Characterization of Large Unilamellar Vesicles as Models for Studies of Lipid Peroxidation Initiated by Azocompounds

The aim of this work was to characterize large unilamellar vesicles (LUVETs) prepared by a hand-driven extrusion device in order to use them for studies of lipid peroxidation and antioxidant activity. Vesicle structure and size were examined by electron microscopy. Lipid and antioxidant content was determined before and after the extrusion procedure. Then LUVETs were subjected to autoxidation initiated by both the lipid-soluble 2,2'-azobis(2,4-dimethylvaleronitrile) and the water-soluble 2,2'-azobis(2-amidinopropane hydrochloride) azocompounds. The results demonstrated that: i) LUVETs prepared with lipid concentrations ranging between 25 and 150 mM were essentially unilamellar and reasonably homogeneous, with an average diameter of 90 nm; ii) the phospholipid, cholesterol and antioxidant amounts retained by filters were about 10-15%; iii) LUVETs were suitable for autoxidation studies initiated by the water-soluble azocompound both in the absence and presence of antioxidants. The lipid-soluble azocompound could be used only at low concentrations and its vesicle content had to be determined since part of the initiator was not incorporated into the lipid bilayer. These data suggest that LUVETs seem to be recommended for studies of lipid peroxidation and antioxidant activity.     

Pore formation by the sea anemone cytolysin equinatoxin II in red blood cells and model lipid membranes

Summary The interaction ofActinia equina equinatoxin II (EqT-II) with human red blood cells (HRBC) and with model lipid membranes was studied. It was found that HRBC hemolysis by EqT-II is the result of a colloid-osmotic shock caused by the opening of toxin-induced ionic pores. In fact, hemolysis can be prevented by osmotic protectants of adequate size. The functional radius of the lesion was estimated to be about 1.1 nm. EqT-II increased also the permeability of calcein-loaded lipid vesicles comprised of different phospholipids. The rate of permeabilization rised when sphingomyelin was introduced into the vesicles, but it was also a function of the pH of the medium, optimum activity being between pH 8 and 9; at pH 10 the toxin became markedly less potent. From the dose-dependence of the permeabilization it was inferred that EqT-II increases membrane permeability by forming oligomeric channels comprising several copies of the cytolysin monomer. The existence of such oligomers was directly demonstrated by chemical cross-linking. Addition of EqT-II to one side of a planar lipid membrane (PLM) increases the conductivity of the film in discrete steps of defined amplitude indicating the formation of cation-selective channels. The conductance of the channel is consistent with the estimated size of the lesion formed in HRBC. High pH and sphingomyelin promoted the interaction even in this system. Chemical modification of lysine residues or carboxyl groups of this protein changed the conductance, the ion selectivity and the current-voltage characteristic of the pore, suggesting that both these groups were present in its lumen.     

Biometric Study of Acetoin Production in Hanseniaspora guilliermondii and Kloeckera apiculata

Gas chromatographic analysis by direct injection of samples yielded quantitative data on acetoin content. Ninety-six strains of Hanseniaspora guilliermondii and Kloeckera apiculata were investigated for the ability to produce acetoin in synthetic medium and in must. High-level production of acetoin was found to be a characteristic of both species. In synthetic medium, the two species were not significantly different with respect to sugar utilization and ethanol or acetoin production. In grape must, the two species were significantly different (P = 0.001) in acetoin production and K. apiculata exhibited a significantly negative correlation between acetoin production and either sugar consumption or ethanol production. Use of selected apiculate yeasts in mixed cultures with Saccharomyces cerevisiae seems promising for optimization of wine bouquet.     

Climate change disrupts core habitats of marine species

Driven by climate change, marine biodiversity is undergoing a phase of rapid change that has proven to be even faster than changes observed in terrestrial ecosystems. Understanding how these changes in species composition will affect future marine life is crucial for conservation management, especially due to increasing demands for marine natural resources. Here, we analyse predictions of a multiparameter habitat suitability model covering the global projected ranges of >33,500 marine species from climate model projections under three CO₂ emission scenarios (RCP2.6, RCP4.5, RCP8.5) up to the year 2100. Our results show that the core habitat area will decline for many species, resulting in a net loss of 50% of the core habitat area for almost half of all marine species in 2100 under the high-emission scenario RCP8.5. As an additional consequence of the continuing distributional reorganization of marine life, gaps around the equator will appear for 8% (RCP2.6), 24% (RCP4.5), and 88% (RCP8.5) of marine species with cross-equatorial ranges. For many more species, continuous distributional ranges will be disrupted, thus reducing effective population size. In addition, high invasion rates in higher latitudes and polar regions will lead to substantial changes in the ecosystem and food web structure, particularly regarding the introduction of new predators. Overall, our study highlights that the degree of spatial and structural reorganization of marine life with ensued consequences for ecosystem functionality and conservation efforts will critically depend on the realized greenhouse gas emission pathway.     

Extra-pair paternity and sexual dimorphism in birds

Differences in the strength of sexual selection between males and females can lead to sexual dimorphism. Extra-pair paternity (EPP) can increase the variance in male reproductive success and hence the opportunity for sexual selection. Previous research on birds suggests that EPP drives the evolution of dimorphism in plumage colour and in body size. Because EPP increases the intensity of sexual selection in males, it should lead to increased dimorphism in species with larger or more colourful males, but decreased dimorphism in species with larger or more colourful females. We explored the covariation between EPP and sexual dimorphism in wing length and plumage colouration in 401 bird species, while controlling for other, potentially confounding variables. Wing length dimorphism was associated positively with the frequency of EPP, but also with social polygamy, sex bias in parental behaviour and body size and negatively with migration distance. The frequency of EPP was the only predictor of plumage colour dimorphism. In support of our prediction, high EPP levels were associated with sexual dichromatism, positively in species in which males are more colourful and negatively in those in which females are more colourful. Contrary to our prediction, high EPP rates were associated with increased wing length dimorphism in species with both male- and female-biased dimorphism. The results support a role for EPP in the evolution of both size and plumage colour dimorphism. The two forms of dimorphism were weakly correlated and predicted by different reproductive, social and life-history traits, suggesting an independent evolution.     

Partial characterization ofAspergillus fumigatus polygalacturonases for the degumming of natural fibers

Summary Aspergillus fumigatus strain 4, cultured on citrus pectin as the sole carbon source, produced polygalacturonases whose activity was optimum at 65°C and pH 3.5–4.5. The enzymes presented a bimodal thermostability for 10 min, but not 60 min, of incubation. Polygalacturonases showed pH stability between 3.0 to 9.0. The enzymes were stable when stored at 4–6°C for 90 days, but their activity was reduced by 24% when they were stored at 26–30°C. Orange pulp was the best pectic carbon source tested for the production of pectinases capable of retting ramie fibers. The reutilization of these enzymes was possible, suggesting the viability of industrial use of pectinases for degumming ramie fibers.     

PYK2 is overexpressed in chronic lymphocytic leukaemia: A potential new therapeutic target

Chronic Lymphocytic Leukaemia (CLL) is the most common adult B-cell leukaemia and despite improvement in patients' outcome, following the use of targeted therapies, it remains incurable. CLL supportive microenvironment plays a key role in both CLL progression and drug resistance through signals that can be sensed by the main components of the focal adhesion complex, such as FAK and PYK2 kinases. Dysregulations of both kinases have been observed in several metastatic cancers, but their role in haematological malignancies is still poorly defined. We characterized FAK and PYK2 expression and observed that PYK2 expression is higher in leukaemic B cells and its overexpression significantly correlates with their malignant transformation. When targeting both FAK and PYK2 with the specific inhibitor defactinib, we observed a dose–response effect on CLL cells viability and survival. In vivo treatment of a CLL mouse model showed a decrease of the leukaemic clone in all the lymphoid organs along with a significant reduction of macrophages and of the spleen weight and size. Our results first define a possible prognostic value for PYK2 in CLL, and show that both FAK and PYK2 might become putative targets for both CLL and its microenvironment (e.g. macrophages), thus paving the way to an innovative therapeutic strategy.     

Design and preclinical testing of an anti-CD41 CAR T cell for the treatment of acute megakaryoblastic leukaemia

Acute megakaryoblastic leukaemia (AMkL) is a rare subtype of acute myeloid leukaemia (AML) representing 5% of all reported cases, and frequently diagnosed in children with Down syndrome. Patients diagnosed with AMkL have low overall survival and have poor outcome to treatment, thus novel therapies such as CAR T cell therapy could represent an alternative in treating AMkL. We investigated the effect of a new CAR T cell which targets CD41, a specific surface antigen for M7-AMkL, against an in vitro model for AMkL, DAMI Luc2 cell line. The performed flow cytometry evaluation highlighted a percentage of 93.8% CAR T cells eGFP-positive and a limited acute effect on lowering the target cell population. However, the interaction between effector and target (E:T) cells, at a low ratio, lowered the cell membrane integrity, and reduced the M7-AMkL cell population after 24 h of co-culture, while the cytotoxic effect was not significant in groups with higher E:T ratio. Our findings suggest that the anti-CD41 CAR T cells are efficient for a limited time spawn and the cytotoxic effect is visible in all experimental groups with low E:T ratio.