A novel self-assembled epitope peptide nanoemulsion vaccine targeting nasal mucosal epithelial cell for reinvigorating CD8+ T cell immune activity and inhibiting tumor progression.

Epitope peptides are not suitable for nasal administration immunity due to their poor immunogenicity and low delivery efficiency. Here, we reported an intranasal self-assembled nanovaccine (I-OVA NE), which was loaded with the peptides IKVAV-OVA257-264 (I-OVA), a laminin peptide (Ile-Lys-Val-ala-Val, IKVAV) and OVA257-264 epitope conjugated peptide. This nanovaccine with I-OVA at a concentration of 4 mg/mL showed the average particle size of 30.37 ± 2.49 nm, zeta potential of -16.67 ± 1.76 mV, and encapsulation rate of 84.07 ± 7.59%. Moreover, the mucin did not alter its stability (size, PdI and zeta potential). And it also had no obvious acute pathological changes neither in the nasal mucosa nor lung tissues after nasal administration. Meanwhile, the antigen uptake of I-OVA NE was promoted, and the nasal residence time was also prolonged in vivo. Besides, the uptake rate of this nanovaccine was obviously higher than that of free I-OVA (P < 0.001) after blocking by the integrin antibody, suggesting that the binding of IKVAV to integrin is involved in the epitope peptide uptake. Importantly, this nanovaccine enhanced peptide-specific CD8+T cells exhibiting OVA257-264-specific CTL activity and Th1 immune response, leading to the induction of the protective immunity in E.G7-OVA tumor-bearing mice. Overall, these data indicate that I-OVA NE can be an applicable strategy of tumor vaccine development.

Re-examining the New Product Paradox: How Innovation Ambidexterity Mediates the Market Orientation and New Product Development Performance Relationship.

More and more well-documented failure of established companies which could not respond to rapid market changes, such as Kodak and Nokia, demonstrate the importance of transferring marketing information into real firm performance. While marketing strategy and management literature has long advocated the direct impact of strong firm market orientation (MO) on new product development (NPD) performance, limited research has discussed the mediating mechanism of this MO-NPD performance relationship. Using the traditional source-position-performance (SPP) framework, this study focuses on the innovation ambidexterity perspective to investigate the mediating mechanism between MO and NPD performance. Then, this study proposed a conceptual framework and propositions to examine the MO - NPD performance relationship further. Theoretical and practical implications of the findings are also discussed.

Development of Different Methods for Preparing Acinetobacter baumannii Outer Membrane Vesicles Vaccine: Impact of Preparation Method on Protective Efficacy.

Acinetobacter baumannii (A. baumannii) is becoming a common global concern due to the emergence of multi-drug or pan-drug resistant strains. Confronting the issue of antimicrobial resistance by developing vaccines against the resistant pathogen is becoming a common strategy. In this study, different methods for preparing A. baumannii outer membrane vesicles (AbOMVs) vaccines were developed. sOMV (spontaneously released AbOMV) was extracted from the culture supernatant, while SuOMV (sucrose-extracted AbOMV) and nOMV (native AbOMV) were prepared from the bacterial cells. Three AbOMVs exhibited significant differences in yield, particle size, protein composition, and LPS/DNA content. To compare the protective efficacy of the three AbOMVs, groups of mice were immunized either intramuscularly or intranasally with each AbOMV. Vaccination via both routes conferred significant protection against lethal and sub-lethal A. baumannii challenge. Moreover, intranasal vaccination provided more robust protection, which may be attributed to the induction of significant sIgA response in mucosal sites. Among the three AbOMVs, SuOMV elicited the highest level of protective immunity against A. baumannii infection, whether intramuscular or intranasal immunization, which was characterized by the expression of the most profound specific serum IgG or mucosal sIgA. Taken together, the preparation method had a significant effect on the yield, morphology, and composition of AbOMVs, that further influenced the protective effect against A. baumannii infection.

Chemical Profiles of Twenty-three Monovarietal Olive Oils Produced in Liangshan Region of China.

The main objective of this study was to differentiate twenty-three monovarietal olive oils produced in China (Azappa, Thiaki, Leucocarpa ovoid, manzanillo de labata, etc.) by studying their physicochemical properties, fatty acid profiles and oxidative stability. The majority of analytical indicators had statistically significant differences (p < 0.05). The tested cultivars produced excellent olive oils with an appreciable amount of natural antioxidants and chemical composition within the legal range. Our results demonstrate that Leucocarpa ovoid oil was characterized by high mean levels of total phenols (401.8 mg/kg), oleic acid (79.62%) and oxidative stability (18.64 h). Additionally, five types of olive oil were selected due to their high overall quality. Investigation of virgin olive oils indicates that the differences in the fatty acid profiles, phenols, sterols and tocopherols may be caused by genetic factors. The principal components analysis (PCA) results showed a strong differentiation between various cultivars in regard to their fatty acid composition and phenol, sterol and tocopherol levels. These components can be effective tools for efficient comparison and differentiation of these cultivars. To the best of our knowledge, this is the first work on the differentiation of the chemical constituents of virgin olive oils.

Atomic structure of a toxic, oligomeric segment of SOD1 linked to amyotrophic lateral sclerosis (ALS).

Fibrils and oligomers are the aggregated protein agents of neuronal dysfunction in ALS diseases. Whereas we now know much about fibril architecture, atomic structures of disease-related oligomers have eluded determination. Here, we determine the corkscrew-like structure of a cytotoxic segment of superoxide dismutase 1 (SOD1) in its oligomeric state. Mutations that prevent formation of this structure eliminate cytotoxicity of the segment in isolation as well as cytotoxicity of the ALS-linked mutants of SOD1 in primary motor neurons and in a Danio rerio (zebrafish) model of ALS. Cytotoxicity assays suggest that toxicity is a property of soluble oligomers, and not large insoluble aggregates. Our work adds to evidence that the toxic oligomeric entities in protein aggregation diseases contain antiparallel, out-of-register ß-sheet structures and identifies a target for structure-based therapeutics in ALS.

Structure-based design of non-natural amino-acid inhibitors of amyloid fibril formation.

Many globular and natively disordered proteins can convert into amyloid fibrils. These fibrils are associated with numerous pathologies as well as with normal cellular functions, and frequently form during protein denaturation. Inhibitors of pathological amyloid fibril formation could be useful in the development of therapeutics, provided that the inhibitors were specific enough to avoid interfering with normal processes. Here we show that computer-aided, structure-based design can yield highly specific peptide inhibitors of amyloid formation. Using known atomic structures of segments of amyloid fibrils as templates, we have designed and characterized an all-D-amino-acid inhibitor of the fibril formation of the tau protein associated with Alzheimer's disease, and a non-natural L-amino-acid inhibitor of an amyloid fibril that enhances sexual transmission of human immunodeficiency virus. Our results indicate that peptides from structure-based designs can disrupt the fibril formation of full-length proteins, including those, such as tau protein, that lack fully ordered native structures. Because the inhibiting peptides have been designed on structures of dual-ß-sheet 'steric zippers', the successful inhibition of amyloid fibril formation strengthens the hypothesis that amyloid spines contain steric zippers.

Genetic selection system for improving recombinant membrane protein expression in E. coli.

A major barrier to the physical characterization and structure determination of membrane proteins is low yield in recombinant expression. To address this problem, we have designed a selection strategy to isolate mutant strains of Escherichia coli that improve the expression of a targeted membrane protein. In this method, the coding sequence of the membrane protein of interest is fused to a C-terminal selectable marker, so that the production of the selectable marker and survival on selective media is linked to expression of the targeted membrane protein. Thus, mutant strains with improved expression properties can be directly selected. We also introduce a rapid method for curing isolated strains of the plasmids used during the selection process, in which the plasmids are removed by in vivo digestion with the homing endonuclease I-CreI. We tested this selection system on a rhomboid family protein from Mycobacterium tuberculosis (Rv1337) and were able to isolate mutants, which we call EXP strains, with up to 75-fold increased expression. The EXP strains also improve the expression of other membrane proteins that were not the target of selection, in one case roughly 90-fold.

Mixed polymeric micelles for combination cancer chemotherapy through the concurrent delivery of multiple chemotherapeutic agents.

As a novel drug nanocarrier for combination delivery of multiple anticancer agents, mixed polymeric micelles were developed and characterized in this study. A DNA-damaging anthracycline agent doxorubicin (DOX) and a phosphatidylinositol-3 kinase inhibitor wortmannin (WOR) were conjugated alone or combination onto poly(ethylene glycol)-poly(aspartate hydrazide) block copolymers through a hydrazone bond. Polymer-drug conjugates assembled into a unimodal micelle structure with a <100 nm particle size, in which the drug mixing ratios between DOX/WOR were precisely controlled. Cytotoxicity assay against a human breast cancer MCF-7 cell line showed that drug nanocarrier based on mixed polymeric micelles can reduce DOX required for cytotoxicity while maintaining the biological activity of the independent polymeric micelles. It is postulated that WOR enhances the efficacy of DOX through an efficient combination delivery accompanied by synergistic drug action. These findings, therefore, bring an effective drug delivery methodology that might reduce the effective dose as well as toxicity in vivo compared to the conventional drug formulations.

Poly(aspartate-g-PEI800), a polyethylenimine analogue of low toxicity and high transfection efficiency for gene delivery.

High-molecular-weight polyethylenimine (25 kDa, PEI25k) is one of the most common cationic polymers utilized in non-viral gene therapy. However, its methylene backbone (-CH(2)CH(2)N(x)-) and high charge density can result in poor biodegradability and high toxicity to cells. We hypothesize that optimizing the polymer length and charge density of PEI analogues may result in decreased toxicity and higher transfection efficiency, and improved biocompatibility in vivo. A series of PEI analogues with controlled molecular weight and charge density were synthesized by grafting low-molecular-weight PEI800 (800 Da) to a polyaspartate peptide backbone of varying degrees of polymerization. The optimum polymer had a degree of polymerization of 65 with an average of 16 PEI800 groups conjugated to it. All of the polycations investigated in the study caused inflammation and apoptosis/necrosis in the liver and spleen of rodents 24h post-injection; however, by day 5, the optimized poly(aspartate-g-PEI800) polymer and PEI800 did not show tissue damage or apoptosis, whereas PEI25k exhibited evidence of apoptosis/necrosis in the kidneys and spleen. Our study points to the need to optimize gene carriers to minimize toxicity, especially important for the safe delivery of therapeutic genes to explicit organs.

Lipophilic prodrugs of Hsp90 inhibitor geldanamycin for nanoencapsulation in poly(ethylene glycol)-b-poly(epsilon-caprolactone) micelles.

A solvent and Cremephor free formulation of the anticancer chemotherapeutic geldanamycin was prepared using amphiphilic block co-polymer micelles of poly(ethylene glycol)-b-poly(epsilon-caprolactone) (PEG-b-PCL). Although geldanamycin was not solubilized by PEG-b-PCL micelles, fatty acid prodrugs of geldanamycin were encapsulated in PEG-b-PCL micelles by a co-solvent extraction technique. Resulting PEG-b-PCL micelles were <120 nm in diameter and solubilized >20% w/w geldanamycin prodrugs increasing aqueous solubility to >2 mg/mL. PEG-b-PCL micelles released the geldanamycin prodrugs over several days, t(1/2) 2.2 to 9.6 days. The free prodrugs hydrolyzed rapidly, t(1/2)<6 h, into the geldanamycin analogue 17-beta-hydroxyethylamino-17-demethoxygeldanamycin, which has high activity against MCF-7 breast cancer cells, IC(50) 240 nM.