Mucopolysaccharidosis-like alterations in cardiac valves of rats treated with tilorone

The purpose of this study was to examine whether the aortic and mitral valves of rats are involved in the mucopolysaccharidosis-like disorder induced by tilorone. Rats were treated with large doses of the drug for periods of 1-21 weeks. After chronic drug treatment the leaflets of both heart valves were thickened and opaque. In all treated animals the spongiosa layer of the stroma was crowded with vacuolated cells; the fibrosa layer was altered only after prolonged treatment. Ultrastructurally, the vacuolated cells of the spongiosa could be identified as histiocytes and fibroblasts, the former being the most susceptible cell type. The fibroblasts of the fibrosa represented the least sensitive cell type. The histochemical results showed that the clear cytoplasmic vacuoles in the spongiosa cells were due to lysosomal storage of polyanionic material with staining characteristics similar to cartilage matrix. After discontinuation of drug treatment the alterations persisted for several weeks. The present study shows that heart valves are involved in the mucopolysaccharidosis-like disorder induced by tilorone. The molecular pathomechanism of the disorder and the exact identification of the storage material must await further analysis.     

Tactile localisation: the function of active antennal movements in the crayfishCherax destructor

Video recordings and single frame analysis were used to study the function of the second antennae of crayfish (Cherax destructor) as a sensory system in freely behaving animals. Walking crayfish move their antennae back and forth through horizontal angles of 100 degrees and more, relative to the body long axis. At rest, animals tend to hold their antennae at angular positions between 20 and 50 degrees. Movements of the two antennae are largely independent of each other. Before and during a turn of the body the ipsilateral antenna is moved into the direction of the turn. Solid objects are explored by repeatedly moving the antennae towards and across them. Both seeing and blinded crayfish can locate stationary objects following antennal contact. On antennal contact with a small novel object, a moving animal withdraws its antenna and attacks the object. When the antenna of a blinded crayfish is lightly touched with a brush the animal turns and attacks the point of stimulation. The direction taken and the distance covered during an attack can be correlated with: the angle at which the antenna is held at the moment of contact and the distance along the antennal flagellum at which the stimulus is applied. From behavioural evidence we conclude that crayfish use information about the angular position of their antennae and about the position of stimulated mechanoreceptors along the antennal flagellum to locate objects in their environment. We suggest ways in which an active tactile system-like the crayfish's antennae--could supply animals with information about the three-dimensional layout of their environment.     

Expression of membrane-bound carbonic anhydrases IV, IX, and XIV in the mouse heart.

Expression of membrane-bound carbonic anhydrases (CAs) of CA IV, CA IX, CA XII, and CA XIV has been investigated in the mouse heart. Western blots using microsomal membranes of wild-type hearts demonstrate a 39-, 43-, and 54-kDa band representing CA IV, CA IX, and CA XIV, respectively, but CA XII could not be detected. Expression of CA IX in the CA IV/CA XIV knockout animals was further confirmed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Cardiac cells were immunostained using anti-CA/FITC and anti-alpha-actinin/TRITC, as well as anti-CA/FITC and anti-SERCA2/TRITC. Subcellular CA localization was investigated by confocal laser scanning microscopy. CA localization in the sarcolemmal (SL) membrane was examined by double immunostaining using anti-CA/FITC and anti-MCT-1/TRITC. CAs showed a distinct distribution pattern in the sarcoplasmic reticulum (SR) membrane. CA XIV is predominantly localized in the longitudinal SR, whereas CA IX is mainly expressed in the terminal SR/t-tubular region. CA IV is present in both SR regions, whereas CA XII is not found in the SR. In the SL membrane, only CA IV and CA XIV are present. We conclude that CA IV and CA XIV are associated with the SR as well as with the SL membrane, CA IX is located in the terminal SR/t-tubular region, and CA XII is not present in the mouse heart. Therefore, the unique subcellular localization of CA IX and CA XIV in cardiac myocytes suggests different functions of both enzymes in excitation-contraction coupling.     

Regulation of gene expression by photosynthetic signals triggered through modified CO2 availability

Background  

To coordinate metabolite fluxes and energy availability, plants adjust metabolism and gene expression to environmental changes through employment of interacting signalling pathways.     

Maintaining homeostasis by controlled alternatives for energy distribution in plant cells under changing conditions of supply and demand

Plants depend on light energy for the generation of ATP and reductant as well as on supply of nutrients (inorganic C, N, and S compounds) to successfully produce biomass. Any excess of reducing power or lack of electron acceptors can lead to the formation of reactive oxygen species (ROS). Multiple systems are operating to avoid imbalances and subsequent oxidative stress by efficiently scavenging any formed ROS. Plants can sense an upcoming imbalance and correspondingly adapt to changed conditions not only by an increase of ROS scavengers, but also by using excess incoming light energy productively for assimilatory processes in actively metabolizing cells of growing leaves. CO₂ assimilation in chloroplasts is controlled by various redox-regulated enzymes; their activation state is strictly linked to metabolism due to the effects of small molecules on their actual activation state. Shuttle systems for indirect transfer of reducing equivalents and ATP specifically distribute the energy fluxes between compartments for optimal biomass production. Integration of metabolic and redox signals involves the cytosolic enzyme glyceraldehyde-3-P dehydrogenase (GapC) and some of its many moonlighting functions. Its redox- and metabolite-dependent interactions with the mitochondrial outer membrane, the cytoskeleton, and its occurrence in the nucleus are examples of these additional functions. Induction of the genes required to achieve an optimal response suitable for the respective conditions allows for growth when plants are exposed to different light intensities and nutrient conditions with varying rates of energy input and different assimilatory pathways for its consumption are the required in the long term. A plant-specific respiratory pathway, the alternative oxidase (AOX), functions as a site to convert excess electrons into heat. For acclimation, any imbalance is sensed and elicits signal transduction to induce the required genes. Examples for regulated steps in this sequence of events are given in this review. Continuous adjustment under natural conditions allows for adaptive responses. In contrast, sudden light stress, as employed when analyzing stress responses in lab experiments, frequently results in cell destruction. Knowledge of all the flexible regulatory mechanisms, their responsiveness, and their interdependencies is needed when plant growth is to be engineered to optimize biomass and production of any desired molecules.

     

Fe homeostasis in plant cells: Does nicotianamine play multiple roles in the regulation of cytoplasmic Fe concentration?

The cellular and intracellular localization of the non-proteogenic amino acid nicotianamine (NA) in leaves and root elongation zones was immunochemically investigated in pea (Pisum sativum L.) and tomato (Lycopersicon esculentum Mill.) plants grown under various iron regimes and in three mutants defective in the regulation of iron uptake. Strongest immunostaining was observed in the over-accumulating pea mutants brz and dgl, and in iron-loaded wild-type plants. Fe concentration and NA level paralleled staining intensity, indicating that NA synthesis is induced by high iron availability. While label was mainly present in the cytoplasm under normal (10 microM) Fe supply and under Fe deprivation, most of the labeling was present in the vacuole in iron-loaded plants. This pattern resembled the distribution of NA in Fe over-accumulating mutants, indicating the possible importance of vacuolar sequestration in the detoxification of excess Fe. Based on the dependence of the cellular distribution of NA on the iron nutritional status of the plant, a possible role of NA in buffering free Fe in root and leaf cells was inferred. We show here for the first time that the NA concentration is increased in response to iron overload, indicating that, besides other classes of intracellular metal-binding ligands, NA may play an essential role in iron tolerance.     

Process parameter shifting: Part I. Effect of DOT, pH, and temperature on the performance of Epo-Fc expressing CHO cells cultivated in controlled batch bioreactors

The impact of process environment changes on process performance is one of the most crucial process safety issues when cultivating mammalian cells in a bioreactor. In contrast, directed shifting of process parameters can also be used as an optimization tool providing higher cell and product yields. Compared to other strategies that also aim on the regulation of cell growth and protein expression process parameter shifts can be easily performed without reagent addition or even genetic modification of the host cell line. However, a successful application of changing process conditions implies a profound understanding of the provoked physiological changes within the cells. In a systematic approach we varied the dissolved oxygen tension (DOT), pH, and temperature of CHO cultures in controlled bioreactors and investigated the impact on growth, productivity, metabolism, product quality and cell cycle distribution using a recombinant CHO cell line expressing the highly glycosylated fusion protein Epo-Fc. We found the reduction of cultivation temperature and the reduction of (external) pH to exert the most significant effects on process performance by mainly reducing cell growth and metabolism. With respect to the cell line used we identified a set of parameters capable of affecting cell proliferation in favor of an increased specific productivity and total product yield. The well directed alteration of the process environment has emerged as a tool adequate for further process optimization applying a biphasic cultivation strategy.     

Der Bau der Canini bei den Paulchoffatiidae (Multituberculata; Ober-Jura)

Canines are preserved among Multituberculata only in the upper jaw of the Paulchoffatiidae, the Pinheirodontidae and the North American genusGliodon Engelmann &; Callison, 1999. They resemble the anterior premolars (p^1–3) in the morphology of their crown, but they differ from them by the presence of only one root. In the present paper, 126 isolated canines of the Paulchoffatiidae from the Guimarota coal-pit in Portugal are treated. They show a wide morphological variation, from bicuspid to pentacuspid expression, and they can be grouped into 18 morphological units (Tab. 1). The bicuspid canines (11 specimens) show one buccal and one lingual cusp, arranged side by side, the latter normally being larger than the former. In the tricuspid canines (32 specimens) the cusps (one buccal and two lingual) are arranged in a triangle, with a great variation in the position of the cusps to each other. Tetracuspid canines (69 specimens) are the dominating group. Two buccal and two lingual cusps are present, which differ markedly in their largeness and their position to each other. The teeth differ also much in the shape of their crown. The pentacuspid canines (14 specimens) show two buccal and three lingual cusps. The variation in the arrangement of the cusps and in the shape of the crown is similar as in the tetracuspid specimens. One or two small cuspules can be present at the anterior border of the crown additionally to the main cusps in the canines. Tricuspid and tetracuspid canines are present also in the skulls of the Paulchoffatiidae, whereas bicuspid and pentacuspid canines are known only as isolated specimens. In the Pinheirodontidae the canines are tricuspid or tetracuspid, inGlirodon they are unicuspid. The Paulchoffatiidae and Pinheirodontidae — Paulchoffatiid line sensuKielan-Jaworowska &; Hurum, 2001 — are characterized by increasing premolarization of the upper canines. With that they differ markedly from all other multituberculates where the canini become reduced.     

First isolation of hepatitis E virus genotype 4 in Europe through swine surveillance in the Netherlands and Belgium

Hepatitis E virus (HEV) genotypes 3 and 4 are a cause of human hepatitis and swine are considered the main reservoir. To study the HEV prevalence and characterize circulating HEV strains, fecal samples from swine in the Netherlands and Belgium were tested by RT-PCR. HEV prevalence in swine was 7-15%. The Dutch strains were characterized as genotype 3, subgroups 3a, 3c and 3f, closely related to sequences found in humans and swine earlier. The HEV strains found in Belgium belonged to genotypes 3f and 4b. The HEV genotype 4 strain was the first ever reported in swine in Europe and an experimental infection in pigs was performed to isolate the virus. The genotype 4 strain readily infected piglets and caused fever and virus shedding. Since HEV4 infections have been reported to run a more severe clinical course in humans this observation may have public health implications.