Vaccenic acid-mediated reduction in cytokine production is independent of c9,t11-CLA in human peripheral blood mononuclear cells

The ruminant trans fatty acid vaccenic acid (tVA) favorably alters markers of inflammation. However, it is not yet clear whether these effects are attributed to its endogenous partial conversion to c9,t11-CLA, which is known to possess anti-inflammatory properties. We compared the cytokine reducing potential of tVA to c9,t11-CLA in human T-helper (Th) cells as a main source of cytokine production during inflammation. Secondly, we assessed whether a bioconversion of tVA to c9,t11-CLA via stearoyl-CoA desaturase (SCD) encoded activity takes place in peripheral blood mononuclear cells (PBMC) in order to relate the outcomes of intracellular cytokine measurement to the degree of conversion. TVA reduced the percentage of both IL-2 and TNF-α expressing Th cells significantly, but to a lesser extent compared to c9,t11-CLA, as determined by flow cytometry after alloreactive stimulation of PBMC. Pre-treatment with the selective PPARγ antagonist T0070907 largely re-established the IL-2 and TNF-α positive Th cell population in both tVA and c9,t11-CLA treated cultures. Interestingly, while the portion of tVA dose-dependently increased within the cellular lipid fraction, the initially marginal amount of c9,t11-CLA remained unaltered. However, SCD mRNA although abundantly expressed in PBMC was not regulated by tVA. Conclusively, these results suggest that the cytokine reducing effect of tVA in human T cells is independent of c9,t11-CLA, since no bioconversion occurred. Moreover, the data provide evidence that tVA mechanistically acts in a manner similar to c9,t11-CLA.     

Immunofluorescent Identification of Type 12 Group A Streptococci

The fluorescent antibody (FA) conjugate prepared by labeling streptococcal M type 12 antibody with fluorescein isothiocyanate was found to exhibit considerable nonspecific FA staining with other group A M-serotypes. The cross-reactions could be reduced sufficiently or eliminated by the addition of adsorbed homologous blocking serum (AHB) but not by preimmune serum. The AHB was prepared by adsorbing type 12 antiserum with untreated homologous cells. Comparative staining with unblocked and AHB-blocked FA conjugates enabled type 12 streptococci from clinical specimens to be rapidly and accurately identified.     

The phosphoproteome of a Chlamydomonas reinhardtii eyespot fraction includes key proteins of the light signaling pathway

Flagellate green algae have developed a visual system, the eyespot apparatus, which allows the cell to phototax. In a recent proteomic approach, we identified 202 proteins from a fraction enriched in eyespot apparatuses of Chlamydomonas reinhardtii. Among these proteins, five protein kinases and two protein phosphatases were present, indicating that reversible protein phosphorylation occurs in the eyespot. About 20 major phosphoprotein bands were detected in immunoblots of eyespot proteins with an anti-phosphothreonine antibody. Toward the profiling of the targets of protein kinases in the eyespot fraction, we analyzed its phosphoproteome. The solubilized proteins of the eyespot fraction were treated with the endopeptidases LysC and trypsin prior to enrichment of phosphopeptides with immobilized metal-ion affinity chromatography. Phosphopeptides were analyzed by nano-liquid chromatography-electrospray ionization-mass spectrometry (MS) with MS/MS as well as neutral-loss-triggered MS/MS/MS spectra. We were able to identify 68 different phosphopeptides along with 52 precise in vivo phosphorylation sites corresponding to 32 known proteins of the eyespot fraction. Among the identified phosphoproteins are enzymes of carotenoid and fatty acid metabolism, putative signaling components, such as a SOUL heme-binding protein, a Ca(2+)-binding protein, and an unusual protein kinase, but also several proteins with unknown function. Notably, two unique photoreceptors, channelrhodopsin-1 and channelrhodopsin-2, contain three and one phosphorylation sites, respectively. Phosphorylation of both photoreceptors occurs in the cytoplasmatic loop next to their seven transmembrane regions in a similar distance to that observed in vertebrate rhodopsins, implying functional importance for regulation of these directly light-gated ion channels relevant for the photoresponses of C. reinhardtii.     

Dynamics of the actin-binding protein drebrin in motile cells and definition of a juxtanuclear drebrin-enriched zone

The actin-binding protein (ABP) drebrin, isoform E2, is involved in remodelling of the actin cytoskeleton and in formation of cell processes, but its role in cell migration has not yet been investigated. Therefore, we have studied the organization of drebrin in motile cultured cells such as murine B16F1 melanoma and human SV80 fibroblast cells, using live cell confocal microscopy. In cells overexpressing DNA constructs encoding drebrin linked to EGFP, numerous long, branched cell processes were formed which slowly retracted and extended, whereas forward movement was halted. In contrast, stably transfected B16F1 cells containing drebrin-EGFP at physiological levels displayed lamellipodia and were able to migrate on laminin. Surprisingly, in such cells, drebrin was absent from anterior lamellipodia but was enriched in a specific juxtanuclear zone, the "drebrin-enriched zone" (DZ), and in the tail. In leading edges of SV80 cells, characterized by pronounced actin microspikes, drebrin was specifically enriched along posterior portions of the microspikes, together with tropomyosin. Drebrin knock-down by small interfering RNAs did not impair movements of SV80 cells. Our results confirm the role of drebrin E2 in the formation of branching processes and further indicate that during cell migration, the protein contributes to retraction of the cell body and the tail but not to lamellipodia formation. In particular, the novel, sizable juxtanuclear DZ structure will have to be characterized in future experiments with respect to its molecular assembly and cell biological functions.     

The non-impact of hunting on moose <Emphasis Type="Italic">Alces alces</Emphasis> movement,diurnal activity,and activity range

Previous studies on moose Alces alces have suggested that interactions with humans may trigger anti-predator behaviors and generate a demographical cost. Therefore, we hypothesized that disturbances from small and big game hunting may have negative effects on moose movements, diurnal activity, and activity range. Using location data from 64 moose equipped with GPS collars from three populations (Low Alpine, Inland, Coastal) with different temporal human presence and spatial accessibility, we evaluated the impact of hunting on moose activity rhythms. On average, female moose in the low human population density (Low Alpine) area (<0.5/km²) had significantly lower movement rates during moose hunting season, but variation in movement rates among individuals were higher compared with female moose in regions with denser human populations (6–24/km²). We found no evidence that reproductive status influenced female moose sensitivity to disturbance. As expected, females used smaller activity ranges and were less active nocturnally than males. The high within-group variation suggests that current hunting disturbance levels do not alter moose population behavior in general. Our data indicate that alterations in movement were related to rutting activity, not human disturbance induced by hunting. In line with behavioral theory, our study suggests that some individuals were more sensitive to hunting disturbance than the general population. Our work suggests that individual moose may perceive human predation risk to be similar to other predation risks.     

Do Female Zebra Finches,Taeniopygia guttata,Choose Their Mates Based on Their ‘Personality’?

A major challenge in behavioural and evolutionary ecology is to understand the evolution and maintenance of consistent behavioural differences among individuals within populations, often referred to as animal ‘personalities’. Here, we present evidence suggesting that sexual selection may act on such personality differences in zebra finches (Taeniopygia guttata), as females seem to choose males on the basis of their exploratory behaviour per se, while taking into account their own personality. After observing a pair of males, whose apparent levels of exploration were experimentally manipulated, females that exhibited low‐exploratory tendencies showed no preference during mate choice for males that had appeared to be either ‘exploratory’ or ‘unexploratory’. In contrast, intermediate and highly exploratory females preferred apparently exploratory males over apparently unexploratory ones. Our results suggest that behavioural or genetic compatibility for personality traits might be important for mate choice, at least for exploratory individuals.     

Impairment of LTD and cerebellar learning by Purkinje cell-specific ablation of cGMP-dependent protein kinase I

The molecular basis for cerebellar plasticity and motor learning remains controversial. Cerebellar Purkinje cells (PCs) contain a high concentration of cGMP-dependent protein kinase type I (cGKI). To investigate the function of cGKI in long-term depression (LTD) and cerebellar learning, we have generated conditional knockout mice lacking cGKI selectively in PCs. These cGKI mutants had a normal cerebellar morphology and intact synaptic calcium signaling, but strongly reduced LTD. Interestingly, no defects in general behavior and motor performance could be detected in the LTD-deficient mice, but the mutants exhibited an impaired adaptation of the vestibulo-ocular reflex (VOR). These results indicate that cGKI in PCs is dispensable for general motor coordination, but that it is required for cerebellar LTD and specific forms of motor learning, namely the adaptation of the VOR.     

Erythropoietin improves the survival of fat tissue after its transplantation in nude mice

Background

Autologous transplanted fat has a high resorption rate, providing a clinical challenge for the means to reduce it. Erythropoietin (EPO) has non-hematopoietic targets, and we hypothesized that EPO may improve long-term fat graft survival because it has both pro-angiogenic and anti-apoptotic properties. We aimed to determine the effect of EPO on the survival of human fat tissue after its transplantation in nude mice.

Methodology/Principal Findings

Human fat tissue was injected subcutaneously into immunologically-compromised nude mice, and the grafts were then treated with either 20 IU or 100 IU EPO. At the end of the 15-week study period, the extent of angiogenesis, apoptosis, and histology were assessed in the fat grafts. The results were compared to vascular endothelial growth factor (VEGF)-treated and phosphate-buffered saline (PBS)-treated fat grafts. The weight and volume of the EPO-treated grafts were higher than those of the PBS-treated grafts, whose weights and volumes were not different from those of the VEGF-treated grafts. EPO treatment also increased the expression of angiogenic factors and microvascular density, and reduced inflammation and apoptosis in a dose-dependent manner in the fat grafts.

Conclusions/Significance

Our data suggest that stimulation of angiogenesis by a cluster of angiogenic factors and decreased fat cell apoptosis account for potential mechanisms that underlie the improved long-term survival of fat transplants following EPO treatment.