EPO-independent functional EPO receptor in breast cancer enhances estrogen receptor activity and promotes cell proliferation

The main function of Erythropoietin (EPO) and its receptor (EPOR) is the stimulation of erythropoiesis. Recombinant human EPO (rhEPO) is therefore used to treat anemia in cancer patients. However, clinical trials have indicated that rhEPO treatment might promote tumor progression and has a negative effect on patient survival. In addition, EPOR expression has been detected in several cancer forms. Using a newly produced anti-EPOR antibody that reliably detects the full-length isoform of the EPOR we show that breast cancer tissue and cells express the EPOR protein. rhEPO stimulation of cultured EPOR expressing breast cancer cells did not result in increased proliferation, overt activation of EPOR (receptor phosphorylation) or a consistent activation of canonical EPOR signaling pathway mediators such as JAK2, STAT3, STAT5, or AKT. However, EPOR knockdown experiments suggested functional EPO receptors in estrogen receptor positive (ERα⁺) breast cancer cells, as reduced EPOR expression resulted in decreased proliferation. This effect on proliferation was not seen in ERα negative cells. EPOR knockdown decreased ERα activity further supports a mechanism by which EPOR affects proliferation via ERα-mediated mechanisms. We show that EPOR protein is expressed in breast cancer cells, where it appears to promote proliferation by an EPO-independent mechanism in ERα expressing breast cancer cells.     

Cytokine production using membrane adsorbers: Human basic fibroblast growth factor produced by Escherichia coli

Basic fibroblast growth factor (FGF‐2) is a multifunctional cytokine that regulates various cellular processes both in vitro and in vivo. FGF‐2 is extensively used in embryonic stem cell cultures since it can maintain the cells in an undifferentiated state. However, the high price of FGF‐2 has limited its application in stem cell research. Here we present a fast and efficient process for the purification of FGF‐2 from recombinant Escherichia coli cultures using reusable membrane adsorbers. A high expression level of FGF‐2 (42 mg/g dry cell) was achieved by fed‐batch cultivation of E. coli BL21(DE3). A new combination of cation exchange membrane chromatography and heparin‐sepharose affinity chromatography was used for the purification of the protein. A novel anion exchange membrane chromatography was used in the polishing step to remove endotoxins and DNA. In this new process, about 200 mg soluble FGF‐2 was yielded from 1.9 L culture broth with a purity of 98%. The purified protein was identified to be endotoxin‐free and bioactive. It was successfully tested to keep primate embryonic stem cell and human‐induced pluripotent stem cell pluripotent. Our approach, in which a controlled cultivation process is combined with an optimized fast and versatile downstreaming process, is suitable for low‐cost preparation of bioactive FGF‐2 at bench‐scale and may be beneficial to the effective production of other cytokines.     

Amino acid transport in brush-border-membrane vesicles isolated from human small intestine.

Uptake of L-alanine and L-phenylalanine by purified bursh-border-membrane vesicles isolated from human small intestine was investigated by using a rapid-filtration technique. L-Alanine entered the same osmotically reactive space as D-glucose, indicating that transport into the vesicle rather than binding to the membranes was being observed. The uptake rate for L-alanine was higher in the presence of a Na+ gradient than in the presence of a K+ gradient. In the presence of a Na+ gradient, the lipophilic anion SCN- caused an increase in L-alanine transport, whereas the nearly impermeant SO42- anion decreased the uptake of L-alanine compared with its uptake in the presence of Cl-. The uptake of L-phenylalanine into the brush-border-membrane vesicle was also stimulated by Na+. The results indicate co-transport of Na+ and neutral amino acids inthe human intestinal brush-border membrane.     

Metabolic syndrome in children and adolescents--risk for sleep-disordered breathing and obstructive sleep-apnoea syndrome?

The clinical relevance of the term "metabolic syndrome", the definition criteria, and predictive power are being disputed. Inclusion of sleep-disordered breathing and sleep apnoea into a definition of metabolic syndrome is also controversial once children and/or adolescents are affected. Nevertheless, along with the increasing prevalence of childhood obesity, the prevalence of the metabolic syndrome in obese children is reported at 30%, irrespective of the definition applied. Moreover, childhood obesity is associated with sleep-disordered breathing. Adipocytokines, cytokines secreted from adipose tissue, are thought to play a major role in the pathophysiology of metabolic syndrome. Leptin was initially suggested as a promising "anti-obesity" hormone. New concepts indicate that in humans leptin and its soluble receptor may be more important in states of energy deficiency rather than a predictor of the metabolic syndrome. Adiponectin, on the other hand, is not only related to obesity and insulin resistance, but appears to be the strongest predictor for metabolic syndrome, even in children. In newborns and infants, both adipocytokines occur in high concentrations, even though this cannot completely explain the increased risk for ensuing metabolic disease later in life. Finally, low-grade systemic inflammation may underlie the clustering of metabolic risk factors. Overall factors from the adipose tissue may constitute not only markers but also mediators of metabolic sequelae of obesity.     

Neuropathological changes in scrapie and Alzheimer's disease are associated with increased expression of apolipoprotein E and cathepsin D in astrocytes.

With the rationale that the neuropathological similarities between scrapie and Alzheimer's disease reflect convergent pathological mechanisms involving altered gene expression, we set out to identify molecular events involved in both processes, using scrapie as a model to study the time course of these changes. We differentially screened a cDNA library constructed from scrapie-infected mice to identify mRNAs that increase or decrease during disease and discovered in this way two mRNAs that are increased in scrapie and Alzheimer's disease. These mRNAs were subsequently shown by sequence analysis to encode apolipoprotein E and cathepsin D (EC 3.4.23.5). Using in situ hybridization and immunocytochemistry to define the cellular and anatomic pathology of altered gene expression, we found that in both diseases the increase in apolipoprotein E and cathepsin D mRNAs and proteins occurred in activated astrocytes. In scrapie, the increase in gene expression occurred soon after the amyloid-forming abnormal isoform of the prion protein has been shown to accumulate in astrocytes. In Alzheimer's disease, the increased expression of cathepsin D also occurred in association with beta-amyloid. These studies reveal some of the molecular antecedents of neuropathological changes in scrapie and Alzheimer's disease and accord new prominence to the role of astrocytes in neurodegenerative conditions.     

Genetic differentiation among populations of Cicerbita alpina (L.) Wallroth (asteraceae) in the western Alps

In this study we analyzed the genetic population structure of the hygrophilous tall-herb Cicerbita alpina in the western Alps because this group of mountain plants is underrepresented in the biogeographical literature. AFLP (amplified fragment length polymorphism) fingerprints of 40 samples were analyzed from four populations situated in a transect from the southwestern Alps to the eastern part of the western Alps and one population from the Black Forest outside the Alps. Two genetic groups can be distinguished. The first group (A) comprises the populations from the northern and eastern parts of the western Alps, and the second group (B) comprises the populations from the southwestern Alps and the Black Forest. Group A originates most likely from at least one refugium in the southern piedmont regions of the Alps. This result provides molecular evidence for a humid climate at the southern margin of the Alps during the Würm glaciation. Group B originates presumably from western or northern direction and we discuss two possible scenarios for the colonization of the Alps, i.e. (1) long-distance dispersal from southwestern refugia and (2) colonization from nearby refugia in the western and/or northern Alpine forelands. The study demonstrates that the target species harbours considerable genetic diversity, even on a regional scale, and therefore is a suitable model for phylogeographic research.     

Serological definition of the lentivirus group of retroviruses.

The major polypeptides of visna viruses and other lentiviruses have been isolated and shown to be closely related if not identical in radioimmunoassays. By this criterion the lentiviruses form a distinct group of retroviruses unrelated to spuma viruses, mammalian and avian retroviruses that cause tumors, and unclassified retroviruses of cattle and horses. Two sera obtained from goats immunized with Mason-Pfizer monkey virus or squirrel monkey virus reacted with visna p30. Additional data suggest that this reaction represents infection of goats with a lentivirus or a new retrovirus closely related to the lentiviruses.     

Distant land use affects terrestrial and aquatic habitats of high naturalness

Biotas from all ecosystems need to respond to factors that determine habitat suitability. These factors originate from different scales. Effects can be assumed to be hierarchical in the order large-scale geographic > regional > local > small-scale in-habitat factors. We aimed at the identification of general patterns by comparisons between ecosystems (forest floor snails, hololimnic stream macroinvertebrates) and across scales, and include potential seasonal effects. Sampling sites displayed signs of naturalness, such as high levels of deadwood accumulation in the forests, or a lack of artificial stream bed fixation plus a “good” to “high” score for the assemblage-derived Multimetric Index (MMI) in the streams. Terrestrial and aquatic assemblages of non-emergent taxa fluctuated independent of seasonal effects. They differed in their relative correlation with environmental matrices with quasi-concentric effects in forests, and longitudinal effects in streams. Large-scale factors, namely geographic position, strongly influenced assemblage turnover, but the effect is based on a high covariation between geographic position and environmental factors. We thus extracted variables that best explained species turnover after correcting for spatio-temporal effects. The terrestrial community assembling was habitat-based and mainly responded to soil acidification, distance to disturbances, and regional scale deforestation and deciduous/mixed forest cover. The stream assemblages were structured by regional pasture cover, organic pollution, regional deciduous forest cover and microlithal cover. Apparently, community assembly occurs along with changes in regional forest cover and the transport of nutrients and matter that can originate from a distance, irrespective of ecosystem and assumed “naturalness”.