<Emphasis Type="Italic">In vivo</Emphasis> and <Emphasis Type="Italic">in vitro</Emphasis> antioxidant effects of three Veronica species

The aim of the present study was to examine the antioxidant activity of three Veronica species (Plantaginaceae). The antioxidant potential of various extracts obtained from aerial flowering parts was evaluated by DPPH-free (1,1-diphenyl-2-picrylhydrazyl-free) radical scavenging activity and ferric-reducing antioxidant power assays. Considerable antioxidant activity was observed in the plant samples (FRAP values ranged from 0.97 to 4.85 mmol Fe²⁺/g, and DPPH IC₅₀ values from 12.58 to 66.34 μg/ml); however, these levels were lower than the activity of the control compound butylated hydroxytoluene (BHT) (FRAP: 10.58 mmol Fe²⁺/g; DPPH IC₅₀: 9.57 μg/ml). Also, the in vivo antioxidant effects were evaluated in several hepatic antioxidant systems in rats (activities of glutathione peroxidase, glutathione reductase, peroxidase, catalase, xanthine oxidase, glutathione content and level of thiobarbituric acid reactive substances) after treatment with different Veronica extracts, or in combination with carbon tetrachloride (CCl₄). Pretreatment with 100 mg/kg b.w. of Veronica extracts inhibited CCl₄-induced liver injury by decreasing TBA-RS level, increasing GSH content, and bringing the activities of CAT and Px to control levels. The present study suggests that the extracts analyzed could protect the liver cells from CCl₄-induced liver damage by their antioxidative effect on hepatocytes.     

Differential protein pathways in 1,25-dihydroxyvitamin d(3) and dexamethasone modulated tolerogenic human dendritic cells

Tolerogenic dendritic cells (DC) that are maturation-resistant and locked in a semimature state are promising tools in clinical applications for tolerance induction. Different immunomodulatory agents have been shown to induce a tolerogenic DC phenotype, such as the biologically active form of vitamin D (1,25(OH)(2)D(3)), glucocorticoids, and a synergistic combination of both. In this study, we aimed to characterize the protein profile, function and phenotype of DCs obtained in vitro in the presence of 1,25(OH)(2)D(3), dexamethasone (DEX), and a combination of both compounds (combi). Human CD14(+) monocytes were differentiated toward mature DCs, in the presence or absence of 1,25(OH)(2)D(3) and/or DEX. Cells were prefractionated into cytoplasmic and microsomal fractions and protein samples were separated in two different pH ranges (pH 3-7NL and 6-9), analyzed by 2D-DIGE and differentially expressed spots (p < 0.05) were identified after MALDI-TOF/TOF analysis. In parallel, morphological and phenotypical analyses were performed, revealing that 1,25(OH)(2)D(3)- and combi-mDCs are closer related to each other than DEX-mDCs. This was translated in their protein profile, indicating that 1,25(OH)(2)D(3) is more potent than DEX in inducing a tolerogenic profile on human DCs. Moreover, we demonstrate that combining 1,25(OH)(2)D(3) with DEX induces a unique protein expression pattern with major imprinting of the 1,25(OH)(2)D(3) effect. Finally, protein interaction networks and pathway analysis suggest that 1,25(OH)(2)D(3), rather than DEX treatment, has a severe impact on metabolic pathways involving lipids, glucose, and oxidative phosphorylation, which may affect the production of or the response to ROS generation. These findings provide new insights on the molecular basis of DC tolerogenicity induced by 1,25(OH)(2)D(3) and/or DEX, which may lead to the discovery of new pathways involved in DC immunomodulation.     

Turning the 'mustard oil bomb' into a 'cyanide bomb': aromatic glucosinolate metabolism in a specialist insect herbivore

Plants have evolved a variety of mechanisms for dealing with insect herbivory among which chemical defense through secondary metabolites plays a prominent role. Physiological, behavioural and sensorical adaptations to these chemicals provide herbivores with selective advantages allowing them to diversify within the newly occupied ecological niche. In turn, this may influence the evolution of plant metabolism giving rise to e.g. new chemical defenses. The association of Pierid butterflies and plants of the Brassicales has been cited as an illustrative example of this adaptive process known as 'coevolutionary armsrace'. All plants of the Brassicales are defended by the glucosinolate-myrosinase system to which larvae of cabbage white butterflies and related species are biochemically adapted through a gut nitrile-specifier protein. Here, we provide evidence by metabolite profiling and enzyme assays that metabolism of benzylglucosinolate in Pieris rapae results in release of equimolar amounts of cyanide, a potent inhibitor of cellular respiration. We further demonstrate that P. rapae larvae develop on transgenic Arabidopsis plants with ectopic production of the cyanogenic glucoside dhurrin without ill effects. Metabolite analyses and fumigation experiments indicate that cyanide is detoxified by β-cyanoalanine synthase and rhodanese in the larvae. Based on these results as well as on the facts that benzylglucosinolate was one of the predominant glucosinolates in ancient Brassicales and that ancient Brassicales lack nitrilases involved in alternative pathways, we propose that the ability of Pierid species to safely handle cyanide contributed to the primary host shift from Fabales to Brassicales that occured about 75 million years ago and was followed by Pierid species diversification.     

Regulation of collagen gene expression in the Tsk2 mouse

The tight skin 2 (Tsk2) mutation is an ENU induced dominant mutation localized on mouse chromosome 1. While the molecular defect is unknown, Tsk2/+ mice display cutaneous thickening associated with excessive matrix production and are used as a model of scleroderma. The purpose of this study was to examine the cellular mechanisms associated with the excessive synthesis of matrix macromolecules using a collagen promoter GFP reporter transgene (pOBCol3.6GFP) as a marker of Col1a1 expression. This analysis of pOBCol3.6GFP expression in Tsk2/+ skin showed an increase in transgene activity compared to wild-type (+/+) samples. In addition, an increased area of "high" GFP fluorescence in Tsk2/+ dermis in both 1- and 4-month-old mice was observed that was also associated with an increased number of dermal fibroblasts per unit area of dermis. These data collectively suggest an important mechanism of Tsk2/+ skin fibrosis; an increased number of collagen expressing cells as well as elevated collagen expression on a per cell basis. During this study it was noted that Tsk2/+ mice appeared consistently smaller than wild-type (+/+) siblings and measurements of body length revealed a decrease (5-10%) in 1- and 2-month-old Tsk2/+ mice as well as a decrease in body weight in both age groups as compared to wild-type (+/+) control mice. Femur length was also decreased (2-9%) in Tsk2/+ mice. Finally, in contrast to Tsk/+ mice that display an emphysema-like lung pathology, histological sections of lungs from Tsk2/+ mice were normal and indistinguishable from wild-type (+/+) controls.     

The release of soluble forms of TRAIL and DR5 by neutrophils of oral cavity cancer patients

In the present study we examined the release of the soluble form of TRAIL by neutrophils (PMN) derived from patients with oral cavity cancer. Simultaneously, we estimated the ability of PMNs of these patients to release the soluble form of DR5 receptor, a natural regulatory protein of TRAIL. The obtained results were confronted with the serum levels of sTRAIL and sDR5. The cells were isolated from 21 patients with squamous cell carcinoma of oral cavity at diagnosis and three weeks after surgery treatment. For comparative purposes we performed similar examinations in autologous peripheral blood mononuclear cells (PBMC). Cytoplasmic protein fractions of the cells were analyzed for the presence of TRAIL and DR5 by western blotting. Soluble TRAIL and soluble DR5 concentrations in the culture supernatants of cells were confronted with their serum levels using ELISA kit. PMN and PBMC of the whole cancer patient group expressed decreased TRAIL protein and unchanged expression of DR5 receptor in comparison with the control group. Unchanged release of sTRAIL by PMNs of patients in Stage II was accompanying the decrease of the ability of PBMC to secrete this protein. In patients in Stage IV the secretion of sTRAIL by PMNs and PBMC was impaired. In contrast to changes in sTRAIL secretion by PMN and PBMC of oral cavity cancer patients, the secretion of sDR5 by these cells was unchanged. The serum levels of sTRAIL were increased in patients in Stage II before treatment and decreased in the same patients after treatment. The altered ability of PMN of PBMC to secrete sTRAIL may have different implications for the immune response of patients with oral cavity cancer cells at different stages of disease.     

The contribution of single and colonial cells of Phaeocystis pouchetii to spring and summer blooms in the north-eastern North Atlantic

Studies of the phytoplankton ecology in different localities in north-Norwegian fjords, the White Sea and the Barents Sea were carried out in spring and early summer to investigate the contribution of single and colonial stages of Phaeocystis pouchetii to phytoplankton abundance. Three different types of flagellated and four colonial cells were observed in all localities. P. pouchetii was rare under the ice of the Barents and White Seas, but their abundance increased rapidly during ice retreat. Single cell C dominated over colonial cell C, often by 50 times or more. The highest share of colonial cells was encountered in April in northern Norwegian fjords, in May in the Barents Sea and in May–June in the White Sea. At times the single cell dominated the total P. pouchetii biomass in Balsfjord (April 1999, 2001) with hardly any colonies present. In the White Sea colonies of P. pouchetii were less abundant than in the other regions. Cell carbon of P. pouchetii colonies appears never to be as dominating in the north-eastern North Atlantic as P. globosa blooms in coastal regions such as the southern North Sea. However, the lobal matrix of P. pouchetii colonies appears to be less solid than that of P. globosa and partly dissolution of the colony matrix during handling and storage of fixes samples induces uncertainty about the absolute numbers of P. pouchetii colonial cell counts. Despite of that, single cells of P. pouchetii seem to dominate significantly over colonial cell biomass at most sites and during some years and in some regions colonial cells seem rare. We speculate that top-down regulation of Phaeocystis spp. blooms possibly determines the ratio between single and colonial cells.