Correlation between frequencies of blood monocytic myeloid-derived suppressor cells,regulatory T cells and negative prognostic markers in patients with castration-resistant metastatic prostate cancer

Myeloid-derived suppressor cells (MDSC) are believed to play a role in immune suppression and subsequent failure of T cells to mount an efficient anti-tumor response, by employing both direct T-cell inhibition as well as induction of regulatory T cells (Tregs). Investigating the frequency and function of immune suppressive cell subsets in the peripheral blood of 41 patients with prostate cancer (PC) and 36 healthy donors (HD) showed a significant increase in circulating CD14⁺ HLA-DR^low/neg monocytic MDSC (M-MDSC) and Tregs in patients with PC compared to HD. Furthermore, M-MDSC frequencies correlated positively with Treg levels. In vitro proliferation assay with autologous T cells confirmed M-MDSC-mediated T-cell suppression, and intracellular staining of immune suppressive enzyme iNOS revealed a higher expression in M-MDSC from patients with PC. Increased frequencies of M-MDSC correlated with known negative prognostic markers in patients with PC including elevated levels of lactate dehydrogenase and prostate-specific antigen. Accordingly, high levels of M-MDSC were associated with a shorter median overall survival. Our data strongly suggest that M-MDSC, possibly along with Tregs, play a role in establishing an immune suppressive environment in patients with PC. Moreover, correlation of M-MDSC frequency with known prognostic markers and the observed impact on OS could reflect a possible role in tumor progression. Further insight into the generation and function of MDSC and their interplay with Tregs and other cell types may suggest ways to tackle their induction and/or function to improve immunological tumor control.     

Prevalence of tick-borne encephalitis virus antibodies in dogs from Denmark

Background  

Large regions of central and eastern Europe are recognized as areas where tick-borne encephalitis virus (TBEV) is endemic, including countries neighbouring Denmark. It is therefore timely and relevant to determine if TBEV infections occur in Denmark. This study investigates the presence of antibodies against TBEV in a cross-section of the Danish canine population to assess the level of exposure to TBEV and possibly identify TBEV microfoci in Denmark.     

Proteomic profiling of the Baltic Sea cyanobacterium Nodularia spumigena strain AV1 during ammonium supplementation

The cyanobacterium Nodularia spumigena dominates the annual, toxic summer blooms in the Baltic Sea. Although Nodularia has been receiving attention due to its production of the hepatotoxin nodularin, molecular data regarding the regulation of nitrogen fixation is lacking. We have previously reported that N. spumigena strain AV1, unlike model filamentous cyanobacteria, differentiates heterocysts in the absence of detectable nitrogen fixation activity. To further analyze the uncoupling between these two linked processes, we assessed the impact of ammonium ions on the N. spumigena metabolism using a proteomic approach. Proteomic profiling was performed at three different times during ammonium supplementation using quantitative 2-dimensional gel electrophoresis followed by MS/MS analysis. Using this approach, we identified 34 proteins, 28 of which were unique proteins that changed successively in abundance during growth on ammonium. Our results indicate that N. spumigena generally exhibits lower energy production and carbon fixation in the presence of ammonium and seems to be inefficient in utilizing ammonium as an external nitrogen source. The possibility of ammonium toxicity due to PSII damage was investigated and the results are discussed. Our findings have implications in regard to the strategies considered to manage the cyanobacterial blooms in the Baltic Sea.     

Percutaneous cryoablation of colorectal liver metastases: potentiated by two consecutive freeze-thaw cycles

Cryoablation may be beneficial for selected patients with liver tumours. Two freeze-thaw cycles at the same location have been recommended during treatment as this potentiate the effect of ablation in experimental studies. However, single freeze ablations are used by some as double freeze procedures are time-consuming and have been associated with increased risk of complications. Estimation of ice-ball volume is difficult using regularly used monitoring techniques. Magnetic resonance imaging, however, allows excellent and multiplanar visualisation of the frozen region during ablation. We comment on the effect of double freeze cycles in regard to ice-ball volume as estimated from magnetic resonance imaging during percutaneous cryoablation of colorectal liver metastases. The ice-ball volume at the end of the second freeze cycle was median 42% larger than the volume at the end of the first freeze. Double freeze cycles may thus facilitate tumour destruction.     

FCS cell surface measurements--photophysical limitations and consequences on molecular ensembles with heterogenic mobilities.

BACKGROUND: Fluorescence Correlation Spectroscopy is a powerful method to analyze densities and diffusive behavior of molecules in membranes, but effects of photodegradation can easily be overlooked. METHOD: Based on experimental photophysical parameters, calculations were performed to analyze the consequences of photobleaching in fluorescence correlation spectroscopy (FCS) cell surface experiments, covering a range of standard measurement conditions. RESULTS: Cumulative effects of photobleaching can be prominent, although an absolute majority of the fluorescent molecules would pass the laser excitation beam without being photo-bleached. Given a distribution of molecules on a cell surface with different diffusive properties, the fraction of molecules that is actually analyzed depends strongly on the excitation intensities and measurement times, as well as on the size of the reservoir of freely diffusing molecules. Both the slower and the faster diffusing molecules can be disfavored. CONCLUSIONS: Apart from quantifying photobleaching effects, the calculations suggest that the effects can be used to extract additional information, for instance about the size of the reservoirs of free diffusion. By certain choices of measurement conditions, it may be possible to more specifically analyze certain species within a population, based on their different diffusive properties, different areas of free diffusion, or different kinetics of possible transient binding.     

Disruption of the Cytokeratin Cytoskeleton and Inhibition of Hepatocytic Autophagy by Okadaic Acid

To learn whether autophagy might be dependent on any of the major cytoskeletal elements, the effect of various cytoskeleton inhibitors on autophagy and cytoskeletal organization was studied in isolated rat hepatocytes. Autophagy, measured as the sequestration of endogenous lactate dehydrogenase, was completely inhibited in isolated rat hepatocytes by the protein phosphatase inhibitor okadaic acid (30 nM). Only small effects were seen with vinblastine (10 μM) or cytochalasin D (10 μM). Indirect immunofluorescence microscopy with antibody to a 55-kDa cytokeratin, corresponding to human cytokeratin 8 (CK8), revealed that whereas control cells contained a well-organized network of cytokeratin intermediate filaments, okadaic acid disrupted this network into small spherical aggregates. Treatment with cytochalasin D or vinblastine, which disrupt microfilaments and microtubules, respectively, had no detectable effect on the cytokeratin filament distribution. Neither the microtubule network (detected by indirect immunofluorescence with antibodies against α- and β-tubulin) nor the actin microfilament network (detected by rhodamine-palloidin) was disrupted by okadaic acid. Naringin (100 μM), a putative protein kinase-inhibitory flavonoid, offered complete protection against the autophagy-inhibitory and cytokeratin-disruptive effects of okadaic acid. Two other flavonoids, genistein (100 μM) and prunin (100 μM) as well as KN-62 (10 μM), a specific inhibitor of Ca²⁺/calmodulin-dependent kinase II), likewise displayed a good ability to protect against the effect of okadaic acid upon cytokeratin organization, while no such protection was seen with H-89 (20 μM), an inhibitor of the cyclic nucleotide-dependent protein kinases, or with H-7 (100 μM), which in addition inhibits protein kinase C. The results suggest that the cytokeratin cytoskeleton of hepatocytes is subject to rapid control by phosphorylation and dephosphorylation and that cytokeratin filaments may somehow be involved in the autophagic process.     

Linker insertion analysis of the FimH adhesin of type 1 fimbriae in an Escherichia coli fimH-null background

Abstract The gene encoding the Escherichia coli FimH adhesin of type 1 fimbriae has been subjected to linker insertion mutagenesis. Amino acid changes were introduced at a number of positions spanning the entire sequence in order to probe the structure-function relationship of the FimH protein. The effect of these mutations on the ability of bacteria to express a D-mannose binding phenotype was assessed in a fimH null mutant (MS4) constructed by allelic exchange in the E. coli K-12 strain PC31. Mutations mapping at amino acid residues 36, 58 and 279 of the mature FimH protein were shown to completely abolish binding to D-mannose receptors. Differences in the level of fimbriation were also observed as a result of some of the mutations in the fimH gene. These mutants may prove useful in dissecting receptor-ligand interactions by defining regions of the FimH protein that are important in erythrocyte binding.     

Molecular Phylogeny of Some European Heteronemertean (Nemertea) Species and the Monophyletic Status ofRiseriellus, Lineus,andMicrura

The 16S rRNA mitochondrial gene was used to reconstruct the relationships among 10 heteronemertean species (subclass Heteronemertea, phylum Nemertea);Lineus ruberandL. viridisare represented by more than one specimen to assess intraspecific variation in these enigmatic species, and the analysis includes in total 14 terminal taxa incorporating one palaeonemertean species (Tubulanus annulatus) for outgroup rooting. The aligned sequences were subjected to maximum parsimony, maximum-likelihood, and neighbor-joining analyses to estimate the phylogenetic relationship of the species. The results were concordant from all analyses and indicate that neitherLineusnorMicruraare monophyletic taxa, and that there is no support from a phylogenetic point of view to establish the monotypic genusRiseriellus.     

Effect of grazing by a neotropical copepod, Notodiaptomus, on a natural cyanobacterial assemblage and on toxic and non-toxic cyanobacterial strains

The neotropical, freshwater copepod Notodiaptomus iheringi decreasedthe growth of small colonies in cyanobacteria-dominated plankton,indicating that they can be efficiently ingested and that theymay represent an important food source for this copepod in reservoirsdominated by cyanobacteria. Cultured, colonial, toxic Microcystisaeruginosa was not affected, indicating that toxicity may causea decrease in its ingestion by N. iheringi.