Immunomodulatory,insulinotropic, and cytotoxic activities of phylloseptins and plasticin‐TR from the Trinidanian leaf frog Phyllomedusa trinitatis

The aim of the study was to determine the in vitro immunomodulatory, cytotoxic, and insulin‐releasing activities of seven phylloseptin‐TR peptides and plasticin‐TR, first isolated from the frog Phyllomedusa trinitatis. The most cationic peptides, phylloseptin‐1.1TR and phylloseptin‐3.1TR, showed greatest cytotoxic potency against A549, MDA‐MB231, and HT‐29 human tumor‐derived cells and against mouse erythrocytes. Phylloseptin‐4TR was the most hydrophobic and the most effective peptide at inhibiting production of the proinflammatory cytokines TNF‐α and IL‐1β by mouse peritoneal cells but was without effect on production of the antiinflammatory cytokine IL‐10. Phylloseptin‐2.1TR and phylloseptin‐3.3TR were the most effective at stimulating the production of IL‐10. The noncytotoxic peptide, plasticin‐TR, inhibited production of TNF‐α and IL‐1β but was without effect on IL‐10 production. The results of CD spectroscopy suggest that the different properties of plasticin‐TR compared with the immunostimulatory activities of the previously characterized plasticin‐L1 from Leptodactylus laticeps may arise from greater ability of plasticin‐TR to oligomerize and adopt a stable helical conformation in a membrane‐mimetic environment. All peptides stimulated release of insulin from BRIN‐BD11 rat clonal β cells with phylloseptin‐3.2TR being the most potent and effective and phylloseptin‐2.1TR the least effective suggesting that insulinotropic potency correlates inversely with helicity. The study has provided insight into structure‐activity relationships among the phylloseptins. The combination of immunomodulatory and insulinotropic activities together with low cytotoxicity suggests that phylloseptin‐3.3TR and plasticin‐TR may represent templates for the development of agents for use in antiinflammatory and type 2 diabetes therapies.     

Evidence for the plasticity of arthropod signal transduction pathways

Metazoans are known to contain a limited, yet highly conserved, set of signal transduction pathways that instruct early developmental patterning mechanisms. Genomic surveys that have compared gene conservation in signal transduction pathways between various insects and Drosophila support the conclusion that these pathways are conserved in evolution. However, the degree to which individual components of signal transduction pathways vary among more divergent arthropods is not known. Here, we report our results of a survey of the genome of the two-spotted spider mite Tetranychus urticae, using a set of 294 Drosophila orthologs of genes that function in signal transduction. We find a third of all genes surveyed absent from the spider mite genome. We also identify several novel duplications that have not been previously reported for a chelicerate. In comparison with previous insect surveys, Tetranychus contains a decrease in overall gene conservation, as well as an unusual ratio of ligands to receptors and other modifiers. These findings suggest that gene loss and duplication among components of signal transduction pathways are common among arthropods and suggest that signal transduction pathways in arthropods are more evolutionarily labile than previously hypothesized.     

Macrophage colony stimulating factor (CSF-1) is a central growth factor of goldfish macrophages

We recently characterized macrophage colony stimulating factor (CSF-1) of fish (the goldfish). Here, we report for the first time that goldfish CSF-1 acts through the CSF-1 receptor by showing loss of CSF-1 function in CSF-1R knockdown monocytes using RNAi, and demonstrate that goldfish CSF-1 administration in vivo increases the amount of circulating monocytes in blood. We also show that conditioned supernatants from goldfish fibroblast cultures induced the proliferation of goldfish monocytes indicating that, like in mammals, teleost fibroblasts are an important producer of CSF-1. The continuous addition of recombinant CSF-1 to primary goldfish macrophage cultures stabilized and extended their longevity and resulted in a long-term culture of functional macrophages capable of mounting a potent nitric oxide response upon activation with goldfish recombinant TNF-alpha.     

An excess vessel in the posterior part of the human cerebral arterial circle (CAC): a case series

Background  

As a continuation of the previous findings in human fetuses, accidental finding of an accessory vascular component in the posterior part of CAC of human adult cadavers inspired the authors to present and compare its posterior part configuration.     

Repeated use of yeast biomass in ethanol fermentation of juniper berry sugars

Summary The object of this study was to establish the possibility of using the yeast biomass separated from the fermentation broth at the end of ethanol fermentation of juniper berry sugars as an inoculum in successive batch fermentation processes. A part of the fermentation broth (10% v/v) and a suspension of yeast biomass (separated from the same broth) into the water extract of juniper berries (2 g of wet yeast biomass per liter of water extract) were used as inocula. It was shown that the suspension of yeast biomass could be used as inoculum in successive batch processes without negative effects on the kinetics and ethanol yield, but with positive effects on the capacity and economy of the bioprocess. The addition of ammonium salts at optimum levels did not affect the final ethanol concentrations (4.3–4.4% v/v), but enhanced the specific rate of ethanol production, thus reducing the process duration by about five times.     

Recombinant glycoprotein 63 (Gp63) of Trypanosoma carassii suppresses antimicrobial responses of goldfish (Carassius auratus L.) monocytes and macrophages

We previously reported that proteins secreted by Trypanosoma carassii play a role in evasion of fish host immune responses. To further understand how these parasites survive in the host, we cloned and expressed T. carassii glycoprotein 63 (Tcagp63), and generated a rabbit polyclonal antibody to the recombinant protein (rTcagp63). Tcagp63 was similar to gp63 of other trypanosomes and grouped with Trypanosoma cruzi and Trypanosoma brucei gp63 in phylogenetic analysis. We showed that rTcagp63 down-regulated Aeromonas salmonicida and recombinant goldfish TNFα2-induced production of reactive oxygen and nitrogen intermediates. Macrophages treated with rTcagp63 also exhibited significant reduction in the expression of inducible nitric oxide synthase (iNOS)-A, TNFα-1 and TNFα-2. Recombinant Tcagp63 bound to and was internalised by goldfish macrophages. The Tcagp63 may act by altering the signalling events important in downstream monocyte/macrophage antimicrobial and other cytokine-induced functions. We believe that this is the first report on downregulation of antimicrobial responses by trypanosome gp63.     

The effect of thermal stress on Ostrini00a nubilalis HBN (Lepidoptera, Pyralidae)—I. The orle of the neurosecretory cells in the survival of diapausing larvae

1. 1.|The effect of short-term exposure (up to 12 h) to upper limiting temperature (45°C) on A-type medial neurosecretory cells (MNSC) of the protocerebrum of Ostrinia nubilalis larvae was studied.

2. 2.|For evaluation of the activity of MNSC, cytological parameters were used: quantity of the neurosecretory material, size of the neurosecretory granules, shape of the nuclei and size of the nucleoli.

3. 3.|After 1 h exposure to 45°C the activity of the MNSC was increased, after 3 h it was decreased, while after 6 h exposure to the same temperature some renewed increase in the activity was observed.

4. 4.|The significance of oscillatory changes in the activity of A-type MNSC in survival of the thermal stress of O. nubilalis diapausing larvae is discussed.

Transformation of the naturally occurring frog skin peptide, alyteserin-2a into a potent, non-toxic anti-cancer agent

Alyteserin-2a (ILGKLLSTAAGLLSNL.NH₂) is a cationic, amphipathic α-helical cell-penetrating peptide, first isolated from skin secretions of the midwife toad Alytes obstetricans. Structure–activity relationships were investigated by synthesizing analogs of alyteserin-2a in which amino acids on the hydrophobic face of the helix were replaced by l-tryptophan and amino acids on the hydrophilic face were replaced by one or more l-lysine or d-lysine residues. The Trp-containing peptides display increased cytotoxic activity against non-small cell lung adenocarcinoma A549 cells (up to 11-fold), but hemolytic activity against human erythrocytes increases in parallel. The potency of the N15K analog against A549 cells (LC₅₀ = 13 μM) increases sixfold relative to alyteserin-2a and the therapeutic index (ratio of LC₅₀ for erythrocytes and tumor cells) increases twofold. Incorporation of a d-Lys¹¹ residue into the N15K analog generates a peptide that retains potency against A549 cells (LC₅₀ = 15 μM) but whose therapeutic index is 13-fold elevated relative to the native peptide. [G11k, N15K] alyteserin-2a is also active against human hepatocarcinoma HepG2 cells (LC₅₀ = 26 μM), breast adenocarcinoma MDA-MB-231 cells (LC₅₀ = 20 μM), and colorectal adenocarcinoma HT-29 cells (LC₅₀ = 28 μM). [G11k, N15K] alyteserin-2a, in concentrations as low as 1 μg/mL, significantly (P < 0.05) inhibits the release of the immune-suppressive cytokines IL-10 and TGF-β from unstimulated and concanavalin A-stimulated peripheral blood mononuclear cells. The data suggest a strategy of increasing the cationicity while reducing the helicity of naturally occurring amphipathic α-helical peptides to generate analogs with improved cytotoxicity against tumor cells but decreased activity against non-neoplastic cells.     

Prenatal glucocorticoid exposure in the sheep alters renal development in utero: implications for adult renal function and blood pressure control

Treatment of the pregnant ewe with glucocorticoids early in pregnancy results in offspring with hypertension. This study examined whether glucocorticoids can reduce nephron formation or alter gene expression for sodium channels in the late gestation fetus. Sodium channel expression was also examined in 2-mo-old lambs, while arterial pressure and renal function was examined in adult female offspring before and during 6 wk of increased dietary salt intake. Pregnant ewes were treated with saline (SAL), dexamethasone (DEX; 0.48 mg/h) or cortisol (CORT; 5 mg/h) over days 26-28 of gestation (term = 150 days). At 140 days of gestation, glomerular number in CORT and DEX animals was 40 and 25% less, respectively, compared with SAL controls. Real-time PCR showed greater gene expression for the epithelial sodium channel (α-, β-, γ-subunits) and Na(+)-K(+)-ATPase (α-, β-, γ-subunits) in both the DEX and CORT group fetal kidneys compared with the SAL group with some of these changes persisting in 2-mo-old female offspring. In adulthood, sheep treated with dexamethasone or cortisol in utero had elevated arterial pressure and an apparent increase in single nephron glomerular filtration rate, but global renal hemodynamics and excretory function were normal and arterial pressure was not salt sensitive. Our findings show that the nephron-deficit in sheep exposed to glucocorticoids in utero is acquired before birth, so it is a potential cause, rather than a consequence, of their elevated arterial pressure in adulthood. Upregulation of sodium channels in these animals could provide a mechanistic link to sustained increases in arterial pressure in cortisol- and dexamethasone-exposed sheep, since it would be expected to promote salt and water retention during the postnatal period.