In vitro and in vivo characterization of an interleukin‐15 antagonist peptide by metabolic stability, 99mTc‐labeling,and biological activity assays

Interleukin (IL)–15 is an inflammatory cytokine that constitutes a validated therapeutic target in some immunopathologies, including rheumatoid arthritis (RA). Previously, we identified an IL‐15 antagonist peptide named [K6T]P8, with potential therapeutic application in RA. In the current work, the metabolic stability of this peptide in synovial fluids from RA patients was studied. Moreover, [K6T]P8 peptide was labeled with ^99mTc to investigate its stability in human plasma and its biodistribution pattern in healthy rats. The biological activity of [K6T]P8 peptide and its dimer was evaluated in CTLL‐2 cells, using 3 different additives to improve the solubility of these peptides. The half‐life of [K6T]P8 in human synovial fluid was 5.88 ± 1.73 minutes, and the major chemical modifications included peptide dimerization, cysteinylation, and methionine oxidation. Radiolabeling of [K6T]P8 with ^99mTc showed a yield of approximately 99.8%. The ^99mTc‐labeled peptide was stable in a 30‐fold molar excess of cysteine and in human plasma, displaying a low affinity to plasma proteins. Preliminary biodistribution studies in healthy Wistar rats suggested a slow elimination of the peptide through the renal and hepatic pathways. Although citric acid, sucrose, and Tween 80 enhanced the solubility of [K6T]P8 peptide and its dimer, only the sucrose did not interfere with the in vitro proliferation assay used to assess their biological activity. The results here presented, reinforce nonclinical characterization of the [K6T]P8 peptide, a potential agent for the treatment of RA and other diseases associated with IL‐15 overexpression.     

Physical factors driving intertidal macroalgae distribution: physiological stress of a dominant fucoid at its southern limit

Climate change is driving species range shifts worldwide. However, physiological responses related to distributional changes are not fully understood. Oceanographers have reported an increase in ocean temperature in the northwest Iberian Peninsula that is potentially related to the decline in some cold-temperate intertidal macroalgae in the Cantabrian Sea, namely Fucus serratus. Low tide stress could also play a role in this decline. We performed one mensurative (in situ) and two manipulative (in culture) experiments designed to evaluate the interactive effects of some physical factors. The first experiment analysed field response to low tide stress in marginal (mid-Cantabrian Sea and northern Portugal) versus central (Galicia) populations of F. serratus. Then a second experiment was performed that utilized either harsh or mild summer conditions of atmospheric temperature, irradiance, humidity, and wind velocity to compare the responses of individuals from one marginal and one central population to low tide stress. Finally, the combined effect of sea temperature and the other factors was evaluated to detect interactive effects. Changes in frond growth, maximal photosynthetic quantum yield (F (v)/F (m)), temperature, and desiccation were found. Three additive factors (solar irradiation, ocean and air temperatures) were found to drive F. serratus distribution, except under mildly humid conditions that ameliorated atmospheric thermal stress (two additive factors). Mid-Cantabrian Sea temperatures have recently increased, reaching the inhibitory levels suggested in this study of F. serratus. We also expect an additive secondary contribution of low tide stress to this species decline. On the northern Portugal coast, ocean warming plus low tide stress has not reached this species' inhibition threshold. No significant differential responses attributed to the population of origin were found. Mechanistic approaches that are designed to analyse the interactive effects of physical stressors may improve the levels of confidence in predicted range shifts of species.     

Ecology,morphology and physiology of Chroothece richteriana (Rhodophyta,Stylonematophyceae) in the highly calcareous Río Chícamo,south-east Spain

The ecology of Chroothece was studied in the highly calcareous Río Chícamo, south-east Spain, in order to explain its success there, but rarity elsewhere. The river, which originates mainly from an underground aquifer, has water with high conductivity, sulphate and nitrate but low phosphate concentrations, the latter mainly organic. Chroothece occurs in mats and in lobed colonies reaching 4 cm in the broadest dimension. The colony surface consists of one layer of cells, each of which is attached to a stalk, which dichotomizes when the cell divides; stalks often extend to the colony base. The central region of many mat cells and almost all colony cells has a yellow to orange-brown colour, associated with the numerous lipid droplets densely covering the surface of the pyrenoid and arms of the star-shaped chloroplast. Field material and laboratory isolates indicate that stalk formation occurs under moderate P limitation and both stalks and cell sheath show high phosphatase activities. This also occurred in a culture collection strain maintained for 30 years in a very P-rich medium, but then transferred to a moderately P-limiting medium (c. 0.9 mg l^?1). We suggest that colony formation is initiated by aggregation of motile cells following P pulses in the water. Comparisons are made with Rivularia, a competitor in this nitrate-rich river, in spite of being a N₂-fixer. One difference is that Chroothece cells lie at the periphery of the colonies and are therefore exposed to maximum sunlight, whereas Rivularia trichomes grow inside colonies with photoprotection by scytonemin. However, the ability to withstand heavy grazing pressure may be an especially important factor favouring Rivularia here.     

Differential effect of pertussis toxin on the affinity state for agonists of renal alpha 1- and alpha 2- adrenoceptors

The effect of pertussis toxin treatment on the guanine nucleotide-induced modulation of the affinity of renal alpha 1- and alpha 2-adrenergic receptors was investigated. Pretreatment of rats with pertussis toxin did not induce any change in the number of or affinity for antagonists of alpha 1- or alpha 2-receptors studied using [3H]prazosin and [3H]yohimbine, respectively. Guanyl-5'-yl imidodiphosphate induced an "up-shift" in the number of alpha 2-adrenergic receptors; this up-shift was not observed for alpha 1-adrenergic receptors. Pertussis toxin treatment decreased the affinity of epinephrine for the [3H]yohimbine-binding sites and reduced the ability of guanine nucleotides to modulate alpha 2-adrenoceptor agonist affinity. The regulation by guanine nucleotides of alpha 1-adrenoceptor affinity for agonists was not altered. These results suggest that the modulation of alpha 1- and alpha 2-adrenoceptors by guanine nucleotides is probably exerted through different molecular entities.     

Relationships between leaf morphological traits, nutrient concentrations and isotopic signatures for Mediterranean woody plant species and communities

Background and aims

Soil factors are driving forces that influence spatial distribution and functional traits of plant species. We test whether two anchor morphological traits—leaf mass per area (LMA) and leaf dry matter content (LDMC)—are significantly related to a broad range of leaf nutrient concentrations in Mediterranean woody plant species. We also explore the main environmental filters (light availability, soil moisture and soil nutrients) that determine the patterns of these functional traits in a forest stand.

Methods

Four morphological and 19 chemical leaf traits (macronutrients and trace elements and δ¹³C and δ¹⁵N signatures) were analysed in 17 woody plant species. Community-weighted leaf traits were calculated for 57 plots within the forest. Links between LMA, LDMC and other leaf traits were analysed at the species and the community level using standardised major axis (SMA) regressions

Results

LMA and LDMC were significantly related to many leaf nutrient concentrations, but only when using abundance-weighted values at community level. Among-traits links were much weaker for the cross-species analysis. Nitrogen isotopic signatures were useful to understand different resource-use strategies. Community-weighted LMA and LDMC were negatively related to light availability, contrary to what was expected.

Conclusion

Community leaf traits have parallel shifts along the environmental factors that determine the community assembly, even though they are weakly related across individual taxa. Light availability is the main environmental factor determining this convergence of the community leaf traits.     

A moderate intake of phytosterols from habitual diet affects cholesterol metabolism

Cholesterol metabolism homeostasis is the result of a balance between synthesis, degradation and intestinal absorption. It is well established that intestinal cholesterol absorption efficiency can be modified by the intake of phytosterol-enriched food and, therefore, have a serum cholesterol-lowering effect. Recent epidemiological and clinical studies have shown that presence of phytosterols at normal diet levels could also be effective on lowering total and LDL serum cholesterol since they affect whole-body cholesterol metabolism even at those moderate doses. The aim of this study was to analyze the effect of the levels of the naturally-occurring phytosterols in the diet on cholesterol metabolism parameters. In order to do that a group of 99 healthy volunteers was studied for their dietary habits and surrogate markers of cholesterol synthesis and absorption. The mean daily dietary intake of phytosterols, measured by a food semiquantitative frequency questionnaire, was found to be 494 mg being β-sitosterol the major contributor to it. Subjects were classified into tertiles according to their total phytosterol intake and comparisons were done between subgroups. No statistical differences were observed for surrogate markers of intestinal cholesterol absorption, but a significant increase in the cholesterol synthesis surrogate marker lathosterol-to-cholesterol ratio associated to highest dietary phytosterol intake was observed. Regardless of this, only a non significant trend toward a less atherogenic lipid profile was observed in the upper tertile. In conclusion, the intake of moderate amounts of phytosterols naturally present in habitual diet may affect cholesterol metabolism and specially the rate of cholesterol synthesis as estimated by the surrogate marker lathosterol-to-cholesterol ratio in serum.     

Agrobacterium-mediated transformation of tomatillo (Physalis ixocarpa) and tissue specific and developmental expression of the CaMV 35S promoter in transgenic tomatillo plants

A protocol for the Agrobacterium-mediated transformation of tomatillo was developed. Up to 40 transgenic plants could be obtained in experiments using 60 cotyledon expiants. The transformed nature of the regenerated plants was confirmed by NPT II and Southern blot hybridization analysis. Using the b-glucuronidase system the tissue specific and developmental patterns of expression of the Cauliflower Mosaic Virus 35S promoter were determined in transgenic tomatillo plants. It was found that this promoter is developmentally regulated during fruit and seed formation.     

19q distal trisomy due to a de novo (19;22)(q13.2;p11) translocation

A 2 4/12-year-old girl whith a de novo 46,XX,-22 + der(22), t(19;22)(q13.2;p11) karyotype is described. From this and other eight similar cases previously published, a typical phenotype in distal 19q trisomy is concluded.