Role of glutamine synthetase in angiogenesis beyond glutamine synthesis

Glutamine synthetase, encoded by the gene GLUL, is an enzyme that converts glutamate and ammonia to glutamine. It is expressed by endothelial cells, but surprisingly shows negligible glutamine-synthesizing activity in these cells at physiological glutamine levels. Here we show in mice that genetic deletion of Glul in endothelial cells impairs vessel sprouting during vascular development, whereas pharmacological blockade of glutamine synthetase suppresses angiogenesis in ocular and inflammatory skin disease while only minimally affecting healthy adult quiescent endothelial cells. This relies on the inhibition of endothelial cell migration but not proliferation. Mechanistically we show that in human umbilical vein endothelial cells GLUL knockdown reduces membrane localization and activation of the GTPase RHOJ while activating other Rho GTPases and Rho kinase, thereby inducing actin stress fibres and impeding endothelial cell motility. Inhibition of Rho kinase rescues the defect in endothelial cell migration that is induced by GLUL knockdown. Notably, glutamine synthetase palmitoylates itself and interacts with RHOJ to sustain RHOJ palmitoylation, membrane localization and activation. These findings reveal that, in addition to the known formation of glutamine, the enzyme glutamine synthetase shows unknown activity in endothelial cell migration during pathological angiogenesis through RHOJ palmitoylation

Tolerance of benthic diatoms from temperate aquatic and terrestrial habitats to experimental desiccation and temperature stress

Soils differ from aquatic sediments in environmental characteristics such as moisture availability and temperature fluctuations, and it is therefore believed that soil-inhabiting diatoms have a broader tolerance range to these stresses than aquatic diatoms. To test this hypothesis, we assessed the survival capacities of vegetative cells of 34 benthic diatom species from terrestrial and aquatic habitats in Belgium when exposed to desiccation and temperature stress. Six different stress conditions were studied: gradual heating up to +30°C and +40°C, abrupt heating to +40°C, freezing. to -20°C and desiccation with and without preconditioning at +30°C. All six conditions resulted in a significantly decreased survival of cells compared to control conditions. Desiccation killed all tested strains, freezing was survived by only three species and abrupt heating was significantly more lethal than gradual heating, suggesting a generally high sensitivity of vegetative diatom cells to these three stress factors. While tolerance to temperature extremes (+40°C and -20°C) was to a large extent species-specific, habitat-specific differences in cell survival were also detected. Only terrestrial species survived freezing, and aquatic diatoms were less tolerant to gradual heating to +40°C, both pointing at a higher tolerance of terrestrial diatoms to temperature extremes. Moreover, in two species with both aquatic and terrestrial isolates, only the terrestrial strains survived +40°C. We conclude that vegetative cells of benthic diatoms (1) are very sensitive to desiccation, freezing and abrupt heating and (2) have a habitat-dependent tolerance to temperature extremes. The consequences of these observations for the dispersal capacities and the subsequent biogeographical patterns of diatoms are discussed

In vitro selection and characterization of putative probiotics isolated from the gut of Acipenser baerii (Brandt, 1869)

To select and characterize potential probiotic bacteria from the gut microbiota of Siberian sturgeon (Acipenser baerii), 129 strains isolated from the hindgut were screened for antagonistic activity against five fish pathogens. Ten isolates showed antagonism towards three or more pathogens. Nine of these isolates were Gram-positive, belonging to Lactococcus (seven) and Bacillus (two), and a single strain belonging to the Gram-negative Citrobacter. These inhibitory isolates were identified using genetic, phentotypic and biochemical traits, and further characterized by in vitro tests assessing the adhesion and growth in mucus and resistance to gastric and intestinal fluids. The candidate probiotics were determined to be non-pathogenic through an in vivo study. Based on these assays, Lactococcus lactis ssp. lactis STG45 and STG81 showed the broadest inhibitory potential, a high viability in simulated gastrointestinal juice and the highest adhesion capacity to mucus. They were therefore selected as the most promising candidate probiotics. This is the first study screening probiotics among the gut microflora of Siberian sturgeon

Effects of arabinoxylan-oligosaccharides (AXOS) on juvenile Siberian sturgeon (Acipenser baerii) performance, immune responses and gastrointestinal microbial community

Arabinoxylan-oligosaccharides (AXOS) are a newly discovered class of candidate prebiotics that exert different properties depending on their structure. In this study the effects of two different structures of AXOS, namely AXOS-32-0.30 (average degree of polymerization: 32, average degree of substitution: 0.30) and AXOS-3-0.25, were investigated on growth performance, immune responses, gut microbial fermentation and gut bacterial composition of juvenile Siberian sturgeon (Acipenser baerii). After a two weeks acclimation, fish (25.9 ± 0.9 g) were distributed over 24 aquariums (8 replicates per treatment) and fed a control diet or a diet containing 2% AXOS-32-0.30 or AXOS-3-0.25 for 12 weeks. Growth performance and feed utilization tend to improve in sturgeon fed on diets supplemented with AXOS-32-0.30, however not significant. Survival was high in all groups. Both AXOS preparations significantly enhanced the phagocytic activity of fish macrophages compared to the control group, while the alternative haemolytic complement activity and total serum peroxidase content improved only in the group fed AXOS-32-0.30 (P <0.05). The lysozyme activity was not affected by AXOS addition. Simultaneously, the amount of short-chain fatty acids (SCFAs) was highest in the hind gut of sturgeon fed AXOS-32-0.30. The concentrations of acetate, butyrate and total SCFAs in fish fed AXOS-32-0.30 was significantly higher than in the groups fed the control diet or AXOS-3-0.25. Study of the bacterial community in the sturgeon hindgut using PCR-denaturing gradient gel electrophoresis (PCR-DGGE) revealed that both preparations of AXOS induced changes in the bacterial composition. According to redundancy analysis (RDA), hindgut microbiota of each treatment group clustered apart from one another (P = 0.001). DNA sequencing of the dominant DGGE bands recovered from the different treatments showed that AXOS mainly stimulated the growth of lactic acid bacteria and Clostridium sp., with more pronounced effects of AXOS-32-0.30. It is concluded that AXOS improves sturgeon health through prebiotic action, but the induced effects depend on the specific structure of AXOS. A higher degree of polymerization of AXOS had a stronger beneficial impact in this sturgeon specie