A novel chimeric influenza virosome containing <Emphasis Type="Italic">Vesicular stomatitis</Emphasis> G protein as a more efficient gene delivery system

Objectives

To enhance the efficiency of influenza virosome-mediated gene delivery by engineering this virosome.

Results

A novel chimeric influenza virosome was constructed containing the glycoprotein of Vesicular stomatitis virus (VSV-G), along with its own hemagglutinin protein. To optimize the transfection efficiency of both chimeric and influenza cationic virosomes, HEK cells were transfected with plasmid DNA and virosomes and the transfection efficiency was assessed by FACS analysis. The chimeric virosome was significantly more efficient in mediating transfection for all amounts of DNA and virosomes compared to the influenza virosome.

Conclusions

Chimeric influenza virosome, including VSV-G, is superior to the conventional influenza virosome for gene delivery.

     

Angiogenic activity of endothelial progenitor cells through angiopoietin-1 and angiopoietin-2

Angiogenesis is a regulated process involving the proliferation, migration, and remodeling of different cell types particularly mature endothelial cells and recently discovered progenitor cells, named as endothelial progenitor cells (EPCs). Up to now, many attempts have been made to understand the dynamic balance of pro- and anti-angiogenic factors on EPCs on different milieu. It has been accepted that Ang-1, -2 and Tie-1, -2 signaling play a key role on angiogenesis pathways in endothelial lineage cells. In the current experiment, the angiogenic/angio-modulatory potency of Ang-1 and -2 was investigated on isolated EPCs. Freshly isolated EPCs were exposed to different concentrations of Ang-1 and -2 (25 and 50?ng/ml) over a course of 7 and 14 days. Corroborating to our results, a superior effect of Ang-1 on angiogenic properties, including an increased concentration of vascular endothelial growth factor, in vitro tubulogenesis, EPC migratory, Tie-2 expression and clonogenicity, was determined. A large amount of positive mature endothelium markers was achieved in EPCs being-exposed to Ang-1 peptide. Nonetheless, the number of CD133 positive cells increased in the presence of Ang-2. Collectively, we conclude that Ang-1 potentially induces functional and mature vascular-like behavior in EPCs more than Ang-2.     

Taxonomy and phylogenetic position of <Emphasis Type="Italic">Phyllactinia takamatsui</Emphasis>, a newly described powdery mildew on cotoneaster,based on molecular and morphological data

The newly recognised powdery mildew species Phyllactinia takamatsui on Cotoneaster nummularius (Rosaceae) is described and illustrated. This species, collected in Kerman Province, Iran, is well characterised by its conidial morphology and rDNA ITS sequences clearly different from allied species. Conidia are broadly ellipsoid to subcylindrical, i.e. they are not clavate-spathuliform as in most Phyllactinia species. The rDNA ITS sequence analysis showed that this species is closely allied to other species described on hosts belonging to Rosaceae, such as Ph. mali and Ph. pyri-serotinae. The ITS sequence of P. takamatsui was 92 to 94 % similar to that of the closest known relatives. The new species is described in detail, illustrated and compared with other similar taxa.     

Optimization of conformational stability and catalytic efficiency in chondroitinase ABC Ι by protein engineering methods

Chondroitinase ABC Ι can promote the recovery of spinal cord injuries by depolimerization of glycosaminoglycans. However, low thermal stability is one of the limitations regarding its clinical application. In order to increase the conformational stability of the enzyme, Leu⁶⁷⁹ at the starting point of a short helix located at the C‐terminal domain of the protein was replaced by serine (L679S mutant) and aspartic acid (L679D mutant). Theoretical and spectroscopic studies showed that the stability of enzyme increased upon mutation. Based on the activity measurements, the catalytic efficiency of L679S was improved in comparison with the wild‐type protein; while that of L679D (a more stabilized protein) was not changed. According to the structural and kinetic data, we proposed a model in which a higher conformational stability results in a slower rate of the formation of the open conformation. On the other hand, a higher flexibility slows down the rate of the formation and holding of the closed conformation. Therefore, the L679S mutant, which is structurally stable relative to the wild‐type protein and is destabilized compared to the L679D mutant, exhibited the best catalytic efficiency. However, it was also found that the L679D mutant was more suitable for long‐term storage of the enzyme.     

Phylogenetic diversity of lichen-associated homobasidiomycetes

The vast majority of lichenicolous fungi are relatively host-specific, nonvirulent ascomycetes and heterobasidiomycetes. A few known lichenicolous homobasidiomycetes (mushroom-forming fungi) generally exhibit broad host ecologies and in some cases, high virulence. Many produce conspicuous sclerotia or bulbils, thought to be adaptive in dispersal and survival. To more clearly understand the evolution of these atypical behaviors in lichenicolous basidiomycetes, we isolated or acquired specimens or cultures of 23 lichenicolous homobasidiomycetes and their relatives, from which we obtained mainly nuclear and some mitochondrial rDNA sequences. Phylogenetic analyses in this study indicate that a lichenicolous habit arose in four major clades. In two of these clades the habit represents a major evolutionary theme linked to the origin of well-known basidiolichens. The phylogenetic diversity of these fungi indicates that the lichenicolous habit arose recently and independently in the mushroom-forming fungi.     

Application of multivariate curve resolution-alternating least squares (MCR-ALS) for secondary structure resolving of proteins

Fourier transform infrared (FTIR) spectroscopy is the most common spectroscopic technique used for study of protein structure. Initially, band deconvolution techniques were applied to determine the secondary structure of proteins. Recently, several multivariate regression methods have been used to predict the secondary structure of proteins as an alternative to the previous methods. Multivariate curve resolution-alternating least squares (MCR-ALS) was applied on the FTIR spectra of proteins to resolve the fraction and spectral profiles of different structural motifs. Initial estimates of spectral profiles of different protein motifs were built using orthogonal projection approach (OPA). Predicted fractions of α-helix and β-sheet obtained by MCR-ALS technique were compared with those from partial least squares (PLS) modeling which revealed superiority of the former. If we consider the possibility of pure spectra prediction in addition to the prediction of secondary structure from the data set, MCR-ALS can be proposed as a very valuable alternative for qualitative and quantitative study of protein structures.     

Variation in Plumage Microbiota Depends on Season and Migration

Migratory birds can be efficient dispersers of pathogens, yet we know little about the effect of migration and season on the microbial community in avian plumage. This is the first study to describe and compare the microbial plumage community of adult and juvenile migratory birds during the annual cycle and compare the plumage community of migrants to that of resident birds at both neotropical and nearctic locations. We used length heterogeneity PCR (16S rRNA) to describe the microbial assemblage sampled from the plumage of 66 birds in two age classes and from 16 soil samples. Resident birds differed significantly in plumage microbial community composition from migrants (R ≥ 0.238, P < 0.01). Nearctic resident birds had higher plumage microbial diversity than nearctic migrants (R = 0.402, P < 0.01). Plumage microbial composition differed significantly between fall premigratory and either breeding (R ≥ 0.161, P < 0.05) or nonbreeding stages (R = 0.267, P < 0.01). Six bacterial operational taxonomic units contributed most to the dissimilarities found in this assay. Soil microbial community composition was significantly different from all samples of plumage microbial communities (R ≥ 0.700, P < 0.01). The plumage microbial community varies in relation to migration strategy and stage of the annual cycle. We suggest that plumage microbial acquisition begins in the first year at natal breeding locations and reaches equilibrium at the neotropical wintering sites. These data lead us to conclude that migration and season play an important role in the dynamics of the microbial community in avian plumage and may reflect patterns of pathogen dispersal by birds.     

Inositol pyrophosphates inhibit Akt signaling, thereby regulating insulin sensitivity and weight gain

The inositol pyrophosphate IP7 (5-diphosphoinositolpentakisphosphate), formed by a family of three inositol hexakisphosphate kinases (IP6Ks), modulates diverse cellular activities. We now report that IP7 is a physiologic inhibitor of Akt, a serine/threonine kinase that regulates glucose homeostasis and protein translation, respectively, via the GSK3β and mTOR pathways. Thus, Akt and mTOR signaling are dramatically augmented and GSK3β signaling reduced in skeletal muscle, white adipose tissue, and liver of mice with targeted deletion of IP6K1. IP7 affects this pathway by potently inhibiting the PDK1 phosphorylation of Akt, preventing its activation and thereby affecting insulin signaling. IP6K1 knockout mice manifest insulin sensitivity and are resistant to obesity elicited by high-fat diet or aging. Inhibition of IP6K1 may afford a therapeutic approach to obesity and diabetes.     

Electrospinning of chitosan nanofibers: Processing optimization

In this study, the electrospinning of chitosan has been investigated. The problem of chitosan high viscosity, which limits its spinability, is resolved through the application of an alkali treatment which hydrolyzes chitosan chains and so decreases its their molecular weight. Solutions of the treated chitosan in aqueous 70–90% acetic acid produce nanofibers with appropriate quality and processing stability. Decreasing the acetic acid concentration in the solvent increases the mean diameter of the nanofibers. Optimum nanofibers are achieved with chitosan which is hydrolyzed for 48 h. Such nanofibers result in a moisture regain which is 74% greater than that of treated and untreated chitosan powder. The diameter of this nanofiber, 140 nm, is strongly affected by the electrospinning conditions as well as by the concentration of the solvent. FTIR investigations prove that neither the alkali treatment nor the electrospinning process change the chemical nature of the polymer.