Variable domain antibodies specific for viral hemorrhagic septicemia virus (VHSV) selected from a randomized IgNAR phage display library

Phage display libraries are used to screen for nucleotide sequences that encode immunoglobulin variable (V) regions that are specific for a target antigen. We previously constructed an immunoglobulin new antigen receptor (IgNAR) phage display library. Here we used this library to obtain an IgNAR V region that is specific for viral hemorrhagic septicemia virus (VHSV). A phage clone (clone 653) was found to be specific for VHSV by the biopanning method. The V region of clone 653 was used to construct a 6 × His tagged recombinant IgNAR-653 V protein (rIgNAR-653) using the Escherichia coli pET system. The rIgNAR-653 protein bound specifically to VHSV, confirming its activity.     

Experimental Verification of the Effects on Normal Domestic Cats by Feeding Prescription Diet for Decreasing Stress

The objective of this study was to evaluate the effects of diet on the feline stress response by measuring plasma and urinary cortisol. A study diet was developed with a unique combination of nutrients that supports the management of stressful situations. The specific formulation of the diet included alpha-casozepine, which is believed to have an anxiolytic effect, and tryptophan supplementation. Tryptophan is the precursor for the synthesis of the neurotransmitter serotonin. Twenty-one indoor cats were fed with the study diet (n = 10) or a control diet (n = 11) for 8 weeks, after which physiological responses were evaluated. The study diet significantly increased the ratio of plasma tryptophan to large neutral amino acids and decreased urinary cortisol concentrations after being consumed daily for 8 weeks, but there was no effect on plasma cortisol levels following a stressful event (veterinary examination and blood draw). Further studies, such as behavioral analyses, are needed to clarify the effects of the study diet.     

Nitrotyrosine formation and its role in various pathological conditions

The formation of peroxynitrite and nitrotyrosine was examined in a variety of in vitro and in vivo animal models and its relation to cell or tissue damage was examined. In polymorphonuclear leukocyte (PMN)-induced injury to cardiac myocytes or endothelial cells, activated PMN produced peroxynitrite. Peroxynitrite appears to be responsible for the injury but it was not a major mediator of endothelial cell injury. In the experiment of ischemia-reperfusion injury of the rat brain nitrotyrosine was formed in the peri-infarct and core-of infarct regions. The degradation curve of nitrotyrosine revealed that its t1/2 was about 2.2 hours. In the radiation-induced lung injury of rats, nitrotyrosine was also formed but it was not the sole mechanism for the injury. Levels of nitrotyrosine correlated with the severity of myocardial dysfunction in the canine model of cytokine-induced cardiac injury. Inhibition of NO generation abolished the formation of peroxynitrite and nitrotyrosine in all experiments. In conclusion; although nitrotyrosine is formed in a variety of pathological conditions where the generation of NO is increased, its presence does not always correlate with the severity of injury.     

Myeloperoxidase has Directly-opposed Effects on Nitration Reaction--Study on Myeloperoxidase-deficient Patient and Myeloperoxidase-knockout Mice

Myeloperoxidase (MPO) catalyzes a nitration reaction to form nitrotyrosine in the presence of high nitrite, the metabolite of NO. Human leukocyte was shown to cause phenolic nitration using released MPO as a catalyst in the presence of nitrite. It opposes our previous finding that inhibition of MPO was essential for phenol nitration in human leukocyte study. To clarify the role of MPO, we utilized MPO-deficient human leukocytes and MPO-knockout mice. Even in the absence of exogenously added nitrite, high nitration product was observed in MPO-deficient leukocytes. In liver subjected to ischemia/reperfusion injury, a significantly higher amount of nitrotyrosine was produced in MPO-knockout mice than in normal mice. These results clearly demonstrate that MPO inhibits the accumulation of nitration products in vivo . Further experiments showed that MPO could degrade nitrotyrosine in the presence of glutathione. Thus, MPO-induced degradation of nitration products may cause the underestimation of the nitration product generated in vivo . We conclude that MPO may act predominantly to scavenge nitrotyrosine under physiological nitrite condition, and protect against injurious effect of nitrotyrosine.     

Tailoring the Peptide-Binding Specificity Of An RNA by Combinations of Specificity-Altering Mutations

In this study, the ability to tailor the peptide-binding specificity of an RNA was investigated. First, variants of the Rev-response element (RRE) RNA with different specificities toward the natural binding partner, Rev, and two RRE-binding aptamers, the RSG-1.2 and the Kl peptides, were identified. Next, hybrid RRE mutants with combinations of two sets of specificity-altering substitutions were tested for peptide-binding specificity. It was shown that in most cases the results of the combination of individual mutations were of an additive nature, therefore providing a way to manipulate the peptide-binding specificity of an RNA in a predictable manner.     

Metabolome analysis of food-chain between plants and insects

Evolution has shown the co-dependency between host plants and predators (insects), especially inevitable dependency of predators on plant biomass for securing their energy sources. It was postulated that NAD⁺ source used for major energy producing pathway is the glycerol-3-phosphate shuttle in insects. Using high throughput metabolomics approach, we found that larva of leaf beetle (Gastrophysa atrocyanea), which feed oxalate-rich plants (Rumex obtusifolius), possessed a unique mechanism for accumulating unusually high amounts of lactate. Similarly, larva of butterfly (Papilio machaon) fed with fennel (Foeniculum vulgare) accumulated lactate. Same butterfly also showed the elevated level of glycerol-3-phosphate equivalent to lactate. These evidences provide new insights into the mechanism underlying metabolite alteration between host plants and insect herbivores.