Amino acid metabolism in leg muscle after an endotoxin injection in healthy volunteers

Decreased plasma amino acid concentrations and increased net release of amino acids from skeletal muscle, especially for glutamine, are common features in critically ill patients. A low dose of endotoxin administered to healthy volunteers was used as a human model for the initial phase of sepsis to study the early metabolic response to sepsis. Six healthy male volunteers were studied in the postabsorptive state. Blood samples from the forearm artery and femoral vein were taken during 4 h before and 4 h after an intravenous endotoxin injection (4 ng/kg body wt). In addition, muscle biopsies from the leg muscle were taken. Plasma concentration of the total sum of amino acids decreased by 19% (P = 0.001) and of glutamine by 25% (P = 0.004) the 3rd h after endotoxin administration. At the same time, muscle concentrations of the sum of amino acids and glutamine decreased by 11% (P = 0.05) and 9% (P = 0.09), respectively. In parallel, the efflux from the leg increased by 35% (P = 0.004) for the total sum of amino acids and by 43% (P = 0.05) for glutamine. In conclusion, intravenous endotoxin administration to healthy volunteers, used as a model for the initial phase of sepsis, resulted in a decrease in plasma amino acid concentrations. At the same time, amino acid concentrations in muscle tissue decreased, whereas the efflux of amino acids from leg skeletal muscle increased.     

Investigating movement in the laboratory: dispersal apparatus designs and the red flour beetle,Tribolium castaneum

The natural dispersal of Tribolium castaneum Herbst (Coleoptera: Tenebrionidae) has been emulated in the laboratory for more than 50 years, using a simple dispersal apparatus. This has typically comprised of a starting container (initial resource or patch) connected by tubing, which contains thread for the animals to climb into a tube and hence to an end container. That is, beetles move to a new viable resource or patch from an inter‐patch zone or non‐viable habitat. We modified this basic apparatus design to test the effect of tubing length and tubing insertion angle on the dispersal rate and proportion of successful dispersers. We expected that the proportion of successful dispersers would be repeatable within each apparatus design, and that increasing tubing length and steepness of the insertion angle would reduce dispersal rate and success across apparatus designs. Dispersal increased linearly through time, similarly so for both males and females. The design with the most vertical tubing insertion angle had a lower proportion of successful dispersers. Tubing length also had a negative relationship with dispersal success (as judged by insects reaching the end container), but a significant reduction in dispersal success was only apparent between the shortest and longest tubing between containers. We suggest that locating and climbing the vertical section of string before they can enter the tubing between containers restricts dispersal and that at higher densities, insects exhibit greater inclination to climb. This type of apparatus has flexible design tolerances and further potential to study the dispersal of other small insect species that primarily use pedestrian locomotion.     

Identification of an Effective Early Signaling Signature during Neo-Vasculogenesis In Vivo by Ex Vivo Proteomic Profiling

Therapeutic neo-vasculogenesis in vivo can be achieved by the co-transplantation of human endothelial colony-forming progenitor cells (ECFCs) with mesenchymal stem/progenitor cells (MSPCs). The underlying mechanism is not completely understood thus hampering the development of novel stem cell therapies. We hypothesized that proteomic profiling could be used to retrieve the in vivo signaling signature during the initial phase of human neo-vasculogenesis. ECFCs and MSPCs were therefore either transplanted alone or co-transplanted subcutaneously into immune deficient mice. Early cell signaling, occurring within the first 24 hours in vivo, was analyzed using antibody microarray proteomic profiling. Vessel formation and persistence were verified in parallel transplants for up to 24 weeks. Proteomic analysis revealed significant alteration of regulatory components including caspases, calcium/calmodulin-dependent protein kinase, DNA protein kinase, human ErbB2 receptor-tyrosine kinase as well as mitogen-activated protein kinases. Caspase-4 was selected from array results as one therapeutic candidate for targeting vascular network formation in vitro as well as modulating therapeutic vasculogenesis in vivo. As a proof-of-principle, caspase-4 and general caspase-blocking led to diminished endothelial network formation in vitro and significantly decreased vasculogenesis in vivo. Proteomic profiling ex vivo thus unraveled a signaling signature which can be used for target selection to modulate neo-vasculogenesis in vivo.     

Influence of phosphine resistance genes on flight propensity and resource location in Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae): the landscape for selection

Phosphine resistance in Tribolium castaneum Herbst (Coleoptera: Tenebrionidae) has evolved through changes to enzymes involved in basic metabolic pathways. These changes impose metabolic stress and could affect energy‐demanding behaviours. We therefore tested whether phosphine resistance alleles impact the movement of these insects in their quest for new resources. We measured walking and flight parameters of four T. castaneum genotypes: (1) a field‐derived population, (2) a laboratory cultured, phosphine‐susceptible reference strain, (3) a laboratory cultured, phosphine‐resistant reference strain, and (4) a resistant introgressed strain that is almost identical genetically to the susceptible population. The temporal pattern of flight was identical across all populations, but resistant beetles took flight significantly less, walked more slowly, and located resources less successfully than did susceptible beetles. Also, the field‐derived beetles (proved not to be carrying resistance genes) walked significantly faster and more directly towards food resources, and had a higher propensity for flight when compared to the susceptible laboratory beetles. These negative effects suggest survival of beetles with the resistance alleles will be compromised should they leave phosphine application sites. The field for selection therefore extends beyond the site at which phosphine fumigant imposed its effect, and other mutations are also likely to be affected in this way.     

A multiparametric activity profiling platform for neuron disease phenotyping and drug screening

Patient stem cell–derived models enable imaging of complex disease phenotypes and the development of scalable drug discovery platforms. Current preclinical methods for assessing cellular activity do not, however, capture the full intricacies of disease-induced disturbances and instead typically focus on a single parameter, which impairs both the understanding of disease and the discovery of effective therapeutics. Here, we describe a cloud-based image processing and analysis platform that captures the intricate activity profile revealed by GCaMP fluorescence recordings of intracellular calcium changes and enables the discovery of molecules that correct 153 parameters that define the amyotrophic lateral sclerosis motor neuron disease phenotype. In a high-throughput screen, we identified compounds that revert the multiparametric disease profile to that found in healthy cells, a novel and robust measure of therapeutic potential quite distinct from unidimensional screening. This platform can guide the development of therapeutics that counteract the multifaceted pathological features of diseased cellular activity.     

Screening and isolation of halophilic bacteria producing extracellular hydrolyses from Howz Soltan Lake,Iran

Screening of bacteria from different areas of Howz Soltan playa, a hypersaline lake in the central desert zone of Iran, led to the isolation of 231 moderately halophilic bacteria, which were able to grow optimally in media with 5–15% of salt, and 49 extremely halophilic microorganisms that required 20–25% of salt for optimal growth. These isolates produced a great variety of extracellular hydrolytic enzymes. A total of 195, 177, 100, 95, 92, 68, 65, 33, and 28 strains produced lipases, amylases, proteases, inulinases, xylanases, cellulases, pullulanases, DNases, and pectinases, respectively. In comparison with gram-negative bacteria, the gram-positive halophilic rods, showed more hydrolytic activities. Several combined activities were showed by some of these isolates. One strain presented 9 hydrolytic activities, 4 strains presented 8 hydrolytic activities, 10 strains presented 7 hydrolytic activities and 29 strains presented 6 hydrolytic activities. No halophilic isolate without hydrolytic activity has been found in this study. According to their phenotypic characteristics and comparative partial 16S rRNA sequence analysis, the halophilic strains were identified as members of the genera: Salicola, Halovibrio, Halomonas, Oceanobacillus, Thalassobacillus, Halobacillus, Virgibacillus, Gracilibacillus, Salinicoccus, and Piscibacillus. Most lipase and DNase producers were members of the genera Gracilibacillus and Halomonas, respectively, whereas most of the isolates able to produce hydrolytic enzymes such as amylase, protease, cellulose (CMCase) and inulinase, belonged to gram-positive genera, like Gracilibacillus, Thalassobacillus, Virgibacillus, and Halobacillus.     

Comparison of Bacterial Biodiversity and Enzyme Production in Three Hypersaline Lakes; Urmia,Howz-Soltan and Aran-Bidgol

This research is a comparative study on the diversity of halophilic bacteria with hydrolytic activities in three significant hypersaline lakes; Urmia in the northwest and Howz-Soltan and Aran-Bidgol in the central desert in Iran. Isolated strains from these saline lakes were found to be halotolerant, moderately and extremely halophilic bacteria. The bacteria in each saline lake were able to produce different hydrolytic enzymes including amylase, protease, lipase, DNase, inulinase, xylanase, carboxy methyl cellulase, pectinase and pullulanase. 188, 302, 91 halophilic strains were isolated from Urmia Lake, Howz-Soltan and Aran-Bidgol playa, respectively. The numbers of Gram-positive strains were more than Gram-negatives, and among Gram-positive bacteria; spore-forming bacilli were most abundant. Due to the unique physico-chemical conditions of the lake environments, the hydrolytic activities of isolated strains were significantly different. For instance, isolated strains from Howz-Soltan playa did not produce pectinase, DNase, amylase, lipase and inulinase, while the isolates from Aran-Bidgol playa had a great ability to produce pectinase and DNase. The strains from Urmia Lake were also good producers of DNase but failed to show any chitinase activity. The diversity of halophilic bacteria from the mentioned three saline lakes was also determined using PCR-amplified 16S rRNA followed by phylogenetic analysis of the partial 16S rRNA sequences.

Electronic supplementary material

The online version of this article (doi:10.1007/s12088-014-0481-9) contains supplementary material, which is available to authorized users.