Implication of apoE isoforms in cholesterol metabolism by primary rat hippocampal neurons and astrocytes

Apolipoprotein E (apoE) has been genetically linked to late-onset Alzheimer's disease. From the three common alleles (epsilon2, epsilon3 and epsilon4), epsilon4 has been suggested to promote amyloid beta (Ass) plaque fibrillation, one hallmark of Alzheimer's disease. It has been demonstrated that altered lipid content of hippocampal plasma membrane coincides with the disease. In this study, we show for the first time that the apoE dependent cholesterol metabolism in hippocampal neurons is higher than that of hippocampal astrocytes. Further, apoE-bound cholesterol is highly incorporated in membranous compartments in hippocampal neurons, whereas hippocampal astrocytes show higher intracellular distribution. This is an effect that coincides with cell-type dependent difference of low density lipoprotein receptor (LDLR) family member expression. Hippocampal neurons express high levels of the LDLR related protein (LRP), whereas hippocampal astrocytes are highly positive for LDLR. We could also demonstrate an apoE isoform (apoE2, apoE3 and apoE4) dependent cholesterol uptake in both cells types. In hippocampal neurons, we could find a decreased apoE4-bound cholesterol uptake. In contrast, hippocampal astrocytes show decreased internalization of apoE2-bound cholesterol. In addition, lipidated apoE4 is little associated with neurites in hippocampal neurons in comparison to the other two isoforms. In contrary, hippocampal astrocytes show faint apoE2 immunocytostaining intensity. Data presented indicate that the role of apoE4 in cholesterol homeostasis and apolipoprotein cell association is more pronounced in hippocampal neurons, showing significant alterations compared to the other two isoforms, suggesting that hippocampal neurons are affected by apoE4 associated altered cholesterol metabolism compared to hippocampal astrocytes.     

Identification and characterization of keyhole limpet hemocyanin N-glycans mediating cross-reactivity with Schistosoma mansoni

Keyhole limpet hemocyanin (KLH) of the mollusc Megathura crenulata is known to serologically cross-react with Schistosoma mansoni glycoconjugates in a carbohydrate-dependent manner. To elucidate the structural basis for this cross-reactivity, KLH glycans were released from tryptic glycopeptides and fluorescently labeled. Cross-reacting glycans were identified using a polyclonal antiserum reacting with soluble S. mansoni egg antigens, isolated by a three-dimensional fractionation scheme and analyzed by different mass spectrometric techniques as well as linkage analysis and exoglycosidase treatment. The results revealed that cross-reacting species comprise approximately 4.5% of released glycans. They all represent novel types of N-glycans with a Fuc(alpha1-3)GalNAc(beta1-4)[Fuc(alpha1-3)]GlcNAc motif, which is known to occur also in schistosomal glycoconjugates. The tetrasaccharide unit is attached to the 3-linked antenna of a trimannosyl core, which can be further decorated by galactosyl residues, a xylose residue in 2-position of the central mannose and/or a fucose at the innermost N-acetylglucosamine. This study provides for the first time detailed structural data on the KLH carbohydrate entities responsible for cross-reactivity with glycoconjugates from S. mansoni.     

Large-scale production of a therapeutic protein in transgenic tobacco plants: effect of subcellular targeting on quality of a recombinant dog gastric lipase

A recombinant dog gastric lipase with therapeutic potential for the treatment of exocrine pancreatic insufficiency was expressed in transgenic tobacco plants. We targeted the protein using two different signal sequences for either vacuolar retention or secretion. In both cases, an active glycosylated recombinant protein was obtained. The recombinant enzymes and the native enzyme displayed similar properties including acid resistance and acidic optimum pH. The proteolytic maturation and the specific activity of the recombinant proteins, however, were found to be dependent on subcellular compartmentalization. Expression levels of recombinant dog gastric lipase were about 5% and 7% of acid extractable plant proteins for vacuolar retention and secretion respectively. This expression system already has allowed the production of tens of grams of purified lipase through open-field culture of transgenic tobacco plants.     

Functional epitope of common gamma chain for interleukin-4 binding.

Interleukin 4 (IL-4) can act on target cells through an IL-4 receptor complex consisting of the IL-4 receptor alpha chain and the common gamma chain (gamma(c)). An IL-4 epitope for gamma(c) binding has previously been identified. In this study, the gamma(c) residues involved in IL-4 binding were defined by alanine-scanning mutational analysis. The epitope comprises gamma(c) residues I100, L102, and Y103 on loop EF1 together with L208 on loop FG2 as the major binding determinants. These predominantly hydrophobic determinants interact with the hydrophobic IL-4 epitope composed of residues I11, N15, and Y124. Double-mutant cycle analysis revealed co-operative interaction between gamma(c) and IL-4 side chains. Several gamma(c) residues involved in IL-4 binding have been previously shown to be mutated in X-linked severe combined immunodeficiency. The importance of these binding residues for gamma(c) function is discussed. These results provide a basis for elucidating the molecular recognition mechanism in the IL-4 receptor system and a paradigm for other gamma(c)-dependent cytokine receptor systems.     

Zur Frage der Selektion von Kindern in ABO-blutgruppenunverträglichen Ehen

Zusammenfassung In der Annahme, daß Blutgruppeninkompatibilität sich häufig erst nach Immunisierung der Mutter durch eine vorausgegangene Schwangerschaft für die nachfolgenden Kinder deletär auswirken dürfte, kamen wir zu der Vermutung, daß die der Selektion verfallenen Blutgruppen bei nachgeborenen Kindern seltener angetroffen werden müßten als bei Erstgeburten, während vor allem die Blutgruppe O eher vermehrt zu finden wäre. Wir haben daher die Blutgruppen-verteiling bei 1186 estgeborenen derjenigen von 2112 nachgeborenen Kindern gegenübergestellt. Es zeigt sich, daß die Blutgruppe O bei den weiteren Kindern häufiger vorkommt als bei den erstgeborenen, die Blutgruppen B and AB dagegen seltener. Ein vermehrter Ausfall von Knabengeburten als Folge der größeren Anfälligkeit des männlichen Geschlechtes scheint nach unseren Ermittlungen nur für Erstgeburten zuzutreffen. Gleichartige Veränderungen der Häufigkeitsverteilung Rh-positiver Individuen scheinen in viel geringerem Ausmaße vorzukommen.

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The ABO blood group distribution was compared in 1186 first born and 2112 later born children. Among the later born children group 0 is significantly more frequent, whereas groups B and AB are rarer. Additionally, the number of firstborn males seems to be reduced due to selection. The influence of selection by incompatibility seems to be smaller in the Rh than in the ABO system.

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Ärztl. Leiter: Prim. Dr. W. Kircher     

Intensification of agriculture in southwestern Germany between the Bronze Age and Medieval period,based on archaeobotanical data from Baden-Württemberg

Vegetation History and Archaeobotany - A system of farming with an alternation of land use between being cultivated or left fallow as grassland (Feldgraswirtschaft) developed in southwestern...     

Streptomonospora litoralis sp. nov., a halophilic thiopeptides producer isolated from sand collected at Cuxhaven beach

Antonie van Leeuwenhoek - Strain M2T was isolated from the beach of Cuxhaven, Wadden Sea, Germany, in course of a program to attain new producers of bioactive natural products. Strain M2T produces...     

Arguments against the significance of the Fenton reaction contributing to signal pathways under in vivo conditions

One of the common explanations for oxidative stress in the physiological milieu is based on the Fenton reaction, i.e. the assumption that radical chain reactions are initiated by metal-catalyzed electron transfer to hydrogen peroxide yielding hydroxyl radicals. On the other hand — especially in the context of so-called “iron switches” — it is postulated that cellular signaling pathways originate from the interaction of reduced iron with hydrogen peroxide.

Using fluorescence detection and EPR for identification of radical intermediates, we determined the rate of iron complexation by physiological buffer together with the reaction rate of concomitant hydroxylations of aromatic compounds under aerobic and anaerobic conditions. With the obtained overall reaction rate of 1,700 M^-1s^-1 for the buffer-dependent reactions and the known rates for Fenton reactions, we derive estimates for the relative reaction probabilities of both processes.

As a consequence we suggest that under in vivo conditions initiation of chain reactions by hydroxyl radicals generated by the Fenton reaction is of minor importance and hence metal-dependent oxidative stress must be rather independent of the so-called “peroxide tone”. Furthermore, it is proposed that — in the low (subtoxic) concentration range — hydroxylated compounds derived from reactions of “non-free” (crypto) OH radicals are better candidates for iron-dependent sensing of redox-states and for explaining the origin of cellular signals than the generation of “free” hydroxyl radicals.     

Culture of a Gastric Non‐Helicobacter pylori Helicobacter from the Stomach of a 14‐Year‐Old Girl

Helicobacter felis belongs to the fastidious gastric non‐Helicobacter pylori helicobacter species that are typically found in the stomach of cats and dogs. These bacteria have the potential to colonize the human stomach and are then associated with gastritis, gastroduodenal ulcers, and MALT lymphoma. Strains cultured from the human stomach are rare. Here, we present the first isolation of H. felis from a gastric biopsy specimen of a 14‐year‐old girl who presented with persistent epigastric pain. The strain was cultured using our routine protocol for H. pylori and identified by phylogenetic analyses of partial urease AB and gyrB gene sequences.