Identification of a novel mechanism for LFA-1 organization during NK cytolytic response

The elimination of transformed and viral infected cells by natural killer (NK) cells requires a specialized junction between NK and target cells, denominated immunological synapse (IS). After initial recognition, the IS enables the directed secretion of lytic granules content into the susceptible target cell. The lymphocyte function-associated antigen (LFA)-1 regulates NK effector function by enabling NK-IS assembly and maturation. The pathways underlying LFA-1 accumulation at the IS in NK cells remained uncharacterized. A kinase anchoring protein 350 (AKAP350) is a centrosome/Golgi-associated protein, which, in T cells, participates in LFA-1 activation by mechanisms that have not been elucidated. We first evaluated AKAP350 participation in NK cytolytic activity. Our results showed that the decrease in AKAP350 levels by RNA interference (AKAP350KD) inhibited NK-YTS cytolytic activity, without affecting conjugate formation. The impairment of NK effector function in AKAP350KD cells correlated with decreased LFA-1 clustering and defective IS maturation. AKAP350KD cells that were exclusively activated via LFA-1 showed impaired LFA-1 organization and deficient lytic granule translocation as well. In NK AKAP350KD cells, activation signaling through Vav1 was preserved up to 10 min of interaction with target cells, but significantly decreased afterwards. Experiments in YTS and in ex vivo NK cells identified an intracellular pool of LFA-1, which partially associated with the Golgi apparatus and, upon NK activation, redistributed to the IS in an AKAP350-dependent manner. The analysis of Golgi organization indicated that the decrease in AKAP350 expression led to the disruption of the Golgi integrity in NK cells. Alteration of Golgi function by BFA treatment or AKAP350 delocalization from this organelle also led to impaired LFA-1 localization at the IS. Therefore, this study characterizes AKAP350 participation in the modulation of NK effector function, revealing the existence of a Golgi-dependent trafficking pathway for LFA-1, which is relevant for LFA-1 organization at NK-lytic IS.     

Same species,different population dynamics: Spatio-temporal differences of Undaria pinnatifida (Ochrophyta,Phaeophyceae) in the intertidal of North Patagonia,Argentina

Population dynamics can be influenced by physical and biological factors, particularly in stressful environments. Introduced species usually have great physiological plasticity, resulting in populations with different traits. Undaria pinnatifida, a macroalga originally described from northeast Asia, was introduced in Northern Patagonia, Argentina (San Matías Gulf) around 2010. To describe the spatio-temporal variability in population structure and morphometry of U. pinnatifida, we conducted monthly field samplings for 2 years at the intertidal area of two contrasting sites in the San Matías Gulf. Individuals of U. pinnatifida were classified by developmental stage, and their morpho-gravimetric variables were measured. In both intertidal sites juveniles were found in higher proportion during austral autumn and grew and matured during the autumn-winter months (from May onwards), and individuals senesced during early austral summer (December and January). Conversely, density and biomass were largely different between sites, and individuals showed slight morphological variability between sites. Environmental (e.g., nutrient concentration, available substrate) and biological factors (e.g., facilitation, competition) may explain the observed differences. Since there is not a macroalga with U. pinnatifida morphometrical characteristics in the intertidal environments of San Matías Gulf, studying this recent introduction gives us a better understanding of its potential ecological effects.     

Nuove ricerche sulla coltura in vitro di Aucuba japonica Thunb

Abstract

New researches on the culture « in vitro » of Aucuba japonica Thunb, – Caulinar fragments grown in vitro in nutrient media solidified with agar are compared with fragments grown on a filter paper support on unshaken liquid media. Tissue cultures of Aucuba are established putting down caulinar fragments just taken from the plant with isolated callus; subcultures are obtained more easily in July. We have observed that during the months of March or April, caulinar fragments, taken from the plant at various months of the year, of different ages and cultivated on various media, unexpectedly produce many roots.     

Culex pipiens affected by joint infection of a mosquito iridescent virus and Strelkovimermis spiculatus

Dual infections with a mosquito iridescent virus (MIV) and the mermithid nematode, Strelkovimermis spiculatus were recorded in natural Culex pipiens populations around La Plata city, Argentina. S. spiculatus was detected in 82% of samples that were positive for MIV infection. Dissected larvae of Cx. pipiens with patent MIV infection presented 42% infection with S. spiculatus. Larvae of Cx. pipiens exposed to MIV and S. spiculatus under laboratory conditions produced a high joint infection rate (82.5%) while no infection was recorded on larvae exposed to virus suspension only. Field and laboratory results suggest a strong association between S. spiculatus and MIV in natural populations of Cx. pipiens, in which S. spiculatus could be a mode of entry for the virus into the mosquito hemocele.     

Cdc15 is required for spore morphogenesis independently of Cdc14 in Saccharomyces cerevisiae

In Saccharomyces cerevisiae exit from mitosis requires the Cdc14 phosphatase to reverse CDK-mediated phosphorylation. Cdc14 is released from the nucleolus by the Cdc14 early anaphase release (FEAR) and mitotic exit network (MEN) pathways. In meiosis, the FEAR pathway is essential for exit from anaphase I. The MEN component Cdc15 is required for the formation of mature spores. To analyze the role of Cdc15 during sporulation, a conditional mutant in which CDC15 expression was controlled by the CLB2 promoter was used. Cdc15-depleted cells proceeded normally through the meiotic divisions but were unable to properly disassemble meiosis II spindles. The morphology of the prospore membrane was aberrant and failed to capture the nuclear lobes. Cdc15 was not required for Cdc14 release from the nucleoli, but it was essential to maintain Cdc14 released and for its nucleo-cytoplasmic transport. However, cells carrying a CDC14 allele with defects in nuclear export (Cdc14-DeltaNES) were able to disassemble the spindle and to complete spore formation, suggesting that the Cdc14 nuclear export defect was not the cause of the phenotypes observed in cdc15 mutants.     

Enhanced expression of the Epithelial Sodium Channel in neutrophils from hypertensive patients

Hypertension (HTN), i.e. abnormally high blood pressure, is a major risk factor for heart attack, stroke, and kidney failure. The Epithelial Sodium Channel (ENaC), one of the main transporters regulates blood pressure by tightly controlling the sodium reabsorption along the nephron. Recently, we have shown an α-ENaC overexpression in platelets from hypertensive patients compared to platelets from normotensive subjects, suggesting it makes a contribution to the activation state of platelets and the physiopathology of hypertension. However, the involvement of the α-ENaC localized in neutrophils to this disease remains unknown. Neutrophils are the first leukocytes to be recruited to an inflammatory site and are equipped with a strong ability to eliminate intra- or extracellular pathogens using reactive oxygen species or antibacterial proteins contained in their granules.Using the Western blotting (Wb), flow cytometry, and qRT-PCR approaches; we determined α-ENaC neutrophil overexpression at the protein and messenger RNA (mRNA) levels. By confocal and cytometry analysis, we determined the α-ENaC distribution and the heterogeneity of HTN neutrophils population, respectively. Immunoprecipitation and Wb assays demonstrated the presence of both α-ENaC and caveolin-1 phosphorylated forms, compared with neutrophils from healthy individuals. Although neutrophils from hypertensive subjects circulating in an activated state were exhibiting important oxidative stress and modifications registered by confocal, atomic force, and scanning electron microscope, they conserved their defense capabilities. The features described above for neutrophils from hypertensive patients could be attributed to α-ENaC overexpression, as its drug inhibition diminished their activation state modulating the actin cytoskeleton reorganization triggered during the activation process.     

Spatial variation in molluscan assemblages from coralline turfs of Argentinean Patagonia

Patterns of spatial variation of molluscan communities associatedwith coralline algal turfs were evaluated over 1,000 kmof the coast of Argentinean Patagonia. A hierarchically-nestedexperimental design was used to determine the relative importanceof molluscan assemblage variation at three different spatialscales (shores, sites and cores). Hypotheses were also testedabout the potential role of habitat variables (frond density,frond length, sediment and epiphytes) for determining molluscancommunity structure. In total, 38 molluscan species were foundcomprising 16, 18 and 4 species of bivalves, gastropods andpolyplacophorans, respectively. Densities of molluscs in corallineturfs reached ca 77,000 individuals per m² and were dominatedby mussels, especially Perumytilus purpuratus. Multivariateand univariate analyses of assemblage structure consistentlyshowed that variation at scales of metres and hundreds of kilometresdominated, with sites 20–50 m apart always contributingless than 24% of the total. Significant associations betweenmolluscan community structure and both frond density and frondlength demonstrated the potential importance of habitat structurein determining community structure at local scales. Variationin molluscan assemblages at the scale of shores, however, didnot appear to correlate with latitudinal, temperature or waveexposure gradients, indicating that other processes must beoperating. The compositions of molluscan assemblages in corallineturfs on the coast of Argentina were similar to those reportedfor central Chile. Comparisons of the richness of these SouthAmerican assemblages to other parts of the world revealed somestriking biogeographical patterns that warrant further investigation.Overall, this work highlights the general importance of small-scalevariation in molluscan assemblages on rocky shores and the consistentinfluence of habitat complexity in determining the structureof diverse molluscan communities associated with mat-like habitats. (Received 10 August 2006; accepted 20 January 2007)     

Exercise-induced bronchoconstriction alters airway nitric oxide exchange in a pattern distinct from spirometry

Exhaled nitric oxide (NO) is altered in asthmatic subjects with exercise-induced bronchoconstriction (EIB). However, the physiological interpretation of exhaled NO is limited because of its dependence on exhalation flow and the inability to distinguish completely proximal (large airway) from peripheral (small airway and alveolar) contributions. We estimated flow-independent NO exchange parameters that partition exhaled NO into proximal and peripheral contributions at baseline, postexercise challenge, and postbronchodilator administration in steroid-naive mild-intermittent asthmatic subjects with EIB (24-43 yr old, n = 9) and healthy controls (20-31 yr old, n = 9). The mean +/- SD maximum airway wall flux and airway diffusing capacity were elevated and forced expiratory flow, midexpiratory phase (FEF(25-75)), forced expiratory volume in 1 s (FEV(1)), and FEV(1)/forced vital capacity (FVC) were reduced at baseline in subjects with EIB compared with healthy controls, whereas the steady-state alveolar concentration of NO and FVC were not different. Compared with the response of healthy controls, exercise challenge significantly reduced FEV(1) (-23 +/- 15%), FEF(25-75) (-37 +/- 18%), FVC (-12 +/- 12%), FEV(1)/FVC (-13 +/- 8%), and maximum airway wall flux (-35 +/- 11%) relative to baseline in subjects with EIB, whereas bronchodilator administration only increased FEV(1) (+20 +/- 21%), FEF(25-75) (+56 +/- 41%), and FEV(1)/FVC (+13 +/- 9%). We conclude that mild-intermittent steroid-naive asthmatic subjects with EIB have altered airway NO exchange dynamics at baseline and after exercise challenge but that these changes occur by distinct mechanisms and are not correlated with alterations in spirometry.     

Quantitative evaluation of macrophage aggregates in brook trout Salvelinus fontinalis and rainbow trout Oncorhynchus mykiss

Macrophage aggregates (MAs) occur in various organs of fishes, especially the kidney, liver and spleen, and contain melanin, ceroid/lipofuscin and hemosiderin pigments. They have been used as indicators of a number of natural and anthropogenic stressors. Macrophage aggregates occur in salmonids but are poorly organized, irregularly shaped, and are generally smaller than those in derived teleosts. These features complicate quantification, and thus these fishes have seldom been used in studies correlating MAs with environmental stressors. To alleviate these complications, we developed color filtering algorithms for use with the software package ImagePro Plus (Media Cybernetics) that select and quantify pigmented area (i.e. colors ranging from gold to brown to black) in tissue sections. Image analysis results compared well with subjective scoring when tested on brook trout Salvelinus fontinalis and rainbow trout Oncorhynchus mykiss captured from high-elevation lakes or hatcheries. Macrophage aggregate pigments correlated positively with age and negatively with condition factor. Within individual fish, pigmentation correlated positively among organs, suggesting that the kidney, liver or spleen are suitable indicator organs. In age-matched fishes, MA pigments were not different between hatcheries and lakes in the organs examined. Between lakes, differences in pigments were observed in the kidney and spleen, but were not explained by age, condition factor, sex or maturation state. Our results indicate that quantification of the area occupied by MA pigments is an efficient and accurate means of evaluating MAs in salmonid organs and that organ pigmentation correlates with age and condition factor, as seen in studies with more derived fishes.     

Feasibility assessment for the classical biological control of <Emphasis Type="Italic">Tamarix</Emphasis> in Argentina

Saltcedars are woody plants in the genus Tamarix L. (Caryophyllales: Tamaricaceae) and are native to Eurasia and Africa. Several species have become invasive in the Americas, Australia and South Africa. In Argentina there are four species of Tamarix distributed in arid, semi-arid and coastal areas of most provinces. The taxonomic isolation of Tamarix spp. in Argentina, their widespread distribution, negative impact to natural areas and lack of impact from existing natural enemies all indicate that Tamarix is an ideal candidate for classical biological control in Argentina. Biological control of Tamarix spp. has been rapid and highly successful in the USA after the introduction of four Diorhabda spp. (Coleoptera: Chrysomelidae). Biological control of Tamarix spp. in Argentina could be implemented easily, rapidly, and at a low cost by utilizing the information developed in the USA.