlich wurde durch Bestrahlung in Gegenwart von Wasser und Triethylamin als Protonen- bzw. Elektronenlieferant eine Hydrierung von Tolan zu Stilben beobachtet. Mit Hilfe von DFT-Rechnungen wurden mögliche Wege des photoinduzierten Elektronentransfers bei dieser Reaktion analysiert
Analysis of a normalised expressed sequence tag (EST) library from a key pollinator, the bumblebee Bombus terrestris
<p>Abstract</p> <p>Background</p> <p>The bumblebee, <it>Bombus terrestris </it>(Order Hymenoptera), is of widespread importance. This species is extensively used for commercial pollination in Europe, and along with other <it>Bombus </it>spp. is a key member of natural pollinator assemblages. Furthermore, the species is studied in a wide variety of biological fields. The objective of this project was to create a <it>B. terrestris </it>EST resource that will prove to be valuable in obtaining a deeper understanding of this significant social insect.</p> <p>Results</p> <p>A normalised cDNA library was constructed from the thorax and abdomen of <it>B. terrestris </it>workers in order to enhance the discovery of rare genes. A total of 29'428 ESTs were sequenced. Subsequent clustering resulted in 13'333 unique sequences. Of these, 58.8 percent had significant similarities to known proteins, with 54.5 percent having a "best-hit" to existing Hymenoptera sequences. Comparisons with the honeybee and other insects allowed the identification of potential candidates for gene loss, pseudogene evolution, and possible incomplete annotation in the honeybee genome. Further, given the focus of much basic research and the perceived threat of disease to natural and commercial populations, the immune system of bumblebees is a particularly relevant component. Although the library is derived from unchallenged bees, we still uncover transcription of a number of immune genes spanning the principally described insect immune pathways. Additionally, the EST library provides a resource for the discovery of genetic markers that can be used in population level studies. Indeed, initial screens identified 589 simple sequence repeats and 854 potential single nucleotide polymorphisms.</p> <p>Conclusion</p> <p>The resource that these <it>B. terrestris </it>ESTs represent is valuable for ongoing work. The ESTs provide direct evidence of transcriptionally active regions, but they will also facilitate further functional genomics, gene discovery and future genome annotation. These are important aspects in obtaining a greater understanding of this key pollinator species.</p
Transnasal delivery of human A-beta peptides elicits impaired learning and memory performance in wild type mice
Background
Murine models of Alzheimer’s disease (AD) are mainly based on overexpression of pathologic amyloid precursor protein and/or presenilins. Those genes resemble underlying cause of early onset type of AD while about 99 % of all human cases are to be characterized as sporadic, late onset. Appropriate animal models for this type of AD are still missing. We here investigated, if transnasal delivery of A-beta 42 peptides might serve to mimic pathological effects in mice.
Results
A-beta 42 peptides, used for the behavioral study, showed the expected dose-dependent toxicity in neur oblastoma cell line SH-SY5Y and were able to form higher molecular weight species in vitro. Upon delivery into nostrils of wild type mice, protein bands that might represent aggregation products of the exogenously applied human A-beta 42 were only observed in total brain homogenates from mice pre-treated with mannitol. By using TAMRA-labeled A-beta 42 peptides we demonstrated, that transport throughout the brain was achieved already 1 h after administration. FVB/N mice treated with A-beta 42 for 3 days were significantly impaired in the cue-retention condition of the fear conditioning task as compared to controls whereas A-beta-treated C57B6/J mice were impaired in the context condition. In the Morris water maze test, these mice also displayed a delayed learning performance, indicated by significantly longer time to find the platform. Those deficits were also seen for memory performance in the probe trial as measured by number of crossings of the former platform position and time spent in the goal quadrant.
Conclusions
Existing AD mouse models are of genetic origin and need prolonged housing time before onset of pathology. Our short-term treatment induced learning and memory deficits via exogenous application of A-beta peptides comparable to those observed for the transgenic animals. With the transnasal A-beta 42 treatment we present an approach to investigate purely A-beta related changes suitable as a model for symptoms of Alzheimer’s dementia (AD). Resulting behavioral deficits were indicative for familial type of Alzheimer’s disease as well as for the late onset variant
Identification of disulfiram as a secretase-modulating compound with beneficial effects on Alzheimer’s disease hallmarks
ADAM10 is a metalloproteinase acting on the amyloid precursor protein (APP) as an alpha-secretase in neurons. Its enzymatic activity results in secretion of a neuroprotective APP cleavage product (sAPPalpha) and prevents formation of the amyloidogenic A-beta peptides, major hallmarks of Alzheimer’s disease (AD). Elevated ADAM10 levels appeared to contribute to attenuation of A-beta-Plaque formation and learning and memory deficits in AD mouse models. Therefore, it has been assumed that ADAM10 might represent a valuable target in AD therapy. Here we screened a FDA-approved drug library and identified disulfiram as a novel ADAM10 gene expression enhancer. Disulfiram increased ADAM10 production as well as sAPP-alpha in SH-SY5Y human neuronal cells and additionally prevented A-beta aggregation in an in vitro assay in a dose-dependent fashion. In addition, acute disulfiram treatment of Alzheimer model mice induced ADAM10 expression in peripheral blood cells, reduced plaque-burden in the dentate gyrus and ameliorated behavioral deficits. Alcohol-dependent patients are subjected to disulfiram-treatment to discourage alcohol-consumption. In such patients, enhancement of ADAM10 by disulfiram-treatment was demonstrated in peripheral blood cells. Our data suggest that disulfiram could be repurposed as an ADAM10 enhancer and AD therapeutic. However, efficacy and safety has to be analyzed in Alzheimer patients in the future
A novel blood-brain barrier co-culture System for drug targeting of Alzheimer’s disease : establishment by using acitretin as a model drug
In the pathogenesis of Alzheimer’s disease (AD) the homeostasis of amyloid precursor protein (APP) processing in the brain is impaired. The expression of the competing proteases ADAM10 (a disintegrin and metalloproteinase 10) and BACE-1 (beta site APP cleaving enzyme 1) is shifted in favor of the A-beta generating enzyme BACE-1. Acitretin–a synthetic retinoid–e.g., has been shown to increase ADAM10 gene expression, resulting in a decreased level of A-beta peptides within the brain of AD model mice and thus is of possible value for AD therapy. A striking challenge in evaluating novel therapeutically applicable drugs is the analysis of their potential to overcome the blood-brain barrier (BBB) for central nervous system targeting. In this study, we established a novel cell-based bio-assay model to test ADAM10-inducing drugs for their ability to cross the BBB. We therefore used primary porcine brain endothelial cells (PBECs) and human neuroblastoma cells (SH-SY5Y) transfected with an ADAM10-promoter luciferase reporter vector in an indirect co-culture system. Acitretin served as a model substance that crosses the BBB and induces ADAM10 expression. We ensured that ADAM10-dependent constitutive APP metabolism in the neuronal cells was unaffected under co-cultivation conditions. Barrier properties established by PBECs were augmented by co-cultivation with SH-SY5Y cells and they remained stable during the treatment with acitretin as demonstrated by electrical resistance measurement and permeability-coefficient determination. As a consequence of transcellular acitretin transport measured by HPLC, the activity of the ADAM10-promoter reporter gene was significantly increased in co-cultured neuronal cells as compared to vehicle-treated controls. In the present study, we provide a new bio-assay system relevant for the study of dru
Identification of disulfiram as a secretase-modulating compound with beneficial effects on Alzheimer’s disease hallmarks
ADAM10 is a metalloproteinase acting on the amyloid precursor protein (APP) as an alpha-secretase in neurons. Its enzymatic activity results in secretion of a neuroprotective APP cleavage product (sAPPalpha) and prevents formation of the amyloidogenic A-beta peptides, major hallmarks of Alzheimer’s disease (AD). Elevated ADAM10 levels appeared to contribute to attenuation of A-beta-Plaque formation and learning and memory deficits in AD mouse models. Therefore, it has been assumed that ADAM10 might represent a valuable target in AD therapy. Here we screened a FDA-approved drug library and identified disulfiram as a novel ADAM10 gene expression enhancer. Disulfiram increased ADAM10 production as well as sAPP-alpha in SH-SY5Y human neuronal cells and additionally prevented A-beta aggregation in an in vitro assay in a dose-dependent fashion. In addition, acute disulfiram treatment of Alzheimer model mice induced ADAM10 expression in peripheral blood cells, reduced plaque-burden in the dentate gyrus and ameliorated behavioral deficits. Alcohol-dependent patients are subjected to disulfiram-treatment to discourage alcohol-consumption. In such patients, enhancement of ADAM10 by disulfiram-treatment was demonstrated in peripheral blood cells. Our data suggest that disulfiram could be repurposed as an ADAM10 enhancer and AD therapeutic. However, efficacy and safety has to be analyzed in Alzheimer patients in the future
Absence of autoantibodies against correctly folded recombinant fibrillin-1 protein in systemic sclerosis patients
Autoantibodies against short recombinant fragments of fibrillin-1 produced in bacterial expression systems have been found in tight-skin mouse, systemic sclerosis, mixed connective tissue disease, and primary pulmonary hypertension syndrome. In patients with scleroderma, the frequency of anti-fibrillin-1 antibodies was 42% in Caucasians. Until now it has been unclear whether this immune response has a primary function in disease pathogenesis or is a secondary phenomenon. In the present study we analyzed the frequency of autoantibodies against two overlapping recombinant polypeptides spanning the N-terminal and C-terminal halves of human fibrillin-1, which were produced in human embryonic kidney (HEK-293) cells. Correct three-dimensional structures of the recombinant fibrillin-1 polypeptides were shown by electron microscopy and immunoreactivity with antibodies. Screening of fibrillin-1 antibodies was performed in 41 sera from systemic sclerosis patients and in 44 healthy controls with a Caucasian background. Microtiter plates were coated with the recombinant polypeptides of fibrillin-1 and incubated with 1:100 diluted sera. Positive binding was defined as being more than 2 SD above the mean of the control group. ELISAs showed that none of the sera of patients with systemic sclerosis contained autoantibodies against the N-terminal or C-terminal recombinant fibrillin-1 polypeptide. The data show the absence of autoantibodies against recombinant fibrillin-1 protein in Caucasian systemic sclerosis patients. Because the correct three-dimensional folding of the recombinant proteins has been substantiated by several independent methods, we conclude that autoantibodies against correctly folded fibrillin are not a primary phenomenon in the pathogenesis of systemic sclerosis
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