The solution molecular weight and shape of the bacterial exopolysaccharides amylovoran and stewartan

Amylovoran, the acidic exopolysaccharide (EPS) of Erwinia amylovora, and stewartan, the capsular EPS of E. stewartii, were characterized by analytical ultracentrifugation and by size exclusion chromatography connected to dual detection of light scattering and mass. The average molecular weights of amylovoran and stewartan were determined as 1.0×10⁶ and 1.7×10⁶ Da, with polydispersity values (M_w/M_n) of 1.5 and 1.4, respectively. Based on the sugar composition and their molecular weight, both exopolysaccharides consist of approximately 1000 repeating units per molecule, this suggests a similar mechanism for chain length determination during biosynthesis of EPS in both organisms.     

Highly potent inhibitors of proprotein convertase furin as potential drugs for treatment of infectious diseases

Optimization of our previously described peptidomimetic furin inhibitors was performed and yielded several analogs with a significantly improved activity. The most potent compounds containing an N-terminal 4- or 3-(guanidinomethyl)phenylacetyl residue inhibit furin with K(i) values of 16 and 8 pM, respectively. These analogs inhibit other proprotein convertases, such as PC1/3, PC4, PACE4, and PC5/6, with similar potency, whereas PC2, PC7, and trypsin-like serine proteases are poorly affected. Incubation of selected compounds with Madin-Darby canine kidney cells over a period of 96 h revealed that they exhibit great stability, making them suitable candidates for further studies in cell culture. Two of the most potent derivatives were used to inhibit the hemagglutinin cleavage and viral propagation of a highly pathogenic avian H7N1 influenza virus strain. The treatment with inhibitor 24 (4-(guanidinomethyl)phenylacetyl-Arg-Val-Arg-4-amidinobenzylamide) resulted in significantly delayed virus propagation compared with an inhibitor-free control. The same analog was also effective in inhibiting Shiga toxin activation in HEp-2 cells. This antiviral effect, as well as the protective effect against a bacterial toxin, suggests that inhibitors of furin or furin-like proprotein convertases could represent promising lead structures for future drug development, in particular for the treatment of infectious diseases.     

Acute myocardial infarction complicating subarachnoid haemorrhage

An acute myocardial infarction is a rare complication of a subarachnoid haemorrhage. The combination of these two conditions imposes important treatment dilemmas. We describe two patients with this combination of life-threatening conditions. Patient 1 was treated with emergency percutaneous coronary intervention followed by clipping of the anterior communicating artery aneurysm. Six months after discharge the patient's memory and orientation had almost completely recovered. Patient 2 was treated with aspirin until coiling of the aneurysm could be performed. After successful coiling low-molecular-weight heparin was added. One week later the patient died due to a free wall rupture. (Neth Heart J 2009;17:284–7.)     

Autoradiographic studies of glial proliferation in different areas of the brain of the 14-day-old rat

Cell cycle parameters, as well as the mode of proliferation of glial cells, in four different areas of the brain of the 14-day-old rat (cortex, corpus callosum, nucleus caudatus putamen and commissura anterior) were studied using different cell kinetic methods after injection of [3H]TdR and/or [14C]TdR. The duration of the S phase (tS) was found to be about 10 hr and that of the cycle time (tC) about 20 hr, tG2 is less than 2 hr and t(G2 + M) about 4 hr. These values are valid for glial cells in all four brain areas studied. However, the labelling index (LI) of the glial cells differs by a factor of 3, between 1.8 and 5.4% in the different brain areas. Accordingly, the growth fraction of the glial cell population in the four areas varies between 0.04 and 0.12. Glial cells (astrocytes as well as oligodendrocytes) proliferate according to a steady state system. Furthermore, the proliferation of glial cells is associated with continuous cell loss. After each mitosis about 3% of the daughter cells become pyknotic and die. In addition, a permanent exchange of glial cells occurs between the proliferating and non-proliferating pool.     

Protein-free transfection of CHO host cells with an IgG-fusion protein: selection and characterization of stable high producers and comparison to conventionally transfected clones

In order to improve the current techniques of cell cultivation in the absence of serum, we have developed a protein-free transfection protocol for CHO cells, based on the Nucleofector technology. After starting with a heterogeneous pool of primary transfectants which express the fusion protein EpoFc, we isolated single clones and compared them with parallel clones generated by lipofection in serum-dependent cultivation. Our intensive characterization program was based on determination of specific productivity (q(p)) and analysis of genetic parameters. In two nucleofection experiments, transfection with 5 microg of DNA resulted in best productivities of the primary cell pools. After subcloning, the q(p) could be raised up to 27 pg x cells(-1) x day(-1). While the serum-dependent transfectants exhibited specific productivities up to 57 pg x cells(-1) x day(-1) in serum-dependent cultivation, a significant decrease that resulted in the range of q(p) of the protein-free transfectants was observed after switching to protein-free conditions. Investigation of genetic parameters revealed higher mRNA levels and gene copy numbers (GCN) for the protein-free adapted serum-dependent transfectants. Therefore, we assume that problems during protein-free adaptation (PFA) lead to a less efficient translation machinery after serum deprivation. We describe the generation of stable-producing recombinant CHO clones by protein-free transfection of a protein-free adapted host cell line, which reduces the risk of adverse clonal changes after PFA. The main advantage of this approach is the earlier predictability of clone behavior, which makes the generation of production clones by protein-free transfection, a viable and highly efficient strategy for recombinant cell line development.     

Identification of potential neuromotor mechanisms of manual therapy in patients with musculoskeletal disablement: rationale and description of a clinical trial

Background  

Many health care practitioners use a variety of hands-on treatments to improve symptoms and disablement in patients with musculoskeletal pathology.     

Effect of long-term organic and mineral fertilization strategies on rhizosphere microbiota assemblage and performance of lettuce

Long-term agricultural fertilization strategies gradually change soil properties including the associated microbial communities. Cultivated crops recruit beneficial microbes from the surrounding soil environment via root exudates. In this study, we aimed to investigate the effects of long-term fertilization strategies across field sites on the rhizosphere prokaryotic (Bacteria and Archaea) community composition and plant performance. We conducted growth chamber experiments with lettuce (Lactuca sativa L.) cultivated in soils from two long-term field experiments, each of which compared organic versus mineral fertilization strategies. 16S rRNA gene amplicon sequencing revealed the assemblage of a rhizosphere core microbiota shared in all lettuce plants across soils, going beyond differences in community composition depending on field site and fertilization strategies. The enhanced expression of several plant genes with roles in oxidative and biotic stress signalling pathways in lettuce grown in soils with organic indicates an induced physiological status in plants. Lettuce plants grown in soils with different fertilization histories were visibly free of stress symptoms and achieved comparable biomass. This suggests a positive aboveground plant response to belowground plant–microbe interactions in the rhizosphere. Besides effects of fertilization strategy and field site, our results demonstrate the crucial role of the plant in driving rhizosphere microbiota assemblage.